Download Review A model for chromosome structure during the mitotic

Chromosome Research 9: 175^198, 2001.
# 2001 Kluwer Academic Publishers. Printed in the Netherlands
175
Review
A model for chromosome structure during the mitotic and meiotic cell cycles
Stephen M. Stack & Lorinda K. Anderson
Department of Biology and Cell and Molecular Biology Program, Colorado State University, Fort Collins,
CO 80523, USA; Fax: 970-491-0649; Tel: 970-491-6802; E-mail: [email protected]
Received 12 October 2000; received in revised form and accepted for publication by H. Macgregor 2 January 2001
Key words: chiasmata, chromosome cores, chromosome structure, coiling, crossing over, meiosis, mitosis,
model, presynaptic alignment
Abstract
The chromosome scaffold model in which loops of chromatin are attached to a central, coiled chromosome
core (scaffold) is the current paradigm for chromosome structure. Here we present a modi¢ed version of the
chromosome scaffold model to describe chromosome structure and behavior through the mitotic and
meiotic cell cycles. We suggest that a salient feature of chromosome structure is established during
DNA replication when sister loops of DNA extend in opposite directions from replication sites on nuclear
matrix strands. This orientation is maintained into prophase when the nuclear matrix strand is converted
into two closely associated sister chromatid cores with sister DNA loops extending in opposite directions.
We propose that chromatid cores are contractile and show, using a physical model, that contraction
of cores during late prophase can result in coiled chromatids. Coiling accounts for the majority of chromosome shortening that is needed to separate sister chromatids within the con¢nes of a cell. In early prophase I
of meiosis, the orientation of sister DNA loops in opposite directions from axial elements assures that DNA
loops interact preferentially with homologous DNA loops rather than with sister DNA loops. In this context, we propose a bar code model for homologous presynaptic chromosome alignment that involves weak
paranemic interactions of homologous DNA loops. Opposite orientation of sister loops also suppresses
crossing over between sister chromatids in favor of crossing over between homologous non-sister
chromatids. After crossing over is completed in pachytene and the synaptonemal complex breaks down
in early diplotene (ˆ diffuse stage), new contractile cores are laid down along each chromatid. These
chromatid cores are comparable to the chromatid cores in mitotic prophase chromosomes. As an aside,
we propose that leptotene through early diplotene represent the `missing' G2 period of the premeiotic
interphase. The new chromosome cores, along with sister chromatid cohesion, stabilize chiasmata. Contraction of cores in late diplotene causes chromosomes to coil in a con¢guration that encourages subsequent
syntelic orientation of sister kinetochores and amphitelic orientation of homologous kinetochore pairs on
the spindle at metaphase I.
Introduction
In the 1950s, the general model for chromosome
structure included a coiled chromosome core
embedded in a matrix that is covered by a thin
pellicle (e.g. Schrader 1953, DeRobertis et al.
1954, see Ohnuki 1968 for a review). However,
electron microscopic observations in the 1960s
were interpreted to indicate that chromosomes
consist of a dense tangle of 10^30-nm chromatin
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¢bers with neither a core to the inside nor a pellicle
to the outside, i.e. the `folded-¢ber model' (e.g.
DuPraw 1968, and see Belmont & Bruce 1994
for a newer version of this model). The existence
of coiling in some chromosomes was acknowledged but the only recognized chromosome cores
were axial elements and lateral elements related
to the synaptonemal complex (SC; see Moses 1968
for a contemporary review).
Subsequently, new observations and experiments resulted in a new paradigm for chromosome structure (the chromosome scaffold
model) that in many ways resembles the preelectron microscope chromosome model from
the 1950s: Condensed chromosomes are coiled
(e.g. Ohnuki 1968), and each chromatid has a
scaffold (core) of non-histone proteins running
along its length (e.g. Paulson & Laemmli 1977,
Earnshaw & Laemmli 1983) that is also coiled
(Rattner and Lin 1985, Boy de la Tour & Laemmli
1988). The single most abundant protein
component that has been identi¢ed so far in the
scaffold is topoisomerase II, an enzyme that has
the potential to decatenate tangled DNA by passing one double helix through the other (Berrios
et al. 1985, Earnshaw & Heck 1985, Gasser et
al. 1986). Loops of DNA (associated with protein
to form chromatin ˆ matrix in the old terminology) extend laterally from the scaffold (e.g.
Gasser & Laemmli 1987, Earnshaw 1988). The
bases of DNA loops are bound to chromosome
scaffolds by SARs (scaffold attachment regions)
that are AT-rich segments of bent DNA to which
one or more scaffold proteins bind (e.g.
Homberger 1989, von Kries et al. 1991). SARs
are also attached to the nuclear matrix during
interphase when SARs are called MARs (matrix
attachment regions; Mirkovitch et al. 1988,
Boulikas 1995). Chromosome scaffolds and
nuclear matrix strands appear to be related structures that share some proteins, including topoisomerase II and p62 (e.g. Gasser et al. 1989,
Berezney et al. 1995). In this regard, many SARs
include the 15-bp consensus sequence for
topoisomerase II, indicating that SARs are sites
where DNA catenations are resolved (Gasser et
al. 1989, Gimënez-Abiän et al. 1995). Finally,
chromosomes are surrounded by an RNA-protein
S. M. Stack & L. K. Anderson
coat (ˆ pellicle in the old terminology) that largely
derives from the disassembly of the nucleolus and
application of some nucleolar components to condensed chromosomes during mitosis (e.g. Moyne
& Garrido 1976, see Hernandez-Verdun & Gautier
1994 for a review).
During interphase, the nuclear matrix is probably the counterpart of the chromosome scaffold
of mitotic chromosomes (see Gasser et al. 1989
for a review). Newly synthesized DNA and RNA,
as well as enzymes involved in these syntheses,
are attached to the nuclear matrix. This suggests
that loops of DNA are reeled through relatively
stationary replication and transcription complexes
on the nuclear matrix (see Cook 1999 for a review).
Replication origins occur roughly every 15^30 mm
of DNA, although the range of replicon size is
probably enormous (Gasser et al. 1989, Boulikas
1995, Berezney et al. 2000). Starting from a
replication origin, two replication forks proceed
in opposite directions until they encounter
replication forks advancing from adjacent
replicons (Huberman & Riggs 1968, Jackson &
Pombo 1998). Clusters of synchronous replicons
can be labeled and observed as £uorescent foci
by light microscopy or as electron-dense masses
of chromatin by electron microscopy (Nakayasu
& Berezney 1989, Hozäk et al. 1993, Newport
& Yan 1996). If the association of DNA with
the matrix were mediated by replication complexes
only, then one would expect a changing sample of
the DNA to be associated with the nuclear matrix
during S phase. However, Gasser et al. (1989)
reported that MARs are associated with the
matrix/scaffold throughout the cell cycle, so this
subset of DNA never leaves the nuclear
matrix/scaffold.
Assuming that these descriptions of chromosome structure and DNA replication are generally
accurate, here we present an extension of these
ideas to describe a model for some aspects of
the structure and behavior of chromosomes
through the mitotic and meiotic cell cycles. In
the descriptions that follow, ideas that we view
as more speculative are in italics. See Earnshaw
(1988), Manuelidis & Chen (1990), Cook (1995),
and Heslop-Harrison (2000) for reviews and other
models for chromosome structure.
Chromosome structure through mitosis and meiosis
Model for the structure of chromosomes through
the mitotic cell cycle
G1 phase
During the G1 phase, each chromosome consists of
only one chromatid that relaxes and uncoils
(Belmont & Bruce 1994). Each chromatid carries
a single DNA double helix that is supercoiled
around clusters of histones to form nucleosomes
(Kornberg 1974, Olins & Olins 1974). Each
nucleosome consists of two molecules each of
histone H2A, H2B, H3, and H4. Tandem
nucleosomes form a strand of chromatin with a
diameter of 10 nm. Through interaction with
H1 histones, these 10-nm ¢laments are often
supercoiled to form 30 nm chromatin ¢bers (Finch
& Klug 1976, Losa et al. 1984, but see Woodcock
& Horowitz 1991 who present evidence for a different organization of nucleosomes in transcriptionally active cells). The 30-nm chromatin
¢bers form loops attached at their bases (via
MARs, replication origins, and promoters) to
strands of nuclear matrix consisting of protein
and RNA (Nickerson et al. 1995, Boulikas 1995,
Jackson et al. 1996, Ma et al. 1999, see Cremer
et al. 1995 for different models of the nuclear
matrix). Loops of DNA (chromatin) extend from
only one side of a matrix strand in G1 (Figures 1,
12A). (The basis for assuming this arrangement
of loops is explained later.) These nuclear matrix
strands to which chromatin loops are attached were
derived from single, relaxed and extended
chromatid cores from the previous anaphase
(Figure 1, 12A), which show relic coiling from
the previous anaphase (e.g. Vejdovsk 1911^1912
for early illustrations, Manuelidis & Chen 1990,
Belmont & Bruce 1994). Interconversion of
nuclear matrix strands and chromosome cores is
suggested by premature chromosome condensation experiments in which G1 cells are fused with
metaphase cells (e.g. Rao & Johnson 1970). While
G1 nuclear matrix strands from unfused cells
do not stain with silver, each long prematurely
condensed G1 chromosome develops a single
silver-stainable core throughout its length
(Gimënez-Abiän et al. 1995). This core is comparable to the single silver-stained core in each
chromatid of metaphase and anaphase chromosomes (Gimënez-Abiän et al. 1995). Acquisition
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of silver stainability must be due to addition of
new proteins and/or modi¢cation of existing proteins that takes place during prophase. The
relationship between matrix strands and/or
chromosome cores is further supported by reports
that DNA loops are attached to matrix strands
and chromosome cores by the same AT-rich
sequences (MARs and SARs ^ Mirkovitch et al.
1984, Gasser et al. 1986) and by reports that
chromosome cores and nuclear matrix strands
share some of the same proteins (Detke & Keller
1982, Peters et al. 1982, Gasser et al. 1989), probably including RNA polymerases I and II and
lamins (Matsui et al. 1979, Jackson & Cook 1995).
S phase
During early S phase in mammalian cells, groups
of approximately 40 adjacent replicons initiate
replication together in electron-dense bodies
100^200 nm in diameter (replication factories)
attached to the nuclear matrix (e.g. Jackson &
Cook 1995). MARs are attached to the nuclear
matrix throughout interphase, and pulse labeling
during S-phase demonstrates that nascent DNA
is closely associated with the nuclear matrix as
well (e.g. Pardoll et al.1980, Jackson & Pombo
1998). Thus, at the beginning of S phase,
replication origins must be attached to the nuclear
matrix at the centers of replicons (Figure 2), and
MARs may reside at or near replication origins
(see Boulikas 1995 for a review). When replication
is initiated at the base of a DNA loop, the two
replication forks remain at the replication
complex, and parental DNA is pulled in from
the left and right (Figures 3^5) (Pardoll et al.
1980). Nucleosomes are disassembled at replication forks before their histones reassociate
(along with new histones) with replicated DNA
to form new nucleosomes in the daughter strands
of chromatin (e.g. Randall & Kelly 1992). Replication origins and MARs are duplicated ¢rst
and quickly reattach to the matrix strand, i.e.
replication origins and MARs do not emerge from
the matrix strand on daughter DNA loops (Cook
1991). However, as more parental DNA without
MARs is reeled in from adjacent parental loops,
two sister loops of DNA emerge from the left
and two sister loops of DNA emerge from the right
of the replication complex. The nascent sister
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loops emerge at 180 to each other. Eventually
replication forks from adjacent replicons meet.
At this point, the DNA is released from
replication complexes and matrix strands except
at MARs (Figure 6). This model is in agreement
with observations that sister DNA sequences
are separated from each other by the time
replication is completed (Selig et al. 1992). There
is some tangling (catenation) of sister DNA loops
but this is minimized by orientation of sister loops
in opposite directions with the result that most
of the tangling should be near the bases of the
loops. Also, MARs often include the 15-bp consensus sequence for topoisomerase II (Gasser et
al. 1986), and topoisomerase in the matrix strands
is in the right position to decatenate sister loops of
DNA when they come under tension as the sister
chromatids begin to shorten and separate in
prophase (Earnshaw & Heck 1985, Woessner et
al. 1987, Saitoh & Laemmli 1994, Gimënez-Abiän
et al. 1995). It is also during S phase when cohesin
complexes establish linkage between sister
chromatids (Uhlmann & Nasmyth 1988, Nasmyth
et al. 2000). Since sister chromatid cohesion can
be established only during S phase, cohesion complexes probably have to be inserted when the
bases of sister DNA loops are close together
during or right after replication.
A number of observations indicate that during S
phase there is a major reorganization of chromosome structure. At this time, prematurely condensed chromosomes take on a `pulverized'
appearance (e.g. Sperling & Rao 1974), silverstained cores of prematurely condensed chromosomes show discontinuities (Gimënez-Abiän et
al. 1995), and interphase chromosomes lose distinct borders (Stack et al. 1977, Brown et al.
1978). These observations are probably related
to local dissolution of the single G1 matrix strands
during DNA replication. Groups of adjacent
replicons initiate replication together in factories
(e.g. Hozäk et al. 1993, Jackson & Cook 1995),
indicating coordination of replication units related
to chromosome structure that is more or less maintained even during S phase (Manuelidis 1990). As
batteries of replicons complete replication, a new
matrix strand forms in association with the MARs
of one DNA double helix (chromatid), and another
matrix strand forms in association with the MARs
of the sister DNA double helix. Sister matrix
S. M. Stack & L. K. Anderson
strands are in close association. These segments
of sister matrix strands link up progressively as
batteries complete replication until a continuous
pair of sister matrix strands extends throughout
the length of a duplicated chromosome at the
end of S phase, i.e. the beginning of G2 (Figure 6
& 7; e.g. Gimënez-Abiän et al. 1995). How sister
matrix strands form in close association with each
other and tell one sister DNA strand from the
other remains to be explained, but it may involve
opposite orientation of sister DNA loops.
G2 phase
During G2, chromatin is diffuse, consisting of 10and 30-nm-diameter chromatin ¢bers. Sister
DNA (chromatin) loops extend in opposite directions from duplicated but closely associated sister
matrix strands (Figure 7). A G2 chromosome
occupies a cylinder of space in the nucleus. DNA
loops of each sister chromatid probably occupy a
half cylinder of space with sister nuclear matrix
cores in the center of the cylinder of chromatin
(Figure 7C). Loops of DNA on one chromatid
can extend from the matrix core outward anywhere
within the half cylinder occupied by one chromatid,
with the caveat that the sister loop in the sister
chromatid extends from its matrix core in the
opposite direction within the con¢nes of its own half
cylinder. Matrix strands are £exible, and DNA
loops are not condensed, so even though sister loops
extend in opposite directions, sister DNA loops are
able to come in contact and act as preferred
templates (compared to homologous chromosomes) for repair of DNA damage during S and
G2 (van Heemst & Heyting 2000). Consistent with
this model, premature chromosome condensation
of G2 nuclei results in the appearance of elongate
chromosomes that consist of two parallel sister
chromatids (e.g. Rao & Johnson 1970). Each
prematurely condensed chromatid has a silverstainable core, and sister cores are closely associated (Gimënez-Abiän et al. 1995, 2000).
See Cook (1991) for a very similar model for
interphase chromosome structure and replication,
except for the stipulation that sister DNA loops
extend in opposite directions from the duplicated
matrix strand.
Chromosome structure through mitosis and meiosis
179
Figures 1^11 (overleaf). Diagrammatic representations of chromosome structure through the mitotic cell cycle. Drawings are not to
scale. Telophase/G1 . (A) Longitudinal view of a segment of a decondensing chromosome during telophase or a decondensed chromosome during G1. The anaphase chromosome core (copper-colored) relaxes and uncoils as it is converted by modi¢cation of its proteins
into a matrix strand during telophase. DNA loops (green lines ^ mostly 30-nm diameter chromatin ¢bers) are attached to the
core/matrix strand via MARS (green dots) with closely associated replication origins (included in green dots). Probable attachments
to transcription complexes are not illustrated for simplicity (Jackson et al. 1996). The DNA loops are present on only one side
of the matrix strand. For diagrammatic purposes, the loops are shown the same length, in one plane, and short relative to the
core/matrix strand. Also see Figure 12A. (B) Cross section of a core/matrix strand with one loop extending upward. (C) End view
of a segment of a core/matrix strand with successive DNA loops extending in various directions within a half-circle (or within
a half-cylinder three-dimensionally). This view is not shown again until Figure 7C. Figure 2. Late G1 . (A) Longitudinal view of
a nuclear matrix strand that includes replication complexes (gray ovals) and MARs (near replication origins) at the bases of
DNA loops. (B) Cross section of a matrix strand showing only one loop. Figure 3. Early S-phase. (A) Longitudinal view of a matrix
strand during DNA synthesis. The matrix strand is partly disassembled (note that the copper-colored strand is missing) at sites actively
undergoing DNA replication. In this string of replication complexes, replication origins and MARS are the ¢rst sequences to be
replicated, but they remain associated with the replication complexes, as do replication forks. Adjacent replicons tend to initiate
and complete replication together in groups associated with dense structures that have been referred to as replication factories (e.g.
Hozäk et al. 1993). (B) Cross section of a replication complex showing two MARs/replication origins as well as an unreplicated
loop extending upward. Figure 4. Mid S-phase. (A) Longitudinal view of a string of replication complexes with newly formed sister
loops of DNA extending upward and downward (in opposite directions) from their bases in replication complexes. For diagrammatic
purposes, the unreplicated (parental) DNA is shown in the same plane as the replicated sister loops but the parental DNA may be at a
90 angle to both newly replicated sister loops. The loops of unreplicated DNA shorten as they are drawn into replication complexes
and fed into replication forks. (B) Cross section of a replication complex showing one long unreplicated loop (above) and shorter
replicated loops above and below. Figure 5. Late S-phase. (A) Longitudinal view of a string of replication complexes near the
end of DNA synthesis. Parental DNA shortens as replicated DNA loops continue to enlarge. (B) Cross section showing sister loops
extending above and below a replication complex. Figure 6. DNA synthesis complete. (A) Longitudinal view of a string of replication
complexes after the last segments of DNA are replicated between adjacent replication complexes (adjacent replication forks meet).
Completely replicated sister loops of DNA are released from the replication complexes except at MAR/replication origins. (B) Cross
section showing sister loops extending in opposite directions from each other and attached to replication complexes at
MAR/replication origins. Figure 7. G 2 . (A) Longitudinal view of assembling sister matrix strands (light copper) after disassembly
of replication complexes. Sister DNA loops extend in opposite directions from adjacent sister cores. Each newly formed matrix strand
is associated with MARs on one DNA double helix. Also see Figure 12B. (B) Cross section of forming sister matrix strands/cores with
two sister loops of DNA extending in opposite directions. (C) End view of a segment of matrix strand showing sister DNA loops (same
color) extending in opposite directions from the duplicated matrix strands/cores. Loops from one chromatid are found within one of
the half circles (e.g. upper gray half circle or half cylinder three-dimensionally), and these loops from one chromatid do not overlap
with DNA loops from the sister chromatid in the other (lower gray) half circle. Figure 8. Early prophase. (A) Longitudinal view
of early prophase chromosome. What were formerly nuclear matrix strands are being modi¢ed by changes in their proteins to become
more substantial chromatid cores (ˆ scaffolds, copper) that are capable of active shortening. Thirty-nm chromatin loops collapse
toward centrally located sister chromatid cores. The loops are connected to chromatid cores by SARs (ˆ interphase MARs). Sister
chromatids and sister cores are held together throughout their length by DNA catenations and cohesins. Also see Figure 12B.
(B) Cross section showing two condensed sister chromatin loops extending in opposite directions from adjacent sister chromatid
cores. Figure 9. Late prophase. Longitudinal view of contracting (actively shortening) chromatid cores. Contraction results in some
shortening of prophase chromosomes. However, as long as sister cores are trapped together in the center of the chromatin mass,
neither extensive shortening nor coiling of chromatids is possible. In this diagram, the SARS are somewhat closer together than
in Figure 8 due to contraction of the sister cores. Also see Figure 12C. Figure 10. Prometaphase/metaphase. Three-dimensional
drawing (at lower magni¢cation than the diagrams in Figures 1^9). Sister chromatid cohesion has relaxed along the length of chromosomes except at centromeres. Whatever the nature of sister chromatid cohesion, sister chromatids remain close to each other along
their length. Active shortening of chromatid cores causes chromatid arms to coil and shorten. Coiling does not occur at the centromere
because the two chromatid cores are still held close together at this location. Because centromeres are not coiled, they look like
constrictions of the chromosome, i.e. primary constrictions. Kinetochores (yellow) face in opposite directions and attach to
kinetochore microtubules extending from opposite poles. At the end of each chromatid, individual chromatin loops coming off
one side of each chromatid core are illustrated. To the right in a transparent view, the coils of both chromatids are loosened
and the paths of the chromatid cores are shown. To the left, note a reversal of coiling direction in the upper chromatid arm. However,
sister chromatids usually coil as mirror images of each other. Also see Figure 12D. Figure 11. Anaphase. Three- dimensional drawing.
Sister chromatid cohesion has been released at the centromere region, and this permits coiling to extend through centromeres. The two
sister chromosomes (chromatids) are moving toward opposite poles. A reversal of coiling is visible in the upper left chromosome (see
Figures 10 & 12E). Within each chromosome, the contracted chromosome core (copper) remains to one side of the chromatin.
To the right in a transparent view, the coils are loosened and the paths of the chromatid cores are shown.
180
S. M. Stack & L. K. Anderson
Chromosome structure through mitosis and meiosis
181
182
Figure 12. Rope and elastic models of mitotic chromosomes.
The chromatin of each chromatid is represented by a segment
of £exible white rope that was initially 53 cm long in all of
the models. The chromatid core is represented by a black elastic
band that was sewn to the white rope. Kinetochores are represented by felt patches labeled with `K.' In these models,
1 cm represents 1 mm, i.e. the ropes are 10 000 : 1 scale models
of chromosomes. (A) G1 chromosome. Here the elastic band
S. M. Stack & L. K. Anderson
was not stretched before it was sewn to the rope because at this
stage the chromosomes are decondensed and the cores are
not contracted. The model is 53 cm long. Also see Figures 1
& 2. (B) G2 early prophase chromosome. Two ropes like those
in A (above) were sewn together with the two (non-stretched)
elastic bands in the center. The bands between the ropes (arrow)
represent the chromatid cores between sister chromatids held
together by cohesins. The model is 53 cm long. Also see
Figure 7. (C) Prophase chromosome. Here, a 53 cm length
of white rope was sewn to a maximally stretched elastic band
53 cm long. If not attached to the rope, this stretched elastic
band will contract to about 22 cm (two-fold) upon release of
the tension. A second 53-cm rope and stretched elastic band
was prepared the same way. Then, while fully stretched to
53 cm, the two ropes were sewn to each other lengthwise with
the stretched elastic bands adjacent between them (arrow).
When tension on this composite of ropes and stretched elastic
is released (as shown), the length decreases from 53 cm to
40 cm (a 32% reduction in length) with no tendency to coil.
This suggests that only modest shortening of a chromosome
can be achieved by a contractile core located between two
closely associated chromatids. Also see Figures 8 & 9. (D)
Metaphase chromosome. Two ropes attached to bands of
stretched elastic were sewn together as in C above. Subsequently, the threads holding the ropes together distal to
the centromeres were cut to free the ropes (chromatid arms)
from each other. The response of the relatively incompressible
rope (representing chromatin) attached on one side to a contractile band (the chromosome core) is to coil with the contractile band taking the shorter, inside track. These coils of
rope are 15 cm long, representing a 72% reduction in length
compared with G2/early prophase chromosomes (see B above),
demonstrating that coiling achieves much more shortening than
would be anticipated by simple contraction of the chromatid
cores (see C above). Coiling occurs everywhere except at the
centromere (= primary constriction) where sister chromatids
are still bound to each other. The arrowhead points to a reversal
of coiling. To the left, the bottom chromatid has been pulled out
so that the inner black elastic band can be seen. Also see Figure
10. (E) Anaphase chromosomes. A metaphase chromosome
model was prepared as described in D above. To mimic the loss
of sister chromatid cohesion from centromeres at the beginning
of anaphase, the threads holding the two ropes together at the
centromere region were removed with the result that both ropes
coiled along their entire length. The two ropes were then
oriented to represent their relationship in mid-anaphase. These
coils are about 11 cm long, representing a reduction in length
of 79% compared with G2-early prophase chromosomes (see
B above). Thus, the majority of reduction in length of mitotic
chromosomes is due to coiling rather than contraction per
se. The arrowhead points to a reversal of coiling. Also see Figure 11.
Chromosome structure through mitosis and meiosis
Mitosis
As cells pass into prophase, any remaining 10-nm
chromatin ¢bers are supercoiled to form 30-nm
¢bers, and most transcription ceases, related to
deacetylation of histones and loss of transcription
factors from chromatin (Mart|¨ nez-Balbäs et al.
1995 and references therein). Thirty-nm ¢bers
are pulled toward their attachment sites on cores
by protein complexes called condensins (see
Hirano 1999 for a review). At the same time,
nuclear matrix strands are converted into more substantial chromatid cores by adding proteins and/or
modifying proteins of the matrix strand that now
can be silver-stained. At the initiation of chromosome condensation in prophase, sister chromatid
cores are close together as observed in prematurely
condensed G2 chromosomes and silver-stained
prometaphase chromosomes (Figures 7, 8 & 12B;
Gimënez-Abiän et al. 1995, 2000).
Chromosomes shorten during prophase (Figure
9). Shortening is initially due to active lengthwise
contraction of the paired sister cores between
as-yet unseparated chromatids that are held
together by DNA catenations and cohesin complexes. Shortening of cores is inhibited to a degree
by their location in the interior of the mass of sister
chromatin that resists being packed more tightly
(see the rope and elastic model shown in Figure
12C).
Sister chromatid cohesion is mediated by
cohesins (see Orr-Weaver 1999 for a review)
and catenations (tangling and interlocking) of
DNA loops. Catenation of sister DNA loops is
most likely at their bases in cores where
topoisomerase II is available to resolve interlocking. Resolution probably occurs when
catenations are brought under tension by chromatin condensation and contraction of cores
during prophase (Downes et al. 1991, Gorbsky
1994, Gimënez-Abiän et al. 1995). If topoisomerase II is inhibited and catenations cannot be
resolved, silver staining reveals intimately paired
sister cores embedded in extended chromosomes
that persist long after sister cores normally would
have separated in prometaphase (Gimënez-Abiän
et al. 1995, 2000). This is strong evidence that
sister cores are located together and to one side
of sister chromatids (Figures 7^9, 12B, C). Without decatenation, full chromosome condensation
183
and sister chromatid separation is not possible
(Uemura et al. 1987, Newport & Spann 1987,
Adachi et al. 1991, Downes et al. 1991, 1994,
Gimënez-Abiän et al. 1995, 2000).
At the transition from prophase to prometaphase, sister chromatid arms separate to some
degree as catenations are eliminated and cohesins
break down. This permits sister chromatid arms
and their cores to separate and coil (GimënezAbiän et al. 1995, 2000, see Manton 1950 for a
review of chromosome coiling; Figures 10, 12D).
Coiling is important because it is the primary
means by which chromosomes are compacted
and shortened enough to be separated from each
other within the con¢nes of the cell (Figure 12).
Sister chromatids usually show mirror image
patterns of coiling (Boy de la Tour & Laemmli
1988) that is probably related to their identical
chromatin structure. This relationship is
supported by the observations of Baumgartner
et al. (1991) that single copy sequences usually
occupy mirror image positions on sister
chromatids in human chromosomes. Because
the position of single copy sequences usually
remained the same on chromosomes that varied
somewhat in length, they concluded that ¢nal
chromosome condensation is due to tighter
packing of the coils rather than continued coiling.
Chromatin structure affects the number and placement of coils based on evidence that the coiling
pattern of individual mammalian chromosomes
is related to their chromomere and G-band
patterns (e.g. Kato & Yosida 1972, Okada &
Comings 1974, Harrison et al. 1981). However,
sister chromatids can differ from each other by
reversals in their coiling pattern (Figures 10, 11
& 12D, Manton 1950, Ohnuki 1968).
There have been a number of different (often
con£icting) proposals to explain coiling (see John
& Lewis 1965 for a review). In our model, the
initial location of a chromosome core along one
side of a chromatid suggests a demonstrable physical mechanism for coiling (see illustrations of the
rope and elastic chromosome model in Figure 12).
The model is based on the behavior of a £exible
rod (ˆ rope) attached along its length to a strip
of contractile material (ˆ stretched elastic). When
the strip contracts, the whole structure spirals with
the contractile strip taking the shorter route along
the inside of the spiral and the £exible rod taking
184
the longer route to the outside of the spiral. While
an elastic strip has been used to prepare the model,
we are not suggesting that the core is elastic but
that it is contractile. Indeed, Gimënez-Abiän et
al. (1995) have shown that chromatid cores
shorten in length by almost two-fold from early
prometaphase to metaphase. Gimënez-Abiän et
al. (1995) and Nokkala & Nokkala (1984) also
illustrate that each chromatid core is in a lateral
position on each sister chromatid from prophase
through prometaphase but they do not suggest
that shortening cores drives coiling. In regard to
this model, it is interesting that contraction and
coiling of the spasmoneme of Vorticella looks like
it uses the proposed mechanism for chromosome
coiling (see Mahadevan & Matsudaira 2000 for
contraction of the spasmoneme, although again
they do not suggest this model for coiling; see
the cover of Science Vol. 288, No. 5463, April
7, 2000).
As the model is described, one might expect a
hollow (chromatinless) center in each contracted
chromatid, but generally hollow centers are not
observed in aldehyde-¢xed and sectioned chromosomes (Manuelidis & Chen 1990). However, if
contraction is strong enough, the twisted core
simply takes up a central position in the coil
without any central open space. With the elimination of a central space, there is little or no change
in the number of gyres but there is a slight decrease
in diameter of the coiled chromatid. We have demonstrated this with a rope and elastic model (not
illustrated). Techniques that demonstrate coiling
usually require a treatment that swells the
chromatin matrix to separate the gyres of the
chromatid cores (e.g. Matsuura 1938, Ohnuki
1968, Rattner & Lin 1985).
At centromeres the two sister chromatids are
held together by a special set of proteins (Figures
10 & 12D; see Rieder & Cole 1999 for a review
of proteins involved in chromatid cohesion). As
long as the contractile sister cores remain in
intimate association in the center of the chromatin
at the centromere, some shortening can occur here
but no coiling. DNA in centromeres is enriched
in SARs, so DNA loops in centromeres are short
(Strissel et al. 1996). Since sister chromatids are
usually mirror images of each other, this means
that sister kinetochores forming on these short
DNA loops face outward, and kinetochores facing
S. M. Stack & L. K. Anderson
in opposite directions are closely anchored to the
underlying cores (Zhao et al. 2000). In contrast
to active centromeres, inactive or dormant
centromeres do not form constrictions (e.g.
Earnshaw & Migeon 1985 and references therein),
suggesting that inactive centromeres do not have
the special proteins that hold active centromeres
together so inactive centromeres are free to coil.
In reference to secondary constrictions, many of
the proteins involved in rRNA transcription and
processing remain in association with rDNA at
NORs throughout mitosis (Jackson & Cook 1995),
and at least some of these proteins can be
selectively silver stained (e.g. Goodpasture &
Bloom 1975, Schubert 1984). However, unlike
active centromeres, sister NORs do not show
any particular tendency to remain associated when
sister chromatid arms begin to condense and coil.
We propose that the accumulation of proteins at
active NORs interferes with contraction of the core
and coiling, and this results in secondary constrictions. Of course, just as in the case of `primary
constrictions' at centromeres, `secondary constrictions' are not really constrictions of chromosomes but rather thinner regions of the
chromosome due to a lack of coiling. NORs that
were not active in the previous interphase lack
most proteins involved in rRNA synthesis and
processing so they are free to coil and do not form
secondary constrictions (e.g. McClintock 1934,
Tanaka & Terasaka 1972, Jordan & Luck 1976).
Cohesins hold centromeres of sister chromatids
together until sister chromatids are completely
separated by proteolysis at the metaphase/
anaphase transition (see Rieder & Cole 1999
and van Heemst & Heyting 2000 for reviews).
At this time, sister centromeres separate, and primary constrictions disappear (Sumner 1991), indicating that coiling now extends through
centromeres as sister chromosomes are drawn to
opposite poles during anaphase (Figures 11 &
12E).
When the nuclear envelope reforms during
telophase, the contracted and coiled chromosome
cores relax. The 30-nm-diameter chromatin ¢bers
that were held near the chromosome cores are
released, except at their bases, to form extended
loops. Proteins are lost or exchanged from
chromosome cores that now become nuclear
matrix strands which no longer stain with silver.
Chromosome structure through mitosis and meiosis
In short, chromosomes revert to an interphase
chromosome structure characteristic of G1
(Figure 1).
Model for the structure of chromosomes through
the meiotic cell cycle
[See Zickler & Kleckner 1998, 1999 for recent
reviews of meiosis.]
Premeiotic interphase, G1 and S
While the events of premeiotic interphase must be
fundamentally similar to mitotic interphase,
premeiotic interphase lasts much longer
(Kofman-Alfaro & Chandley 1970, Bennett 1977,
Holm 1977). Because leptotene chromosomes that
appear after S phase are exceptionally long, extra
time may be needed for establishing this special
chromosome organization. For example, variants
of H1 histones like meiotin 1 are added to plant
primary microsporocyte nuclei (Qureshi &
Hasenkampf 1995) and H1t is added to mammalian primary spermatocyte nuclei (Moens
1995). More recently, it has been shown in
Coprinus cinereus that proteins known to be
involved in forming double strand breaks during
prophase I also seem to function in premeiotic
S phase where they somehow prepare chromosomes for synapsis and crossing over (Merino
et al. 2000, also see Borde et al. 2000). In
addition, SARs which were not utilized during
the mitotic cell cycle may be enlisted during the
meiotic cell cycle, thereby yielding a longer
chromosome with shorter DNA loops, and SARs
may associate with new and/or additional proteins
in matrix strands. In any case, at the end of
premeiotic S phase, sister DNA loops extend in
opposite directions from duplicated nuclear
matrix strands just as at the end of mitotic S phase
(compare Figures 7 & 13).
Leptotene
With little, if any, delay between the end of S phase
and the beginning of leptotene (e.g Crone et al.
1965, Callan & Taylor 1968, King 1970, Stern
et al. 1975), matrix strands between sister DNA
loops are reinforced with additional meiosis-
185
speci¢c proteins (Offenberg et al. 1998) to form
axial elements (cores) from which contracting
sister DNA (chromatin) loops extend in opposite
directions (Figure 13A, B). An axial element probably consists of two intimately associated, parallel
sister cores (Nebel & Coulon 1962, Dresser &
Moses 1980, and see Zickler & Kleckner 1999
p. 649 for other references), although usually
the double nature of axial elements is not obvious
(see Dawe et al. 1994 and references therein for
examples of visibly double leptotene chromosomes). Since leptotene chromosomes are longer
than at any other time during meiosis (except
lampbrush chromosomes), the axial core is probably a comparatively rigid uncoiled core at this
time (Scherthan et al. 1998).
In Bombyx mori and Lilium longi£orum at early
prophase I, chromatin ¢bers have been reported to
be from 20^30 nm in diameter, which is probably
typical of most chromatin ¢bers during interphase
and mitosis (e.g. Ris & Kubai 1970, Rattner &
Hamkalo 1979, Rattner et al. 1980). However,
unlike mitotic chromosomes, there is transcription
on the loops of early prophase I chromosomes
(Kierszenbaum & Tres 1974, Rattner et al. 1980,
Cook 1997 and references therein), suggesting that
DNA in chromatin loops during early prophase I
is fairly accessible. The importance of an open
chromatin conformation for recombination is
suggested by observations in S. cerevisiae that
hot spots for double strand breaks and recombination are often located in or near promoter
regions (whether functional or not). Promoters
are known to be sites where DNA is exposed
and accessible, and experimental disruption of
nucleosome structure leads to double strand
breaks in the exposed DNA (e.g. Wu & Lichten
1994).
During late leptotene (if not before) and early
zygotene, homologous chromosomes are actively
moved in the nucleus, and this probably helps
homologs come in contact so they can recognize
each other and align (pair) in preparation for synapsis (SC formation) (e.g. Parvinen & SÎderstrÎm
1976, Loidl 1993, Dawe et al. 1994, Scherthan
et al. 1996, 1998, but see Schwarzacher 1997 for
association of homologous arms during premeiotic
interphase in wheat). Initial recognition involves
weak, homologous, paranemic DNA/DNA interactions that are stable only when reinforced by
186
S. M. Stack & L. K. Anderson
Figures 13^19. Diagrammatic representations of meiotic chromosome structure from leptotene through anaphase I. See Figures 1^6
for diagrams of chromosome structures that are the same for mitotic and premeiotic interphase. Drawings are not to scale.
Chromosome structure through mitosis and meiosis
other nearby interactions (Figure 13B; e.g. McKee
et al. 1992, Kleckner & Weiner 1993, CameriniOtero & Hsieh 1993, Weiner & Kleckner 1994,
Gaillard & Strauss 1994, see Cook 1997 for other
references and a model for pairing based on
transcription). Breaks in DNA are not required
for these weak interactions, and the interactions
are not directly involved in crossing over, only homologous pairing. Presynaptic alignment (pairing)
is only possible between homologs with the same
chromatin pattern of exposed DNA loops (Rocco
& Nicolas 1996, also see Karpen et al. 1996). This
is analogous to aligning unique bar codes on commercial product labels (see Figure 20 for a descrip-
187
tion of the bar code model and the role of the
bouquet orientation in presynaptic alignment).
It is important that sister loops are contracted
and reside on opposite sides of chromosomes to
avoid saturating homologous DNA^DNA interactions with interactions between sister loops of
DNA. In support of this idea, leptotene through
pachytene is the only period in the life cycle of Saccharomyces cerevisiae when chromatin is reported
to be contracted (Dresser & Giroux 1988). A
related observation involves a 1.7-Mb lambda
transgene that forms a long loop extending from
a mouse SC where its base is anchored (Moens
et al. 1997b). Presumably, the long loop is due
Figure 13. Leptotene. (A) Longitudinal view of a segment of a leptotene chromosome after DNA replication is complete (see Figure 6).
Proteinaceous axial elements (red^violet) assemble in association with MARs (green dots) at the base of sister DNA loops. Although
not usually obvious, each axial element consists of two closely associated sister elements. Chromatin condenses by collapsing 30 nm
diameter chromatin loops toward axial elements. The use of more MARs may additionally shorten chromatin loop lengths. Sister
DNA loops are represented in the same color but on opposite sides of the axial element. (B) End view of segments of two homologous
leptotene chromosomes. For each chromosome, sister DNA loops (same color) extend from axial elements in opposite directions.
Homologous DNA loops are shown in different shades of the same color. For example, the black and gray loops are homologous
as are the light green and dark green loops. Because the chromatin loops have condensed, it is dif¢cult for sister chromatin loops
to contact one another, whereas homologous loops are free to associate. Here, (green) homologous DNA loops have made contact.
Such contacts mediate presynaptic alignment. Figure 14. Zygotene/pachytene. End view of a segment of synaptonemal complex
(SC). SCs form during zygotene^pachytene. When incorporated into SCs, axial elements are renamed lateral elements (red^violet).
Each lateral element is shown divided into two sister elements. Occasionally a lateral element twists so the lower sister element
(as viewed here) moves to the upper side and the upper sister element moves to the lower side (not shown). Twisting of lateral elements
relative to one another is likely because leptotene chromosomes have been reported to be twisted (Oehlkers & Eberle 1957). This
accounts for observations that either sister in each homolog may be involved in a crossover. During SC formation, sister DNA loops
(shown the same color) are de£ected somewhat from a strictly opposite arrangement around lateral elements (LEs) to make room
for transverse ¢laments in the central region of the SC. Even so, sister loops are sterically inhibited from coming into contact with
each other. On the other hand, homologous loops (loops of similar colors, e.g. light green and dark green loops) are in close association
and free to interact for repair and recombination. Here a recombination nodule mediates a crossover between the two homologous
green loops. Figure 15. Pachytene. Frontal view of a segment of synaptonemal complex. Chromatin loops extend from lateral
elements. Sister chromatin loops are represented by thick and thin lines of the same shade of green. At the recombination nodule,
DNA loops from two non-sister chromatids are involved in a crossover. Figure 16. Diffuse stage (early diplotene). Homologs desynapse with the disintegration of the SC. However, one or more lateral element proteins may remain in place and other proteins
may be added to hold sister chromatids together through the diffuse stage (light gray rods). In the ¢gure, proteins (probably including
some from the RN) are associated at the crossover site (gray circle). Figure 17. Late diplotene. (A) Longitudinal view during the
transition from the diffuse stage to late diplotene showing new chromatid cores (copper ^ comparable to those in mitotic chromosomes)
forming in association with MARs (green dots) at the base of loops comprising each continuous DNA strand. These new cores
faithfully follow chromatids though cross overs and thereby form chiasmata. (B) End view of a segment of a diplotene bivalent (not
at a chiasma). Sister chromatid cores are close together due to sister chromatid cohesion and lie to one side of the chromatin as
was the case for lateral elements in pachytene. Even so, sister loops extend roughly in opposite directions. Also see Figure 20. Figure
18. Diakinesis^metaphase I. Three-dimensional drawing of a bivalent at lower magni¢cation than the diagrams in Figures 13^17.
From diakinesis through metaphase I, sister chromatids are held together throughout their length (sister chromatid cohesion) with
sister cores to one side of the chromatin. When sister cores actively shorten, sister chromatids coil together. Between chiasmata,
coiling can be achieved by including roughly the same number of right- and left-hand twists. Between the end of a bivalent and
the ¢rst chiasma, coiling can be all right- or left-handed since the ends can rotate to relieve tension. The bivalent diagrammed
in this ¢gure has two chiasmata, and each chromosome has two coiling reversals between the chiasmata. The two sister cores (copper)
of each chromosome are indicated by arrows. To the lower right, the relation of chromatin loops and the cores is illustrated. Sister
kinetochores (yellow) face the same pole. Also see Figure 22A. Figure 19. Anaphase I. Three dimensional drawing of two homologous
anaphase I chromosomes. Sister chromatid cohesion is lost in the arms but maintained at centromeres. As a result, sister chromatid
arms swing apart, chiasmata are lost, and homologous chromosomes are pulled to opposite poles by kinetochore microtubules. Also
see Figure 22B.
188
S. M. Stack & L. K. Anderson
Figure 20. The bar code model for presynaptic alignment of homologous chromosomes is based on two premises: Each chromosome
displays a unique sequence of DNA loops in a linear pattern that is analogous to bar codes used for scanning commercial products,
and homologous loops of DNA interact weakly to bind them together. (A) Non-homologs (represented by the unlike upper and
lower bar codes) have few, if any, like sequences, so they cannot form stable associations when brought into contact. (B) Homologs
(represented by the upper and lower identical bar codes) cannot form stable associations unless they are aligned at some point.
(C) The bouquet orientation of leptotene and zygotene chromosomes requires movement of chromosomes within the nucleus
and encourages alignment of chromosomes starting at their ends on the nuclear envelope (see Zickler & Kleckner 1998 for a review
of the bouquet orientation). Indeed, heterozygosity for distal de¢ciencies and asymmetry in isochromosome arms seriously interferes
with synapsis, presumably due to dif¢culty in homologous alignment when telomeres are attached to the nuclear envelope (Curtis
et al. 1991, Lukaszewski 1997). Once homologous loop interactions begin, homologs (represented by the upper and lower identical
bar codes) can zip together by an increasing number of weak, but additive, homologous loop interactions.
Chromosome structure through mitosis and meiosis
to a lack of MARs/SARs in the insert. In a
homozygote for the transgene, all four inserts
are nearby and can make contact to form a single
FISH signal, probably based on the same weak
interactions that lead to presynaptic alignment.
In a heterozygote where inserts are on nonhomologs, sister inserts associate to form a single
FISH signal, but inserts on non-homologs yield
separate signals, i.e. there is no ectopic pairing
of inserts on non-homologous chromosomes. This
observation could be explained according to the
model if sister loop interactions do not leave enough
unsaturated sequence to mediate stable ectopic
interactions between non-homologs that otherwise
would not be attracted to each other.
Because chromatin loops are ordinarily contracted, there are always many loops on one
homolog that are sterically unavailable for interactions with loops on the other homolog (Figure
13B). In polysomic and polyploid cells there are
more than two homologs, so unused DNA loops
on an aligned pair of homologs are available to
associate with loops on other homologs. The result
is that presynaptic alignment is not saturated by
associations of chromosome pairs, but can involve
simultaneously as many homologs as are present
in a cell (e.g. Loidl 1988).
Homologous interactions in late leptotene draw
homologs into close enough association that
transverse ¢lament proteins attached at their C
termini to axial cores can interact head to head
(N termini to N termini) to form transverse
¢laments that span the central region between
the axial cores (Westergaard & von Wettstein
1966, Schmekel et al. 1996, Liu et al. 1996, Dong
& Roeder 1999). Once transverse ¢laments are
in place, synapsis has occurred, and axial cores
are now referred to as lateral elements.
Collectively the lateral elements and transverse
¢laments constitute the SC (Figure 14, see
Schmekel et al. 1993 for structure of the SC).
By drawing axial cores together in this way, there
is a tendency for synapsis to proceed in either
direction from an initiation site. While synapsis
may be completed from a single initiation site
(Greenbaum et al. 1986), usually there are multiple
sites of synaptic initiation, especially on large
chromosomes (e.g. Hasenkampf & Menzel 1985).
Generally synapsis is not initiated in heterochromatin (e.g. Stack & Anderson 1986a).
189
Early recombination nodules are ellipsoidal,
darkly-stained, proteinaceous bodies that are
found in association with axial elements during
leptotene and in the central regions of SCs during
zygotene (Figure 14; e.g. Stack & Anderson
1986a). Early nodules are often observed between
segments of axial cores that approach each other
prior to synapsis and at forks where synapsis
has just occurred (Albini & Jones 1987, Anderson
& Stack 1988, Stack & Roelofs 1996). Although
proteins that are capable of mediating DNA^
DNA interactions are components of at least some
early nodules (such as Rad51p and Dmc1p;
Anderson et al. 1997, Moens et al. 1997a, Tarsounas et al. 1999), it remains unclear whether
early nodules are involved in presynaptic alignment or synapsis (von Wettstein et al. 1984, Stack
& Anderson 1986b, Zickler & Kleckner 1999).
An important feature of this model is that sister
DNA loops are contracted on opposite sides of lateral elements. As a result, sister loops are sterically
inhibited from interacting with each other during
crossing over, gene conversion, or other DNA
repair events. In contrast, homologous non-sister
DNA loops can come into contact above, below,
and possibly through the central region of the
SC (Figure 14). This arrangement of sister loops
would explain why non-sister chromatids are
involved in crossing over in preference to sister
chromatids (Haber et al. 1984, Game et al. 1989,
Collins & Newton 1994, Schwacha & Kleckner
1994, 1997). Observations of red1 mutants in S.
cerevisiae are consistent with this model
(Schwacha & Kleckner 1997). The RED1 gene
encodes a lateral element protein. Mutants of
RED1 do not form lateral elements, and they
do not have an interhomolog bias in crossing over.
Schizosaccharomyces pombe and Aspergillus
nidulans are also relevant to this discussion
because both go through meiosis without forming
SCs and yet both have high levels of crossing over
without crossover interference (Olson et al. 1978,
Egel-Mitani et al. 1982). Because axial elements
appear to be formed in S. pombe (Olson et al. 1978,
BÌhler et al. 1993), while axial elements seem not
to be formed in A. nidulans (Egel-Mitani et al.
1982), this model predicts that S. pombe will show
a bias against crossing over between sister
chromatids, while A. nidulans will not. However,
because Scherthan et al. (1994) report that S.
190
pombe is unusual in having no chromatin condensation at prophase I, it is possible that sister loops
can make contact during prophase I to permit a
bias towards sister strand crossing over in this
species as well.
In regard to recombination nodules that are
probably involved in the molecular events associated with gene conversion and crossing over
(Carpenter 1975 and subsequently many additional references), their location on one side or
the other of the central region of an SC would
permit a recombination nodule to be in contact with
homologous non-sister DNA loops much more easily than with sister DNA loops (Figure 14). Again
this would bias crossovers toward non-sister
chromatids. However, plant and animal mutants
that do not form lateral elements or SCs have
essentially no crossing over between homologs
(Moses 1968), and fungal mutants that do not
form lateral elements also have reduced or
eliminated crossing over (e.g. Smith & Roeder
1997, van Heemst & Heyting 2000). Because of
this, one must conclude that lateral elements and
SCs play a more active role in crossing over than
simply suppressing crossing over between sister
chromatids. Most likely this involves association
of recombination nodules with axial/lateral
elements (Figures 14 & 15; e.g. Stack & Anderson
1986a, 1986b, Storlazzi et al. 1996).
SCs are sometimes observed to be twisted
(Zickler & Kleckner 1999). While a variety of
explanations have been offered to account for
twisting of whole SCs, a simple explanation is
based on active shortening (contraction) of the lateral elements that are attached to relatively
incompressible chromatin on one side and rigid
transverse ¢laments on the other side. If transverse
¢laments can swivel or bend to some degree at their
attachment sites to lateral elements, contraction of
lateral elements will cause them to spiral around a
longitudinal axis with the chromatin to the outside
(demonstrated in a rope and elastic model but
not illustrated).
Breakdown of SCs marks the end of pachytene
and the beginning of diplotene. In many
organisms, this point is referred to as the diffuse
stage because nuclei take on an interphase-like
appearance (Figure 16; e.g. Moses 1968, von
Wettstein et al. 1984, see Wilson 1928 and
Kläs̄terskä 1976, 1977 for reviews of the diffuse
S. M. Stack & L. K. Anderson
stage). Breakdown of SCs is a necessary prerequisite for reformation of chromatid cores that re£ect
the results of crossing over, i.e. chiasmata (e.g.
Rufas et al. 1982, Stack 1991). While some SC proteins are lost from chromosomes at this time, a few
lateral element proteins remain with the
chromosomes, particularly at the centromeres (e.g.
Moens et al. 1987, Moens and Spyropoulos 1995,
Molnar et al. 1995, Klein et al. 1999, Zickler
and Kleckner 1999). Some of these lateral element
proteins may be incorporated into newly formed
chromosome cores (e.g. Moens & Earnshaw 1989,
Fedotova et al, 1989, van Heemst & Heyting
2000). Considering the position of the diffuse stage
in the meiotic cell cycle, this interphase-like stage
is probably equivalent to G2 in a mitotic cell cycle.
Indeed, since G2 is de¢ned as that part of
interphase after S phase and before chromosomes
condense at prophase (Howard & Pelc 1953 and
see below), it is attractive to interpret leptotene,
zygotene, pachytene, and the diffuse stage
collectively as a long G2 phase during which structures (axial cores, SCs, early and late recombination nodules) that are needed for crossing
over and gene conversion between homologous nonsister chromatids form, carry out their functions,
and break down (also see Beerman 1963, van
Heemst & Heyting 2000).
There is a great deal of transcription during the
diffuse stage (Kläs̄terskä 1977), and this is the
stage when lampbrush chromosomes appear in
primary oocytes. Indeed, interpreting the diffuse
stage as the end of G2 suggests that lampbrush
chromosomes represent highly extended and extensively transcribed late interphase chromosomes.
For the chromosome model presented here, it is
of interest that sister loops probably extend to
opposite sides of lampbrush chromosome axes
(Figure 21 and see other illustrations in Callan
& Lloyd 1963 and Callan 1986) just as sister loops
are hypothesized to emerge from opposite sides
of mitotic G2 nuclear matrix strands, axial
elements, and lateral elements.
After the diffuse stage, long diplotene chromosomes become visible (Figure 17). As chromosomes shorten in late diplotene and diakinesis, a
pair of closely associated sister cores that can
be stained with silver becomes visible in each
chromosome (Rufas et al. 1982, 1987, Stack 1991,
Gimënez-Abiän et al. 1997). These paired cores
Chromosome structure through mitosis and meiosis
191
Figure 21. Camera lucida drawing of a lampbrush chromosome from a crested newt (Triturus cristatus (Laurenti)) primary oocyte.
Note that sister loops of chromatin extend in opposite directions from the axes. [Callan, HG and Lloyd, L (1963), Lampbrush chromosomes of crested newts Triturus cristatus (Laurenti). Roy Soc Lond Phil Trans Ser B 243: 135^219, Figure 9a; reproduced with permission.]
are comparable to those in paired sister chromatids
of mitotic prophase chromosomes (Gimënez-Abiän
et al. 1995), except diplotene cores trade partners
at chiasmata (compare Figures 9 & 17). While
it is unclear how newly formed chromosome cores
accurately track DNA through chiasmata, the
problem may be no different and the mechanism
the same as when newly formed sister cores track
sister DNA double helices in G2-prophase of
mitosis.
During diplotene^metaphase I, sister chromatid
cohesion maintains chiasmata and keep homologs
together (Darlington 1932, Maguire 1974). In
grasshoppers, sister cores are close together in
the center of the chromatin mass and, as a result,
there is no coiling (Rufas et al. 1987). In Lilium
longi£orum (lily), sister chromatids coil together
during diakinesis-metaphase I (Stack 1991). These
pairs of sister cores are close but separate and
lie to one side of the chromatin of the sister
chromatids (Figure 17B). In this lateral position,
active contraction of the sister cores during
diplotene^diakinesis results in coiling, just as in
mitosis (Figures 18 & 22A). Because there are
two close, parallel sister cores (rather than the separate single sister cores observed during mitosis),
the major coils of lily chromosomes at diakinesis
have larger gyres than lily mitotic chromosomes
(Stack 1991). Distal to chiasmata, the strain of
coiling is relieved by twisting sister chromatids
around each other in the same direction as the
coiling. However, strain caused by chromosome
coiling between chiasmata cannot be relieved like
this. Instead, there are reversals of the direction
of coiling to yield little or no net coiling or twisting
between chiasmata. Active contraction of cores
and coiling of chromosomes during diplotene^
diakinesis, causes them to shorten and thicken. This
puts tension on chiasmata and causes chromosomes
to bulge out between chiasmata, a phenomenon
sometimes referred to as repulsion (Figures 18 &
22A; Ostergren 1943).
It should be mentioned that certain plants (such
as Tradescantia and lily) have very large meiotic
chromosomes that have sometimes been reported
to consist of large (major) coils of a smaller
(minor) coiled strand (e.g. Kuwada 1938, Manton
1950, Taylor 1958, Stack 1991). If, indeed, two
levels of coiling can coexist in the same
chromosome, this is not readily explained with
a single contractile core in each chromatid.
The chromosome model proposed here provides
an explanation for how the two pairs of sister
kinetochores in a metaphase I bivalent are
oriented to opposite poles (amphitelic orientation). The geometry of a coiled bivalent dictates
that one pair of sister kinetochores is located to
one side of the chromatin mass while the other pair
is located to the other side. In addition, the
side-by-side arrangement of sister chromatids
during diakinesis^metaphase I positions both sister
kinetochores on one side of the chromosome so
microtubules from one pole attach to both of them
(syntelic orientation), while the other two sister
kinetochores on the homologous chromosome
attach to microtubules from the opposite pole
(syntelic orientation) to establish amphitelic
orientation for the whole bivalent (Figures 18 &
22A).
At anaphase I, sister chromatid binding is lost in
the arms, but not at centromeres (Figures 19 &
22B; e.g. Moens & Spyropoulos 1995, Sekelsky
192
S. M. Stack & L. K. Anderson
Figure 22. Rope and elastic models of meiotic chromosomes. The chromatin of each chromatid is represented by a segment of white
rope that is 53 cm long in all models. The chromatid core is represented by a black elastic band that was sewn to the white rope.
Kinetochores (K) are indicated by felt patches. In these models, 1 cm represents 1 mm, i.e. the ropes are 10 000 : 1 scale models
of chromosomes. (A) Diakinesis^metaphase I chromosome. To prepare the model, a 53-cm length of white rope was sewn to a length
of elastic maximally stretched to 53 cm. This represents one chromatid. This was repeated three more times for a total of four ropes
ˆ four chromatids. While still maximally stretched, the proximal two-thirds of two lengths of rope/elastic were sewn together
side-by-side to represent one homolog, and the proximal two-thirds of the other two lengths of rope/elastic were similarly sewn
together to represent the other homolog. The ropes were not sewn together elastic band to elastic band because the cores look nearby
but separate in lily prometaphase I chromosomes (Stack 1991). For a distal chiasma, one rope end (ˆ chromatid) from one rope
pair (ˆ homolog) was sewn in the same orientation to another rope end (ˆ chromatid) from the other rope pair (ˆ homolog). This
process was repeated for the other rope ends. The pairs of rope ends distal to a partner trade represent pairs of sister chromatids
held together by sister chromatid cohesion. The same sewing was repeated to represent another distal chiasma at the other end
of the `bivalent'. Then, the tension on all four ropes was released to permit coiling. Reversals of coiling occur between chiasmata
because tension cannot be relieved by rotating the ends of the chromosomes. Two pairs of sister kinetochores (K) face in opposite
directions. Sister cores (double arrows) to the inside of the coils. Also see Figure 18. (B) Separating anaphase I chromosomes.
In this model, each of four 53-cm lengths of white rope was sewn to a 53 cm length of maximally stretched elastic band. While
still maximally stretched, two of the ropes were sewn together side-by-side for a short distance at the centromeres to represent sister
chromatid cohesion that is still present at this location. The ropes were sewn together side-by-side as a continuation of their relationship from metaphase I. When tension on the ropes was released, the ropes coiled along their length with the sister kinetochores
facing outward in the same direction. A second model of an anaphase I chromosome was prepared in the same way using the other
two lengths of rope/elastic. Then the two models were arranged as they would look separating at anaphase I. Also see Figure 19.
Chromosome structure through mitosis and meiosis
& Hawley 1995, Klein et al. 1999). With the loss of
sister chromatid cohesion, each arm coils
independently. However, because the sister centromeres are still held together in a side-by-side
orientation, the coiled arms swing apart in a
manner consistent with Ostergren's (1943) model
of the response of two elastic bodies bound
together at a point. Loss of sister chromatid
cohesion along the arms eliminates chiasmata,
and homologous chromosomes are pulled to
opposite poles by tension on kinetochore
microtubules. Telophase I and a modi¢ed interphase called interkinesis occur between meiotic
divisions. The chromosomes decondense, uncoil,
and lengthen, but there is no DNA synthesis
and no reorganization of chromosome structure.
Chromosomes that recondense at prophase II
are little altered from anaphase I. Indeed, many
animal eggs largely or completely skip telophase
I and interkinesis to move directly into prophase
II (Sharp 1926). In either case, sister kinetochores
still lie to one side of metaphase II chromosomes
(e.g. Gimënez-Abiän et al. 1997) but anaphase
II cannot begin until sister kinetochores are under
tension due to establishing amphitelic orientation
on the spindle (Nicklas et al. 1995). The second
meiotic division is otherwise a typical mitotic division of a haploid cell (Figures 7^11 & 12B^E).
Conclusion
We propose three important additional features to
the chromosome scaffold model: (1) DNA loops
extend from only one side of a chromatid core;
(2) chromatid cores are actively contractile; and
(3) after DNA replication, sister DNA loops
extend in opposite directions from sister cores.
Together, these proposed features provide a mechanical explanation for chromatid coiling and a
steric explanation for the preferential involvement
of non-sister chromatids in crossing over. Based
on these proposals, we make speci¢c testable
predictions about chromosome structure.
Acknowledgements
We thank Paul Kugrens for assistance with the
illustrations and Maria Pigozzi for carefully
193
reading the manuscript. This work was supported
in part by NSF grant MCB-9728673.
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