Download growth polarity and cytokinesis in fission yeast: the role of the

J. Cell Sci. Suppl. 5, 229-241 (1986)
Printed in Great Britain © The Company of Biologists Limited 1986
229
GROWTH POLARITY AND CYTOKINESIS IN FISSION
YEAST: THE ROLE OF THE CYTOSKELETON
JO H N M A R K S , I A I N M . H A G A N
and
JE R E M Y S. H Y A M S *
D epartm ent o f Botany ¿2? Microbiology, University College London, Gower Street,
London WC1E 6BT, U K
S UMMA R Y
The distribution of F-actin in the fission yeast Schizosaccharomyces pombe was investigated by
fluorescence microscopy using rhodamine-conjugated phalloidin. Fluorescence was seen either at
the ends of the cell or at the cell equator. End staining was predominantly in the form of dots whilst
equatorial actin was resolved as a filamentous band. The different staining patterns showed a close
correlation with the known pattern of cell wall deposition through the cell cycle. In small, newly
divided cells actin was localized at the single growing cell end whilst initiation of bipolar cell growth
was coincident with the appearance of actin at both ends of the cell. As cells ceased to grow and
entered cell division, a ring of actin was seen to anticipate the deposition of the septum at
cytokinesis. The relationship between actin and cell wall deposition was further confirmed in three
temperature-sensitive cell division cycle (cdc) mutants; cdc 10, cdc 11 and cdc 13. Immunofluor­
escence microscopy of S. pombe with an anti-tubulin antibody revealed a system of cytoplasmic
microtubules extending between the cell ends. The function of these was investigated in the coldsensitive, benomyl-resistant mutant ben A. In cold-grown cells actin was seen to form conspicuous
filamentous rings around the nucleus. The origin of these and the possible role of microtubules in
the cell-cycle-dependent rearrangements of F-actin are discussed.
INTRODUCTION
The use of simple model systems to study complex problems is a familiar strategy
in biological research. Recent developments in molecular genetics have reinforced
the value of organisms such as yeasts in the study of fundamental cellular processes.
This is particularly true in the case of the cytoskeleton. Although the small size of
yeast cells and the presence of a cell wall present obstacles to traditional approaches
to the organization of cytoplasmic filament systems such as immunofluorescence
microscopy, these have now been largely overcome (Kilmartin & Adams, 1984;
Adams & Pringle, 1984). Coupled with the analysis of cloned actin and tubulin
genes (Thomas et al. 1984; Yanagida et al. 1985) this opens up a combination of
approaches to cytoskeleton structure and function that is possible in few other
organisms.
An additional attraction of yeasts as experimental systems is the availability of a
large number and variety of mutants that affect the yeast cell division cycle (Pringle
& Hartwell, 1981; Nurse, 1981). Thus, the relationship of the cytoskeleton to other
cell cycle events is also open to direct study. Two yeasts in particular have been the
focus of most attention, the budding yeast Saccharomyces cerevisiae and the fission
•A u th or for correspondence.
230
J . Marks, I. M. Hagan andjf. S. Hyams
yeast Schizosaccharomyces pombe. Of the two, 5. pombe may prove to be the better
model, if only because its cell division cycle more closely resembles that of higher
eukaryotes (Nurse, 1985).
Fission yeast cells grow only at their ends (Johnson, 1965; Streiblova & Wolf,
1972; Mitchison & Nurse, 1985). In newly divided cells growth occurs solely at the
old end, that is, the end that existed prior to septation. At a point in the Gz phase of
the cell cycle termed N E T O (new-end take off) bipolar growth is initiated. This is
maintained until the end of Gz, at which point growth ceases and the sequence
of mitosis, cytokinesis and cell separation begins. In this paper we describe the
relationship of the cytoskeleton to these changes in growth polarity. We also address
the role of the cytoskeleton in establishing the division plane in these simple
eukaryotes, both in wild-type cells and in various cell division cycle (cdc) mutants.
MATERIALS AND METHODS
Wild-type S. pombe strain 972h_ and the mutant strains cdclQ, 129h_ , cdc 11, 136h~ and cdc 13,
117h- (Nurse et al. 1976) were kindly supplied by D r Paul Nurse; strain ben A, D3 (Roy & Fantes,
1982) was kindly supplied by D r Peter Fantes. Cultures were grown as previously described
(Marks & Hyams, 1985). Temperature-sensitive cdc strains were grown at 25°C to mid-log phase
prior to arrest at 36°C for 5-7 h. The cold-sensitive mutant ben A was grown at 36°C to mid-log
phase and blocked at 22-5°C for 24h. Intracellular F-actin distribution was determined by
phalloidin staining (Wulf et al. 1979; Wieland & Govindan, 1974) according to Marks & Hyams
(1985). Nuclear morphology, and thus the position of the cell in its mitotic cycle, was determined
using DA PI (4'-6-diamidino-2-phenylindole; Williamson & Fennell, 1975). Calcofluor white was
used to reveal both the cell wall and the septum (Darken, 1961). Tubulin staining of wild-type cells
using the monoclonal antibody to yeast tubulin, Y O L 1/34 (Kilmartin et al. 1982), was performed
essentially after the method of Kilmartin & Adams (1984).
RESULTS
A field of 5. pombe cells stained with rhodamine-conjugated phalloidin as a probe
for F-actin is shown in Fig. IB. Fluorescence is seen either at one end of the cell, at
both ends or at the cell equator. End staining is mainly in the form of dots whereas
equatorial actin is resolved as a filamentous ring. The different staining patterns may
be ordered with respect to the cell cycle by reference to Fig. 1A, which shows the
same field of cells stained with the cell wall stain Calcofluor and the D NA probe
D API. Newly divided 5. pombe cells grow only at the old end (Mitchison & Nurse,
1985). The two cell ends can be distinguished by their affinity for Calcofluor, the old
(growing) end staining brightly whilst the new (non-growing) end appears as a dark,
unstained hemisphere (Fig. 1A, cell 1 ). When new-end growth is initiated (N ETO )
this dark region is internalized and appears as a birth scar on the cell surface
(Fig. 1A, cell 2 ). Comparison of the phalloidin staining patterns reveals that the
distribution of actin coincides precisely with the polarity of cell growth; namely,
actin resides at the single growing end before N ETO and at both growing ends after
N E T O (compare cells 1 and 2 in Fig. IB). The transition from one-end to two-end
staining also sees the transient appearance of fine filaments of F-actin possibly
extending the length of the cell (cell J , Fig. IB).
Fission yeast cytoskeleton
231
F ig. 1. T w o views of the same field of asynchronous wild-type S . pombe cells: A, stained
with C alco flu or-D A P I; B, stained with rhodamine-conjugated phalloidin. N um bers
designate the staining patterns referred to in the text. Bar, 10 j.lm.
Double-end actin staining is maintained until the onset of mitosis (cell 4, Fig. 1A),
which coincides with the completion of end growth. Actin now disappears from the
poles but reappears as a ring at the cell equator where it overlies the dividing nucleus
(cell 4, Fig. 1A ,B). T h e position of the actin ring anticipates the deposition of the
septum , which stains intensely with Calcofluor (cell 5, Fig. 1A). As the septum
grows centripetally the appearance of the equatorial actin changes from a filamentous
ring to clusters of dots (cell 6, Fig. IB ). At the completion of septation the remnants
of the equatorial actin lie at the new ends of the two daughter cells (cell 7, Fig. IB ).
Since growth will be initiated at the opposite (old) end, actin must rapidly relocate to
the other end of the cell before the next cell division cycle can begin. This transition
is again accompanied by the transient appearance of fine actin fibres (not shown).
T he complete sequence of actin distribution through the S. pombe cell cycle is
summarized in Fig. 2.
Further evidence for the relationship between actin and cell growth and division
in fission yeast has been obtained from various temperature-sensitive cdc mutants.
These become arrested at a specific point in the cell division cycle when grown at the
restrictive temperature although their metabolic processes are maintained and they
continue to elongate (Bonatti et al. 1972; Nurse et al. 1976). Fig. 3 shows Calcofluor
and phalloidin images of cdc 10, which arrests in G\, i.e. prior to N E T O . Intense
232
J . Marks, I. M. Hagan andy. S. Hyams
actin staining is seen at the growing old end although a few dots are also seen at the
new end (Fig. 3B). Correspondingly, although Calcofluor staining indicates that
growth is predominantly at the old end, a small amount of cell wall deposition at the
new end is also detectable (Fig. 3A). None of the cells display equatorial actin nor do
they form septa. Similar images of cdc 13 are shown in Fig. 4. This mutant arrests in
mitosis although under certain conditions a proportion of the cells leak through the
temperature block and form multiple, aberrant septa (Fig. 4A). Growing cell ends
(as judged by Calcofluor staining) again reveal intense actin fluorescence in the
form of both dots and fibres (cells 1, 2 and 3 in Fig. 4B). Actin dots also occur in
the region of the multiple septa in cell 4 whilst cells 5 and 6 reveal the equatorial
actin ring, which anticipates the septum. The contiguous nature of the ring is
particularly clearly seen in these cells. In cell /, the equatorial actin is largely
dispersed following the formation of a septum and intense staining is again seen at the
cell poles.
Fig. 5 shows the relationship of the equatorial actin ring to nuclear position in an
early septation mutant cdc 11. These cells are unable to undergo cytokinesis at the
restrictive temperature although nuclear division and cell elongation continue. At the
first mitosis following the temperature block the daughter nuclei return to a point
either side of the middle of the cell (cell 1, Fig. 5B). Reinitiation of end growth is
indicated by the bipolarity of actin staining (cell 1, Fig. 5A). Although these cells do
not form a septum, a pair of actin rings forms at the positions occupied by the two
nuclei prior to mitosis (cell 2, Fig. 5A). The nuclei do not appear to be physically
connected to their respective actin rings since they are able to move away from their
original position at the second mitotic division (cell 2, Fig. 5A,B). At the next
mitosis actin rings appear at the position occupied by each of the four daughter nuclei
(not shown).
Although not excluding other possibilities, we have begun to investigate whether
the changing patterns of actin described above are dependent on the presence of
cytoplasmic microtubules. S. pombe cells stained with anti-tubulin antibody are
shown in Fig. 6. During interphase, groups of microtubules extend between the two
ends of the cell. As the cell enters mitosis, these cytoplasmic microtubules disappear,
to be replaced by an intranuclear spindle. The precise details of the cell cycle
rearrangements of tubulin in S. pombe will be presented elsewhere (I. M. Hagan,
P. Nurse & J. S. Hyams, unpublished data). Since anti-microtubule drugs have only
a limited effect on fission yeast (Bums, 1973; Walker, 1982; our unpublished results)
we have attempted to destabilize cytoplasmic microtubules by genetic means. The
cold-sensitive, benomyl-resistant mutant benA is unable to undergo cell division at
20°C (Roy & Fantes, 1982). When cold-grown cells are stained with phalloidin, a
dramatic rearrangement of actin is observed. Instead of multiple small dots at the
ends of the cells, a single large dot is frequently present. Most obviously, however,
much of the cellular F-actin is seen to be associated with the nucleus. Predominantly,
this is in the form of a continuous ring, although sometimes this is twisted to form a
figure eight and often bears a tail like a hangman’s noose (Fig. 7). These various
perinuclear configurations are present in up to 90 % of the cells.
233
Fission yeast cytoskeleton
(Os
O
O
B
-e-
o
$
H
F ig. 2. Schem atic representation of the structural rearrangements of F-actin through the
cell division cycle of 5 . pombe. T h e old end of the cell is at the top of the figure. T h e open
circle in the centre of the cell is the nucleus, dots show the position of actin. Because of the
asym m etric nature of cell growth in S . pombe, the cell wall material inherited by each
daughter cell from its mother is different. The cell depicted in A has a ‘waistcoat’ of old
cell wall material, which acts as a useful marker for the directionality of new wall
deposition. Note that at the end of the sequence (cell-H) only one daughter inherits the
waistcoat. T h e major transitions of actin distribution are seen in cell C, which is at
N E T O , where actin filaments are also transiently observed; cell E , which shows the
com pletion of end growth and the initiation of the actin ring (note that the shape of the
nucleus has changed to signify that these events are coincident with the initiation of
m ito sis); and cell H, where actin left at the cell equator by the breakdown of the actin ring
m ust relocate to the opposite end of each daughter cell in order that new cell growth may
be initiated. Fibres also appear transiently at this stage. (Redrawn from Marks & Hyams
(1985).)
F ig. 3. Tem perature-attested cells of cdc 10 double stained with: A, Calcofluor; and
B , phalloidin. Note that as these cells block before N E T O both growth and the location
of actin are restricted largely to the old ends. Bar, 10 fxm.
234
J . Marks, I. M. Hagan and J . S. Hyams
£
**
2
4A
F ig. 4. Calcofluor (A) and phalloidin (B) images of temperature-arrested cdc 13 cells.
N o te : the intense apical staining associated with the growing cell ends in cells 1 , 2 and 3 ;
the diffuse arrays of actin dots associated with the multiple septa in cell 4 ; the actin ring
preceding septum deposition in cells 5 and 6. Bar, 10 fxm.
235
Fission yeast cytoskeleton
m *.
4%
5A
F ig. 5. P h ase-D A P I (A) and phalloidin (B) images of two temperature-arrested cdc 11
cells. At the restrictive tem perature these cells undergo repeated nuclear divisions but do
not divide. Following the first m itosis the daughter nuclei return to the middle of the cell
and end growth is reinitiated (cell / ) . As the daughter nuclei synchronously enter the
next mitotic division an actin ring appears at the site formerly occupied by the nuclei (cell
2 ) . T h e fact that cell 1 contains two interphase nuclei as opposed to a single mitotic
nucleus is confirmed by the obvious difference in size between the nuclei in these two
cells. Bar, lOjUm.
DISCUSSION
T h e precise manner of cell growth in S. pombe coupled with the availability of
mutants affecting both the cytoskeleton and the cell division cycle make this a most
attractive organism in which to investigate the structural rearrangements and inter­
actions of cytoskeletal proteins through the cell cycle and the mechanisms whereby
these are integrated with other cellular events. In this paper we have shown that the
two major growth transitions in the S. pombe cell cycle, that is, from monopolar
to bipolar cell growth early in Gz (N E T O ), and the cessation of end growth and
the initiation of cell division are accompanied by corresponding rearrangements of
F-actin. T hese findings, which were originally established in wild-type cells (Marks
& Hyam s, 1985), have been confirmed here by the use of cell division cycle mutants
arrested at different points of the cell cycle by growth at the restrictive temperature
(N urse et al. 1976). In cdc 10 both actin and cell growth were predominantly mono­
polar (consistent with the known execution point of this mutant before N E T O ). It
was noticeable, however, that a small amount of actin staining, and a corresponding
degree of Calcofluor staining, was always detectable at the opposite pole. A small
amount of new-end growth prior to N E T O has been detected in wild-type cells
although its extent has been difficult to assess, partly because it represents only a
minor contribution to total cell growth and also because the methods of analysis used
to date are relatively crude (Mitchison & Nurse, 1985). T he problem may be
236
J . Marks, I. M. Hagan and y . S. Hyams
resolved when more sophisticated methods for detecting zones of cell expansion in
yeasts are applied to 5. pombe (Staebell & Soli, 1985), or through the use of mutants
such as cdc 10, which can be held prior to N E T O for extended periods. Preliminary
observations of cd c\\ have shown that the normal relationship between nuclear
division and the formation and disappearance of the equatorial actin ring is
maintained even in the absence of septation. Mitchison & Nurse (1985) noted that
cdc 11 showed pulses of cell growth interspersed with periods of quiescence. The fact
that actin disappears from the cell ends with each cycle of ring formation provides an
explanation for their observations. The relationship between the actin ring and
septation is also clearly shown in cdc 13, where the ring cycle and nuclear cycle
become uncoupled and multiple septa are laid down in the absence of nuclear
division.
At present we cannot distinguish whether the coincidence of actin and polarized
cell growth is a cause or an effect relationship. In the case of septation, however,
the situation is unambiguous, actin appearing at the cell equator prior to the
F ig. 6. Wild-type cells: A, phase-contrast-D A P I; B, immunofluorescence microscopy
using anti-tubulin antibody Y O L 1/34. D uring interphase groups of microtubules extend
between the cell ends (e.g. cells 1 and 2 ) . At mitosis the cytoplasmic microtubules are
replaced by a short intranuclear spindle (cell J ) , which elongates until the nuclei reach
the ends of the cell (cell 4 ). Reinitiation of the interphase array occurs prior to spindle
breakdown by the nucléation of microtubules at the cell equator (cells 4 and 5 ) . Bar,
10 fim .
Fission yeast cytoskeleton
237
F ig. 7. Phalloidin staining of cold-treated benA cells. These contain fewer, larger actin
dots and, in many cases, a conspicuous perinuclear ring. Bar, 10 fim.
deposition of Calcofluor-staining material. Both end growth and septation involve
the deposition of new cell wall macromolecules, albeit of different chemical com­
position (Bush et al. 1974; Horisberger & Rouvet-Vauthey, 1985). In fungi this
requires the mobilization of vesicles containing wall precursors to the growing region
(M cClure et al. 1968; Grove, 1978). Vesicles associated with the poles and septa of
S.pombe cells have been reported (OuleveyeZ al. 1970; Johnson etal. 1973), and this
suggests a possible explanation for the dot-like nature of actin staining. Although at
present there is no evidence for the association of actin with cell wall vesicles in
yeasts, vesicles coated with fine filaments of the approximate dimensions of F-actin
have been observed in a filamentous fungus (Hoch & Howard, 1980). T he role of
actin in wall deposition may not be finally clarified until cell wall vesicles are available
for biochemical study. T he localization of myosin may also provide important clues
(Watts et al. 1985) as will drugs that selectively interfere with cell wall morphology
(Miyata et al. 1985, 1986), and experiments to these ends are in progress. Although
the nature of the dot staining remains to be resolved, it is obviously a common feature
of fungal cells, having been seen in three ascomycetous yeasts (S a . cerevisiae,
Sa. uvarum and S. pombe) (Kilmartin & Adams, 1984; Adams & Pringle, 1984;
M arks & Hyams, 1985) and a filamentous basidiomycete (Utvmycesphaseoli) (Hoch
& Staples, 1983). T h is represents a broader sample than may initially be apparent, in
view of the evolutionary divergence between budding and fission yeasts (Huysmans
238
J . Marks, I. M. Hagan andy. S. Hyams
et al. 1983) and the fact that their mode of division is quite different. Cytokinesis in
budding yeast is assymmetric and involves a chitin ring, whereas fission yeasts divide
symmetrically and lack chitin (Bush et al. 1974).
The nature of actin staining as well as its position undergoes a marked change at
the initiation of mitosis. Dot staining at the cell ends is replaced by a filamentous
ring, which presumably occupies the thin layer of cytoplasm between the nucleus
and the cell membrane (Streiblova & Girbardt, 1980). Our finding that a ring of actin
anticipates the formation of the septum in S. pombe is consistent with earlier studies
of cytokinesis in fungi, which have provided ultrastructural evidence for the presence
of such a structure (Girbardt, 1979). The intimate relationship between the nucleus
and the actin ring is most clearly demonstrated by cdc 11, which can go through
multiple nuclear divisions at the restrictive temperature without undergoing cyto­
kinesis (Nurse et al. 1976). An actin ring forms in association with each daughter
nucleus at the start of mitosis; however, as the nuclei divide this spatial relationship
is lost. The fact that cdc 11 forms apparently normal actin rings and yet fails to lay
down any detectable septal material may well be of value in establishing the coupling
of these two events.
The relationship between the nucleus and the actin ring may also be addressed by
our findings with ben A. This mutant has the classic phenotype of a tubulin gene
mutation, namely, it is benomyl-resistant and cold-sensitive (Roy & Fantes, 1982).
Genetic studies have shown, however, that the ben A gene is distinct from the known
tubulin genes of S. pombe (P. Fantes, personal communication; see Yanagida et al.
1985). The exciting possibility therefore exists that ben A codes for a protein that
interacts with microtubules, i.e. a microtubule-associated protein (MAP). This is
supported by immunofluorescence staining of ben A cells with anti-tubulin anti­
body, which reveals an apparently normal array of cytoplasmic microtubules (our
unpublished results). The most conspicuous cytological feature of cold-treated ben A
cells is the presence of a perinuclear F-actin ring, reminiscent of the rings of
intermediate filaments that form around the nuclei of cultured cells treated with
colchicine (Goldman, 1971). A possible explanation for the origin of these structures
is that, like the /3-tubulin mutant nda3 (Hiraoka et al. 1984), cold-treated ben A cells
arrest at mitotic prophase. Since this is the time at which the equatorial actin ring
appears, the perinuclear rings in ben A may be the equatorial ring displaced from its
normal location. Although Roy & Fantes reported that ben A does not exhibit classical
cell cycle arrest, we have used a different temperature from that used in their study
and this can have a profound effect on the phenotype (Hiraoka et al. 1984). Clearly,
examination of the distribution of actin in known tubulin mutants of. S. pombe will be
of value in further establishing the nature of the ben A mutation as well as clarifying
the relationship between actin and tubulin in this organism.
Investigations of the role of microtubules in 5. pombe will also be considerably
aided by the introduction of the immunofluorescence techniques described here.
Spindle microtubules have previously been demonstrated in this way (Hiraoka
et al. 1984) but this is the first report of the visualization of cytoplasmic micro­
tubules. These are much more abundant than has previously been appreciated from
Fission yeast cytoskeleton
239
electron microscopy (Hereward, 1974; Streiblova & Girbardt, 1980; King & Hyams,
1982a,b\ Tanaka & Kanbe, 1986) and extend from one end of the cell to the
other. Cytoplasmic microtubules are clearly involved in the establishment and
maintenance of cell morphology since treatment of fission yeast with anti-micro­
tubule drugs results in a variety of morphological changes (Walker, 1982), as does the
disruption of microtubules by means of tubulin gene mutations (Toda et al. 1983;
Hiraoka et al. 1984). Whether microtubules are actively involved in the transport of
cell wall precursors or merely establish general cell polarity is at present unknown.
However, the fact that bud expansion in S. cerevisiae proceeds in the absence of
microtubules (Pringle et al. 1984) encourages us to favour the latter alternative.
Irrespective of this, our results have clearly shown that the distribution of F-actin in
fission yeast is controlled in a cell cycle site-specific manner. Whether understanding
of this regulation will emerge through the wider study of cell cycle controls (Hayles &
Nurse, 1986) or, more specifically, through the use of cloned actin genes and the
identification and characterization of the actin binding proteins remains to be seen.
We thank Professor T h. Wieland and D r John Kilmartin for the generous gifts of phalloidin and
tubulin antibody, respectively. This work was supported by Action Research for the Crippled
Child and the Science and Engineering Research Council.
REFERENCES
E. M. & P r i n g l e , J. R. (1984). Relationship of actin and tubulin distribution to bud
growth in wild-type and morphogenetic mutant Saccharomyces cerevisiae. J . Cell Biol. 98,
934-945.
B o n a t t i , S . , S i m i l i , M. & A b b o n d a n d o l o , A . (1972). Isolation of temperature sensitive mutants
of Schizosaccharomyces pom be.J. B a d . 109, 484-491.
B u r n s , R. G . (1973). 3H-Colchicine binding: Failure to detect any binding tosoluble proteins
from various lower organisms. E xpl Cell Res. 81, 285-292.
B u s h , D . A ., H o risberg er , M ., H orman , I. & W u r sc h , P. (1974). The wall structure of
Schizosaccharomyces pombe. jf. gen. Microbiol. 81, 199-206.
D a r k e n , M. A. (1961). Applications of fluorescent brighteners in biological techniques. Science
133, 1704-1705.
G irbard t , M. (1979). A microfilamentous septal belt (F SB ) during induction of cytokinesis in
Trametes versicolor (L . ex Fr). E xplM ycol. 3, 215-228.
G o l d m a n , R. D . (1971). The role of the three cytoplasmic fibres in BHK-21 cell motility.
1. Microtubules and the effect of colchicine. J . Cell Biol. 51, 752-762.
G rove , S. N. (1978). The cytology of hyphal tip growth. In The Filamentous Fungi, vol. 3 (ed.
J . E. Smith & D . R. Berry), pp. 28-50. New York: John Wiley & Sons.
H a y l e s , J . & N u r s e , P. (1986). Cell cycle regulation in yeast. J . Cell Sei. Suppl. 4, 155-170.
H e r e w a r d , F. V. (1974). Cytoplasmic microtubules in a yeast. Planta 117, 355-360.
H iraoka , Y ., T o da , T . & Y an a gida , M. (1984). The ND A3 gene of fission yeast encodes ßtubulin. A cold sensitive nda3 mutation reversibly blocks spindle formation and chromosome
movement in mitosis. Cell 39, 349-358.
H och , H . C. & H ow ard , R . J . (1980). Ultrastructure of freeze-substituted hyphae of the
basidiomycete Laetisaria arvalis. Protoplasma 103, 281-297.
H o c h , H . C. & S t a p l e s , R. C. (1983). Visualization of actin in situ by rhodamine-conjugated
phalloin in the fungus Uromyces phaseoli. Eur. J . Cell Biol. 32, 52-58.
H o r i s b e r g e r , M. & R o u v e t -V a u t h e y , M. (1985). Cell wall architecture of the fission yeast
Schizosaccharomyces pombe. Experientia 41, 748-750.
A dams A .
240
J . Marks, I. M. Hagan a n d j. S. Hyams
E ., D a m s , E ., V a n d e r b e r g h e , A. & d e W a t c h e r , R. (1983). The nucleotide
sequences of the 5 S rRNAs of four mushrooms and their use in studying the phylogenetic
position of basidiomycetes among the eukaryotes. Nucl. Acids Res. 11, 2871-2880.
J o h n s o n , B. F. (1965). Autoradiographic analysis of regional cell wall growth of yeasts. E xpl Cell
Res. 39, 613-624.
J o h n s o n , B. F ., Y o o , B. Y . & C a l l e j a , G. B. (1973). Cell division in yeasts: Movement of
organelles associated with cell plate growth of Schizosaccharomyces pombe. J . B a d . 115,
358-366.
H u y sm an s,
K ilm a r tin , J . V. & A dam s, A. E . M . (1984). Structural rearrangements of tubulin and actin
during the cell cycle of the yeast Saccharomyces. J . Cell Biol. 98, 922-933.
J . V., W r i g h t , B. & M i l s t e i n , C. (1982). Rat monoclonal antitubulin antibodies
derived by using a new nonsecreting rat cell line. J . Cell Biol. 93, 576-582.
K i n g , S. M. & H y a m s , J. S. (1982a). Synchronization of mitosis in a cdc mutant of
Schizosaccharomyces pombe released from temperature arrest. C a n .J. Microbiol. 28, 261-264.
K in g , S. M . & H yam s, J. S. (19826). Interdependence of cell cycle events in Schizosaccharomyces
pombe. Term inal phenotypes of cdc mutants arrested during D N A synthesis and cell division.
Protoplasma 110, 54 -6 2 .
M a r k s , J. & H y a m s , J. S. (1985). Localization of F-actin through the cell division cycle of
Schizosaccharomyces pombe. E u r.J . Cell Biol. 39, 27-32.
M c C lu r e , W. D ., P a r k , D . & R o b in so n , P. M . (1968). Apical organization in the somatic hyphae
of fungi, jf. gen. Microbiol. 50, 177-182.
M i t c h i s o n , J . M . & N u r s e , P. (1985). Growth in cell length in the fission yeast Schizo­
saccharomyces pombe. J . Cell Sci. 75, 357-376.
M i y a t a , M ., K a n b e , T . & T a n a k a , K . (1985). Morphological alterations in the fission yeast
Schizosaccharomyces pombe in the presence of aculeacin A: Spherical wall formation. J . gen.
Microbiol. 131, 611—621.
M i y a t a , M . , M i y a t a , H. & J o h n s o n , B. F. (1986). Assymetric location of the septum in
physiologically altered cells of the fission yeast Schizosaccharomyces pombe. J . gen. Microbiol.
132, 883-891.
N u r s e , P. (1981). Genetic analysis of the cell cycle. In Genetics as a Tool in Microbiology. SG M
Sym p. 31, pp. 291-315. Cambridge University Press.
N u r s e , P. (1985). Cell cycle control genes in yeast. Trends Genet. 2, 51-55.
N u r s e , P., T h u r i a u x , P. & N a s m y t h , K . (1976). Genetic control of the cell division cycle of the
fission yeast Schizosaccharomyces pombe. Molec. gen. Genet. 146, 167-178.
O u l e v e y , N ., D e s h u s s e s , J . & T u r i a n , G. (1970). Étude de la zone septale de Schizo­
saccharomyces pombe en division a ses etapes succesives. Protoplasma 70, 217-224.
P r in g l e , J . R ., C o lem a n , K ., A d a m s , A ., L il l ie , S ., H aarer , B ., J a cobs , C ., R o bin so n , J . &
E v a n s , C . (1984). Cellular morphogenesis in the yeast cell cycle. In Molecular Biology o f the
Cytoskeleton (ed. G . G . Borisy, D . W. Cleveland & D . B. M urphy). New Y ork: Cold Sprin g
K ilm a r tin ,
H arbor Laboratory Press.
J . R. & H a r t w e l l , L . H. (1981). The Saccharomyces cerevisiae cell cycle. In The
M olecular Biology o f the Yeast Saccharomyces (ed. J. N. Strathern, E. W. Jones & J . R. Broach).
New York: Cold Spring Harbor Laboratory Press.
R o y , D . & F a n t e s , P. A. (1982). Benomyl resistant mutants of Schizosaccharomyces pombe coldsensitive for mitosis. Curr. Genet. 6, 195-201.
STAEBELL, M. & S o l l , D . R. (1985). Temporal and spatial differences in cell wall expansion
during bud and mycelial formation in Candida albicans. J . gen. Microbiol. 131, 1467-1480.
S t r e i b l o v a , E. & G i r b a r d t , M. (1980). Microfilaments and cytoplasmic microtubules in cell
division cycle mutants of Schizosaccharomyces pombe. C an.J. Microbiol. 26, 250-254.
S t r e i b l o v a , E. & W o l f , A. (1972). Cell wall growth during the cell cycle of Schizosaccharomyces
pombe. Z . Allg. Mikrobiol. 12, 673—684.
T a n a k a , K . & K a n b e , T . (1986). Mitosis in fission yeast Schizosaccharomyces pombe as revealed
by freeze-substitution electron microscopy. J . Cell Sci. 80, 253-268.
P r in g l e ,
Fission yeast cytoskeleton
241
J . H ., N o v i c k , P. & B o t s t e i n , D . (1984). Genetics of the yeast cytoskeleton. In
M olecular Biology o f the Cytoskeleton (ed. G. G. Borisy, D. W . Cleveland & D. B . Murphy).
New York: Cold Spring Harbor Laboratory Press.
T o d a , T . , U m e s o n o , K . & H ir a o k a , A. (1983). Cold-sensitive nuclear division arrest mutants of
the fission yeast Schizosaccharomyces pom be.jf. molec. Biol. 168, 251-270.
W a l k e r , G . M. (1982). Cell cycle specificity of certain antimicrotubular drugs in Schizo­
saccharomyces pom be.jf. gen. Microbiol. 128, 61-71.
W a t t s , F . Z . , M i l l e r , D . M . & O r r , E . (1985). Identification of myosin heavy chain in
Saccharomyces cerevisiae. Nature, Lond. 316, 83-85.
W i e l a n d , T h . & G o v i n d a n , V. M . (1974). Phallotoxins bind to actins. F E B S L e tt. 46, 351-353.
W i l l i a m s o n , D . H. & F e n n e l l , D. J. (1975). The use of fluorescent D N A binding agent for
detecting and separating yeast mitochondrial DN A. Meth. Cell Biol. 12, 335-351.
W u l f , E . , D e b o b e n , A ., B a u t z , F . A ., F a u l t i s c h , H . & W i e l a n d , T h . (1979). Fluorescent
phallotoxin, a tool for the visualisation of cellular actin. Proc. natn. Acad. Sei. U .S A . 76,
4498-4502.
Y a n a g i d a , M . , H i r a o k a , Y ., U e m u r a , T ., M i y a k e , S. & H i r a n o , T . (1985). Control
mechanisms of chromosome movement in mitosis of fission yeast. In Yeast Cell Biology, UCLA
Sym p. vol. 33 (ed. J . Hicks). New York: Alan R. Liss.
T
h o m a s,