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Chapter2:Genesinpedigrees
«Celuiquinecomprendpas,etquiledit,estcelui
qui fait le plus évidemment preuve d’intelligence,
carilacomprisqu’iln’apascomprisetc’estcequi
est le plus difficile à comprendre ….» Albert
Jacquard.Lascienceàl’usagedesnon-scientifiques,
2001.
Haploidgametesareproducedbymeiosis
Parents transmit their genes to their offspring via
the gametes: the large immotile oocytes of the
mother and the miniature motile spermatozoa of
thefather.Bothtypesofgametesareproducedby
aspecializedtypeofcelldivision,knownasmeiosis,
thatproduceshaploidgametesfromdiploidcellsof
the germ line. Contrary to the somatic cells and
cells of the germ line, gametes only contain one
copy of the “genomic encyclopedia”. The number
of chromosomes found in haploid gametes is said
to be “n”, while diploid cells contain “2n”
chromosomes (corresponding to “n” pairs of
homologouschromosomesorhomologues).
Before entering meiosis, spermatogonia in the
testisandoogoniaintheovaryundergoreplication
oftheirDNAduringanSphasethatisverysimilar
tomitosis-precedingS-phasesexceptforitsslightly
increased duration. After S-phase, each
chromosome in the cell is present as two sister
chromatidsconnectedbycohesins.Gametogenesis
then proceeds with a very unusual type of cell
division: meiosis I. Rather than segregating one
sisterchromatidofeachhomologueineachofthe
two daughter cells, each cell will inherit the two
sister chromatids of one homologue for each
chromosome (ignoring recombination for the
moment). As mitosis, meiosis I is subdivided in a
prophase, metaphase, anaphase and telophase.
Prophase I is itself subdivided in four stages:
leptotene,zygotene,pachytene,anddiplotene.
Leptotene is characterized by the condensation of
thereplicatedchromosomesand-moststrikingly-
by the pairing of homologues. The condensed
sister chromatids of the paternal copy of
chromosome 1 will align themselves with the
homologoussisterchromatidsofthematernalcopy
of chromosome 1 along their entire length. The
same will happen for each chromosome pair,
forming as many bivalents. How are homologous
chromosomes finding each other in the nucleus?
Pairing appears to be mediated by the process of
“homologous recombination”. A specialized
protein (called Spo11 in yeast) introduces
staggered cuts in the DNA known as doublestranded breaks. On either side of the break,
exonucleasesthengeneratelong,single-stranded3’
overhangs. Proteins of the RecA family (called
Rad51 in yeast) cover the single-strands and
catalyzethesearchforhomologousDNAsequences
by strand-invasion, i.e. the formation of threestranded structures comprising a region of
heteroduplex DNA and corresponding displaced
strand. Branch migration extends the
heteroduplex region, while formation of a doubleHolliday junction (involving DNA synthesis and
ligation) seals the local connection between
homologues. Pairing involves tens to hundreds of
exchangesofthiskindperchromosomepair.
During zygotene, the alignment of homologous
chromosomes is stabilized by formation of
synaptonemal complexes: ladder-like structures
comprising a pair of axial cores bridged by
transversefilaments.Pachytenemarkscompletion
ofsynapsis.Double-Hollidayjunctionsareresolved
by formation of DNA strand cuts and re-ligation,
which can happen in two ways. By far the most
common pathway leaves a local segment of
heteroduplex as sole trace of the recombination
process. Approximately one double-Holliday
junction per chromosome arm is resolved using a
distinct pathway that creates a crossing-over
between homologous sister chromatids. Crossingovers result in pairs of double-stranded-helixes
with distinct parental origin on either side of the
crossing over (maternal-CO-paternal and paternalCO-maternal).
Paternalandmaternalsequencesmaydifferwithin
the heteroduplex regions as a result of Single
Nucleotide Polymorphisms (or SNPs) that occur in
all populations (see hereafter). This may cause a
mismatchthatlocallydeformtheDNAdoublehelix
andmaytriggerDNArepair.Inthisprocess,known
as “gene conversion”, the sequence originating
fromoneparentis“converted”tothatoftheother.
Diplotene is characterized by the disassembly of
the synaptonemal complexes, and further
chromosomal condensation. Paired homologs
remainconnectedatchiasmata(singularchiasma),
the sites where crossing-overs occurred. The end
ofprophaseIisreferredtoasdiakinesis.
The remainder of meiosis I (i.e. metaphase I,
anaphaseI,telophaseI)ishighlysimilartomitosis,
exceptthat(i)thekinetochoresofsisterchromatids
bind to microtubules originating from the same
pole (rather than opposite poles as in mitosis), (ii)
chiasmata connect homologues (rather than
cohesins connecting sister chromatids during
mitosis)priortosegregation,enablingalignmentof
thechromosomesonthemetaphaseplate,and(iii)
cohesins at the kinetochores are protected from
Chapter2:GenesinpedigreesPage1/14
the action of separase by proteins called
shugoshins.
Meiosis II is in nearly all respect equivalent to a
mitoticdivision.
The diploid germ cells that undergo meiosis are
calledoogonia(singularoogonium)infemales,and
spermatogonia(singularspermatogonium)inmales.
Both types of cells derive from a population of
primordial germ cells (PGC) that migrate to the
developinggonads.
Inafemalefetus,PGCwilldividemitoticallyinthe
6
ovarytoyield∼5x10 oogonia.Itisestimatedthat
∼22celldivisionsseparateeachoogoniumfromthe
zygote from which they derive. The fetal oogonia
thenentermeiosisI,whichis–however-arrested
at the diplotene stage of prophase I. The
corresponding germ cells are now called primary
oocytes. Only at puberty, stimulated by
gonadotropins secreted by the pituitary gland at
eachmenstrualcycle,willoneorasmallnumberof
primary oocytes complete meiosis I. The
corresponding cytokinesis is strikingly asymmetric,
yielding a large secondary oocyte inheriting
virtuallyallthecytoplasmiccontentattheexpense
oftheminutefirstpolarbody.Secondaryoocytes
are released from the ovary by the process of
ovulation, yet remain arrested in metaphase II.
Only if fertilized by a sperm cell, will secondary
oocytes complete meiosis II, releasing a fertilized
matureegg(orzygote)andsecondpolarbody.
In a male fetus, PGC migrate in the developing
9
testis,wheretheymultiplymitoticallytoyield∼10 spermatogonia lining the basal membrane of the
seminiferoustubulesatthetimeofpuberty.Each
one of these is separated by ∼30 mitotic divisions
fromthezygotefromwhichtheyderive.Starting
atpuberty,spermatogoniawillengageevery∼two
weeksinanasymmetricmitoticdivision,yielding(i)
astemcell-likespermatogoniumtosupportfuture
spermatogenesis,
and
(ii)
a
maturing
spermatogonium that commits to completing
spermatogenesis, implying three more mitotic
divisions (yielding eight primary spermatocytes),
followed by meiosis I (yielding sixteen secondary
spermatocytes), and meiosis II (yielding 32
spermatids that will terminally differentiate in as
manymaturespermcells).The32cellsthatderive
from a maturing spermatogonium remain
connected by cytoplasmic bridges, forming a
syncytium until the very final stages of sperm
maturation. Although genetically haploid,
secondary spermatocytes and spermatids are in
effect functionally diploid. As a result, although
halvethesecondaryspermatocytesandspermatids
are X-bearing and the other halve Y-bearing, all of
them will contain all the gene products derived
fromtheXandYchromosome,ofwhichsomeare
essential to complete spermatogenesis. Contrary
totheegg,whichisseparatedfromthezygotebya
constantnumberof∼23celldivisions,thenumber
ofcelldivisionsleadingfromthezygotetoasperm
cell equals ∼34 + (26 x number of years after
puberty).
Sexualreproduction,involvingthefusionofhaploid
gametescontributedbyanimalsofoppositesex,is
an elaborate and costly process. Yet is utilized by
the vast majority of plants and animals. It
thereforemustconferaselectiveadvantage,which
nature–however-remainspoorlyunderstood.As
a result of the independent segregation of the
maternal and paternal homologues for distinct
chromosomes (Mendel’s second law), as well as
theirreshufflingbytheprocessofcrossing-over,no
two gametes produced by an individual are
identical. For species producing many offspring,
this might ensure that at least some offspring
wouldbewelladaptedtoachangingenvironment.
Sexual reproduction promotes the combination of
multiple favorable mutation having occurred in
independent lineages, while asexual reproduction
requires such favorable mutations to occur
sequentially in the same lineage. Sexual
reproductionhasalsobeenproposedasameansto
eliminate deleterious mutations, which would
otherwiseaccumulateinthepopulation.
In most sexually reproducing organisms, haploid
cellsdonotmultiply.Onlythediploidsomaticand
germline cells multiply mitotically. In some
primitive plants, diploid and haploid cells
proliferate.Insomespeciesofyeast,onlyhaploid
cells proliferate, while the diploid zygote
immediately engages in meiosis to produce new
haploidcells.
Mendel’slaws
When asked what genetics is all about, students
typically respond with “the science of heredity” or
“the study of genes and their action”. Indeed as
apparentfromthetableofcontentsofthiscourse,
genetics studies the behavior and mode of action
of genes in cells (molecular genetics), pedigrees
(factorial and quantitative genetics), and
populations(populationandevolutionarygenetics).
But, equally important, genetics also refers to a
wayofdoingscience,anexperimentalapproachto
unravel complex biological phenomena. If Gregor
Mendel is as much revered by geneticists as he is
today, it is because he was probably the first to
applythismethod,andthisinanexemplaryfashion.
Providing extra appeal to geneticists, he applied it
tothestudyofheredity.
Mendel was born Johann in the now Czech
Republic in 1822. He was raised on a farm, and
wenttoUniversitybeforeenteringpriesthoodand
the Augustinian Abbey of St Thomas in Brno. He
performed research in plant and animal breeding
but also astronomy and meteorology. His now
famousresultsonplanthybridizationswentlargely
Chapter2:GenesinpedigreesPage2/14
unnoticed for ∼35 years (including by Darwin)
beforebeingrediscoveredaround1900.Heisnow
generally considered to be the father of modern
genetics.
Mendel’s most famous experiments exemplify the
key features of a well designed and executed
experiment, and these are still highly relevant in
thewayscienceisconductedtoday.Hefirstsetthe
stage by carefully designing an experimental
system. In his case he used peas, which he could
multiply in a controlled fashion. As peas are
hermaphrodites (the same plant produces both
male and female gametes), he could both “self”
plants. But he could also perform outcrossing at
will, by excising the undeveloped male pollenproducing structures of a plant, and expose the
femalestructurestomaturepollenfromadifferent
plant using a pencil. Repeated selfing of plants
with unique attributes (called phenotypes) let to
pure lines, i.e. plants that would yield uniform
offspring upon self-fertilization. By doing so
Mendel developed pure “parental” lines (F0) that
differed for one or more of seven “binary”
phenotypes, including seed shape (round or
wrinkled)andcolor(yelloworgreen).
Inhisfirstexperiments,Mendelcrossedpure-lines
differingforonesuchbinaryphenotype,producing
aso-calledF1generation.HethenselfedtheF1’s,
producing the F2 generation. The outcome of
thesecrossessharedcommonfeatures:(i)theF1’s
exhibited the phenotype of one of the crossed
parentallines,(ii)¾oftheF2’sexhibitedthesame
phenotype as the F1 generation, while the
remaining¼re-expressedthephenotypeoftheF0
lineunseenintheF1generation.
Mendelthenproposedamodel(Mendel’sfirstlaw
of segregation) that would account for his
observations. He posited that each binary trait is
determined by a hereditary particle (now referred
toas“gene”)(f.i.thegenedeterminingseedshape),
that comes in two forms or alleles (f.i. the round
alleleandthewrinkledallele).Individualshavetwo
copiesofeachgene:oneinheritedfromthefather
the other from the mother. Individuals inheriting
the same allele from both parents are said to be
homozygous or have homozygous genotype (f.i.
wrinkled-wrinkledorround-round)andexpressthe
correspondingphenotype(respectively,wrinkledor
round). Individuals that inherit a different copy
from each parent are heterozygous (f.i. genotype
round-wrinkled). As one allele is dominant (and
the other recessive), heterozygous individuals
express the same phenotype as the homozygotes
forthedominantallele.Gametescontainonlyone
copy of each gene. Gametes from heterozygous
parents have equal probability to harbor either of
theallelesoftheparent.Referringtothedominant
round allele as W and to the recessive wrinkled
allele as w, Mendel’s model applied to crosses
between pure lines of peas with round and
wrinkledseedscanbesummarizedasfollows:
F0:WWxww
F1:Ww
F2
Male
gametes
W(1/2)
w(1/2)
Femalegametes
W(1/2)
w(1/2)
WW(1/4)
Ww(1/4)
Ww(1/4)
Ww(1/4)
Thefinalbutcrucialstagewastotesttheproposed
model. The model indeed makes a number of
predictions, including about the expected
phenotypicproportionsonselfingF2,i.e.intheF3
generation. According to the model, wrinkled F2
are all homozygous ww and should therefore
produce all wrinkled offspring when selfed. More
interestingly,themodelpredictsthat1/3ofround
F2’s should be homozygous WW and hence
produceallroundoffspringonlywhenselfed,while
2/3ofroundF2’sshouldbeheterozygousWwand
hence behave as the F2’s when selfed, i.e. yield ¾
roundand¼wrinkledF3’s.Andthisisindeedwhat
Mendelobservedwhenhetestedhishypothesis.
Another important observation made by Mendel,
wasthe“equivalenceofreciprocalcrosses”,i.e.the
outcome was independent of the sex-byphenotype combination in the F0 generation. In
theexampleoftheseedshape,thismeansthatthe
outcomeofthecrosseswasthesamewhetherthe
wrinkledF0parentwasmaleorfemale.
Inasecondseriesofexperiments,Mendelcrossed
pure-lines that differed for two binary phenotypes
(f.i.round&yellowcrossedwithwrinkled&green).
In agreement with the previous experiments, the
F1 generation only exhibited the dominant
phenotypes as defined before (f.i. round and
yellow). When selfing the corresponding F1’s he
always obtained the four possible phenotypic
combinations in the following proportions in the
F2: 9/16 (dominant-dominant; f.i. round-yellow),
3/16 (dominant-recessive; f.i. round-green), 3/16
(recessive-dominant;f.i.wrinkled-yellow),and1/16
(recessive-recessive;f.i.wrinkled-green).
The model he proposed to account for these
observations, was that alleles of different genes
segregateindependentlyinthegametes(Mendel’s
secondlawofindependentsegregation).Thus,ifa
individual is double heterozygote, he will produce
four types of gametes in equal proportions.
Applied to crosses between wrinkled-yellow and
round-green peas, and referring to the dominant
yellowallelesasGandtotherecessivegreenallele
asg,hismodelcanbesummarizedasfollows:
F0:WWGGxwwgg
Chapter2:GenesinpedigreesPage3/14
F1:WwGg
MaleGametes
F2
Femalegametes
WG
(1/4)
Wg
(1/4)
wG
(1/4)
wg
(1/4)
WG
(1/4)
WWGG
(1/16)
WWGg
(1/16)
WwGG
(1/16)
WwGg
(1/16)
Wg
(1/4)
WWGg
(1/16)
WWgg
(1/16)
WwGg
(1/16)
Wwgg
(1/16)
wG
(1/4)
WwGG
(1/16)
WwGg
(1/16)
wwGG
(1/16)
wwGg
(1/16)
wg
(1/4)
WwGg
(1/16)
Wwgg
(1/16)
wwGg
(1/16)
wwgg
(1/16)
Asbefore,thisextendedmodelmakesanumberof
predictions which Mendel’s verified. One of the
checks he carried out was to cross the F1’s with a
tester. A tester is a pure line that is homozygous
fortherecessiveallelesatallexaminedgenes.Asa
consequence, the phenotype exhibited by the
offspring directly reflects the genotype of the
gametestransmittedbytheF1.Aspredictedbyhis
model, this test-cross yielded the four possible
phenotypiccombinationsinequalproportions.
By performing an apparently simple “bean bag”
experiment, and just on the basis of observed
phenotypicproportions,Mendelproposedamodel
that accurately described essential features of
heredity and meiosis. Make sure to identify the
featuresofmeiosisthatunderlieMendel’sfirstlaw
of segregation and second law of independent
assortment.
Subsequently, other scientists tested Mendel’s
models,includinginnewmodelorganismssuchas
the fruitfly D. melanogaster. In a few (lucky)
instances, Mendel’s predictions proved to be
inaccurate, and predictive models had to be
amended accordingly. As an example, violation of
the equivalence of reciprocal crosses let to the
discovery of the sex chromosomes, and of
Mendel’s second law to the discovery of genetic
linkage.
(BoxNomenclature)
Geneticlinkage
Mendel’ssecondlawstatesthatanindividualthat
is heterozygous for two genes (f.i. AaBb) produces
fourtypesofgametesinequalproportions:AB,Ab,
aB and ab. However, geneticist discovered in the
early20-thcenturythatMendel’ssecondlawdidn’t
apply to all pairs of genes. For such pairs, two
types of gametes were significantly overrepresented at the expense of the two other
classes. The over-represented types corresponded
to the “parental” gametes, i.e. gametes with
genotype identical to the ones inherited by the
double heterozygous individual from its parents.
AssumethattheparentswererespectivelyofAABB
and aabb genotype, the informative AaBb
individual would preferentially transmit “parental”
AB and ab gametes, at the expense of so-called
“recombinant” Ab and aB gametes. If – on the
otherhand–theparentswererespectivelyofAAbb
and aaBB genotype, the AaBb offspring would
preferentially transmit “parental” Ab and aB
gametes at the expense of “recombinant” AB and
ab gametes. Determining which gametes
transmitted by a double heterozygous individual
are parental and which are recombinant is called
“determining the linkage phase”. One way to
deducethelinkagephaseisfromtheanalysisofthe
parental genotypes – as shown above. When the
linkage phase of a double heterozygous individual
is known, this can be indicated by writing the
genotype as follows: AB/ab. Pairs of genes
characterized by an excess of parental and lack of
recombinant gametes are said to be genetically
linkedorexhibitgeneticlinkage.Theproportionof
recombinant gametes was rapidly shown to vary
for different pairs of linked genes, and to be
independent of the phase of the informative
individual (i.e. it would be the same for an AB/ab
andanAb/aBindividual).
It is now well established that linkage occurs
between genes that reside on the same
chromosome. Such genes are said to be syntenic.
ThefactthatMendeldidnotobservelinkageisdue
tothefactthat-bychance–thesevengenesthat
he studied were non-syntenic. In the case of
syntenic genes, recombinant gametes derive from
meiosesinwhichacrossing-overoccurredbetween
the two considered genes. Such event is more
likelyifthetwosyntenicgenesarefarapartthanif
theyareclosetoeachother.Thedegreeoflinkage,
reflected by the proportion of recombinant
gametes (or recombination frequency - RF), can
therefore be viewed as a measure of the distance
separatingsyntenicgenes.
Itwasquicklyrecognizedthatsyntenicgenescould
be positioned relative to each other based on
observed RF, i.e. that one could build genome
mapsbylinkageanalysis.Imaginethattest-crossing
an ABC/abc individual yields the following set of
progeny:
Phenotype
Number
Proportion
ABC
350
.350
abc
360
.360
ABc
6
.006
abC
4
.004
Abc
92
.092
aBC
98
.098
AbC
43
.043
aBc
47
.047
Total
1,000
1
Rememberingthatinatestcrossthephenotypesof
the offspring directly reflect the genotype of the
gamete transmitted by the triple heterozygote
parent, the observed RF between A and B equals
(92+98+43+47)/1000or0.28.Achi-squared-based
Chapter2:GenesinpedigreesPage4/14
goodness-of-fit test indicates that the observed
gametic frequencies differ very significantly from
those expected under Mendel’s second law, with
an excess of parental over recombinant gametes
typicaloflinkedgenes:
O(bs)
E(xp)
2
(O-E) /E
AB
ab
Ab
aB
350+6
360+4
92+43
98+47
250
250
250
250
44.9
52.0
52.9
44.1
2
Chi p
193.9
-6
<10 TheRFbetweenAandCequals(6+4+92+98)/1000
or 0.20, which also departs very significantly
-6
(pgoodness-of-fit<10 ) from Mendelian expectations
indicatingthatAisalsolinkedtoC.Alongthesame
lines, the RF between B and C equals
-6
(6+4+43+47)/1000or0.10(pgoodness-of-fit<10 ).
ThedatathusindicatethatA,BandCaresyntenic,
while analysis of pairwise RF (AB: 0.28; AC: 0.20;
BC:0.10)indicatethatthecorrectgeneorder(out
ofthethreepossibleorders)isA-C-B,i.e.thatgene
CislocatedbetweengenesAandB.
Note that we could have reached the same
conclusions even without a priori (f.i. based on
knowledge of the genotype of its parents)
information about the linkage phase of the
informative triple heterozygous parent. Indeed,
the two most frequently transmitted gametes
(ABC: 350/1000 and abc: 360/1000) indirectly
determinethephase,whilethetwoleastcommon
gametes (ABc: 6/1000 and abC: 4/1000) identify
the double recombinants thereby unambiguously
establishingorder.
It is worthwhile noting that the distances in RFunits between A-C (0.20) and C-B (0.10) add up to
more than the distance between A-B (0.28). This
non-additivityisofcoursenotadesirableproperty
for a distance measure. Closer examination
indicatesthatthelowerthanexpectedRFbetween
A and B is due to the fact that 10 gametes have
been counted as parentals, while being in fact
double-recombinants (having recombined in both
theA-CandC-Bintervals).Thefactthatwehave
informationaboutCbetweenAandBallowedusto
detect recombinational events, which we would
havemissedotherwise.If,ratherthanconsidering
these 10 gametes as parental, we consider that
they jointly contribute 20 recombination events,
the corrected RF between A and B becomes 0.30
which is exactly equal to 0.20 (A-C) + 0.10 (C-B).
Thisindicatesthatadistancemeasurebasedonthe
numberofCOintheconsideredintervalwouldbe
betterthanonebasedontheobservedproportion
of parental gametes (which in fact includes
gameteswithzerobutalsowithanevennumberof
COintheintervalofinterest).Asamatteroffact,
even if one can’t directly observe all CO that may
have occurred between two loci from genotype
data, one can fairly accurately estimate their
frequencyfromtheobservedRF.Themathematical
relationship between the observed RF and the
estimated CO frequency (COF) is called a mapping
function.
ThesimplestmappingfunctionisHaldane’s.Itcan
bederivedasfollows.Assumetwosyntenicloci,A
andB.Assumethemanymeiosesoccurringinthe
gonadsofadoubleheterozygousAB/abindividual.
A proportion of meiosis will be characterized by 0
chiasmata(CH)betweenAandB,aproportionby1
CH, a proportion by 2 CH, a proportion by 3 CH,
etc ... We will assume that these proportions are
accuratelydescribedbyaPoissondistribution,i.e.
p(x;m) =
e−m m x
x!
inwhichmistheaveragenumberofCHoccurring
betweenAandBacrossallmeioses.Notethatthe
-m
proportionofmeiosiswith0CHhenceequalse .
By definition gametes issued from meiosis with 0
CH between A and B can only be parental.
Gametes issued from meioses with one CH
between A and B, have 50% chance to be
recombinantand50%chancetobeparental.This
is due to the fact that CH occur at prophase I,
involving two of the four present chromatids.
Paradoxically, gametes issued from meiosis with
two or more CH between A and B, likewise have
50%chancetoberecombinantand50%chanceto
beparental.Thiscanbeseenformeiosiswithtwo
CHasfollows.Letuslabelthesisterchromatidson
thepaternalhomologue1and2,andthoseonthe
maternalhomologue3and4.EachCHcaninvolve
four possible chromatid combinations (1+3; 1+4;
2+3; 2+4). Meioses with two CH can thus
correspondto16possiblechromaticcombinations,
which are a priori equally probable. For four of
these,thetwoCHinvolvethesamechromatidpair
(f.i. 1+3 & 1+3). All gametes derived from such
meiosisareparental.Foreight,thetwoCHinvolve
three chromatids (f.i. 1+3 & 1+4). Halve the
gametes derived from such a meiosis will be
parental, the other halve recombinant. For the
remainingfour,thetwoCHjointlyinvolvethefour
chromatids (f.i. 1+3 & 2+4). All gametes derived
from such meiosis will be recombinant. Taken
together, half the gametes derived from meioses
withtwoCHbetweenAandBwillbeparental,the
otherhalfrecombinant.Thesameconclusioncan
be reached for meioses with three CH between A
and B (64 possible chromatid combinations), four
CH,etc.
Therefore, the expected RF between loci A and B
equals:
1
RFA−B = (1− e−m ) 2
where (1− e−m ) corresponds to the proportion of
meiosiswithatleastoneCHbetweenAandB.
Chapter2:GenesinpedigreesPage5/14
GeneticistsprefertousetheaveragenumberofCO
(COF)pergameteasdistancemeasure,ratherthan
the average number of CH per meiosis (m), but
thesetwoaresimplyrelatedas2xCOF=m.Hence,
1
RF = (1− e−2*COF ) 2
and
1
COF = − ln(1− 2RF) 2
COFisexpressedinmorgansorcentimorgans(cM).
One cM corresponds to one CO per hundred
gametes.ThusRFbetweenlociaremeasuredfrom
thegenotypedata,andsubsequentlyconvertedto
cM (i.e. an additive measure of genetic distance)
usingmappingfunctions.
Haldane’smappingfunctionassumesthatCHoccur
at random along a bivalent and have no influence
on each other. This does not correspond to the
reality.First,therearevirtuallynonullichiasmatic
meiosis, as chiasmata play a crucial mechanistic
role in linking homologous chromosomes during
meiosis I. This observation is sometimes referred
to as the “obliged crossing-over” rule. Secondly,
multiple CO tend to occur farther apart than
expectedbychancealone.Thelatterphenomenon
is called positive interference, i.e. the fact that a
firstCOdiminishestheprobabilityofasecondone
in its immediate vicinity. The degree of
interferencebetweenintervals(sayA-BvsB-C)can
bemeasuredona0to1scaleas:
I = 1−
DCOOBS
DCOEXP
where DCOOBS/DCOEXP is the ratio between the
observed and expected numbers of doublecrossovers, also known as coefficient of
coincidence. In the previous example, the
coefficientofcoincidencewas((4+6)/1000)/0.1*0.2
=0.5,correspondingtoaninterferenceof0.5.
Mappingfunctionshavebeenmodifiedinorderto
accountforthestronginterferencethatisobserved
for closely linked intervals in many organisms,
including mammals. The most commonly used
mappingfunctionisKosambi’sone.Itassumesthat
thelevelofinterferencediminisheswithincreasing
distance.
Centimorgans, as defined above, are a convenient
additive measure of genetic distance. But what is
the relationship between centimorgans and the
physical measure of genetic distance, i.e. the base
pairs? At small scale the relationship looks like a
staircase: regions with very little recombination
separated by “recombination hotspots”. The
majorityofCOindeedoccurwithin∼25,0001-2Kblong recombination hotspots containing short
recombination-triggering motifs. In human, the
consensus sequence of the most common
recombination-triggering
motif
is:
CCNCCNTNNCCNC.Atlargerscale,oneobservesan
increase in the recombination towards the
telomeres where recombination hotspots tend to
concentrate, and a comparative decrease in
recombination around the centromeres, which are
recombinationcold-spots.
Recombinationvariesbetweenindividuals,withsex
having a major effect: the recombination rate is
typically higher in the homogametic than in the
heterogametic sex. But even within sex,
recombination is variable and this is in part
geneticallydetermined.Ofnote,inwomenglobal
recombinationrate(i.e.theaveragenumberofCO
in a gamete) is positively correlated with
reproductive success. It has also been
hypothesized that domestication has lead to an
increaseinrecombination.
Linkageplaysacentralroleingenetics.Assoonas
recombinant DNA technology permitted the
development of numerous polymorphic DNA
markers (f.i. microsatellites and SNPs) for human
and other species of interest, dense linkage maps
comprising thousands of markers were generated.
These played an essential role as a first step
towards assembling the complete genomic
sequenceofthesespecies,and–mostimportantly
- they permitted the mapping of thousands of
genes underlying inherited defects and other
monogenictraits,aswellhundredsofQuantitative
Trait Loci influencing complex diseases and
quantitativetraitsinhuman,animalsandplants(cfr.
Chapter...).
Chromosomalsexdetermination
Inmanyspecies,thekaryotypediffersbetweenthe
sexes. In placental mammals for instance, while
females have n pairs of matched chromosomes,
males have n-1 pairs of matched chromosomes
(referredtoasautosomes),plusonepaircomposed
of two distinct chromosomes. This unique
chromosomal pair corresponds to the sex
chromosomes or gonosomes. In mammals,
females are the homogametic sex and their n-th
matchedpairofchromosomescorrespondstotwo
identical gonosomes referred to as “X”. The males
are the heterogametic sex and their n-th
unmatchedgonosomalpaircorrespondstoone“X”
(identical to the sex chromosomes of the females)
and “Y” chromosome. The Y chromosome is
typicallyconsiderablysmallerthanitsXcounterpart.
As females are XX, all oocytes carry a X
chromosome.Ontheotherhand,halvethesperm
cells carry a X and will give a female offspring on
fertilization,whiletheotherhalvecarryaYandwill
givemaleoffspringonfertilization.
Chapter2:GenesinpedigreesPage6/14
120 million years ago, i.e. after the separation of
the lineage that would give the metatheria (i.e.
marsupials) and eutheria (i.e. placental mammals)
from that of the prototheria (i.e. monotremes), a
mutation in the SOX3 gene (encoding a
transcription factor) located on a then regular
autosome,generatedanalleleconferringmaleness,
known today as the SRY gene. The corresponding
chromosomes carrying the original SOX3 allele
would become the X chromosome, while those
carrying the newly born male-determining SRY
genewouldbecometheYchromosome.
How the size of the Y chromosome has been
reduced during 120 million years of evolution, is a
very interesting biological question. It is believed
that mutations in genes in the vicinity of SRY
created alleles that were beneficial for male
fertility but deleterious for female fertility. Such
newalleleswouldfarebestinthepopulation(and
hence favored by natural selection) if they were
confinedtomales.Onewaytoachievethiswould
be to block recombination between such malebeneficial haplotypes on the proto-Y chromosome
andtheirXcounterpart.Aswewillseelaterinthis
chapter, chromosomal inversions are one such
recombination-blocking mechanism. It is now
assumed that successive inversions have isolated
increasing portions of the Y chromosome from its
formerXpartner,reducingtheportionalongwhich
the ancestral partners are still able to recombine
(whichisessentialforpropersegregationofthesex
chromosomes during male meiosis) to the
nowadays very short “pseudo-autosomal region”
(PAR).TheprogressivedegradationoftheY-specific
region of the Y chromosome is thought to be a
directconsequenceofthefactthat,contrarytothe
Xchromosome,theYneverproductivelyengagesin
recombination with a homologous partner (as YY
individuals don’t occur) which seem essential for
repair, and probably also of the fact that
deleterious mutations on the Y are systematically
shielded from purifying selection by the X
chromosome.
As the Y chromosome progressively lost an
increasing proportion of its original gene
complement, the ratio between autosomal gene
copies(diploid)andgonosomalgenecopies(diploid
in females, haploid in males) started to differ
betweenmalesandfemales,causingaproblemfor
molecular systems requiring finely tuned
componentdosage.Counteringthisdosageissueis
thought to underlie the selective pressure that let
to the evolution of “dosage compensation”
mechanisms to achieve proper and equal
autosomaltogonosomalbalanceinbothsexes.In
mammals, dosage compensation involves two
components.
The first is “X-inactivation” in females (or
“Lyonisation” in honor of Mary Lyon who
discoveredthismechanism).Althoughfemalecells
are diploid XX, for most genes on the X-specific
portion of the X chromosome, only one copy is
active the other one being epigenetically silenced.
In eutherian females, cells of the epiblast chose
oneoftheXchromosomestoremainactive,while
theotheroneissilenced.Thischoiceistoalarge
extent random, so that some cells will silence the
paternal X while neighboring cells may have
selected the maternal X for silencing. However,
once the decision is made in an embryonic
precursor cells, her choice is epigenetically
transmittedtotheentirecloneofdescendingcells.
Alleutherianfemalesarethusinessence“mosaics”
(i.e. organisms comprising cells that derive from
the same zygote yet differing in their genetic
complement),andfunctionallyhaploidformanyof
the genes on the X chromosome. This functional
mosaicism of females is directly apparent in
tortoiseshell cats. These always female cats are
heterozygous
“B(lack)/O(range)”
for
a
pigmentation gene on the X. The black sectors
correspond to cell populations having inactivated
theXcarryingtheOallele,whiletheorangesectors
correspond to cell populations having inactivated
the X carrying the B allele. The additional white
spots, characterizing so-called calico cats, are due
tothe“Piebald”alleleatadistinctautosomallocus.
OneofthekeyplayersoftheX-inactivationprocess
is the XIST gene, which is the only gene that is
expressed from the inactive X and not from the
activeX.ThecorrespondingXISTtranscriptisone
ofthebeststudiedlongnon-codingRNAgenes.It
coverstheXchromosomefromwhichitisderived,
thereby silencing it by a still rather obscure
mechanism. In addition to its XIST coat, the
inactive X chromosome is characterized by
epigenetically inherited DNA methylation and
chromatin silencing marks. The inactive X
chromosomeadoptsacondensedheterochromatic
conformation that is microscopically visible at the
nuclearperipheryasaso-calledBarrbody.
X inactivation equalizes expression levels from the
X chromosome in males and females. Proper
gonosome-to-autosomebalanceisachievedbythe
second mechanism involved in dosage
compensation,i.e.thedoublingofexpressionlevels
of all genes on the X when compared to genes on
autosomes. The mechanisms underlying this
generalized X-specific boost of expression remains
unknown.
How does the SRY gene cause masculinization of
thedevelopingfetus?Inamalefetus,theSRYgene
is activated in somatic cells of the developing
gonad, which consequently differentiate in Sertoli
cells. Sertoli cells secrete signals which (i) cause
the incoming germ cells to differentiate along the
sperm rather than egg pathway, (ii) cause
neighboring cells to differentiate into Leydig cells
whichsecretethesex-hormonetestosteronewhich
will cause masculinization of several organs, and
Chapter2:GenesinpedigreesPage7/14
(iii) block the development of the Müllerian
anlagen(anti-Müllerianhormone).Intheabsence
of SRY (female fetus), the somatic cells in the
developing gonad differentiate in follicle cells, (i)
letting incoming germ cells develop along the egg
pathway, and (ii) stimulating neighboring cells to
differentiateintoestrogen-secretingthecacells.
Inmammals,themalesexisheterogameticandthe
female sex homogametic. In some organisms,
however, the situation is opposite: males are
homogametic and females heterogametic. The
gonosomesarethenreferredtoasZandW,males
beingZZandfemalesZW.Thissituationappliesin
particulartobirds.AsintheXYsystem,theWhas
a tendency to degenerate over time for what are
assumedtobesimilarreasons.Intheseorganisms,
thesex-determininggeneisnotSRY.
(Cell-autonomoussexdifferentiationinbirds)
(discoveryofthesexchromosomes)
Geneticpolymorphismsandhaplotypeblocks.The
descriptionofmeiosisandgametogenesisinformed
us about the mechanisms that govern the
transmission of chromosomes and genes from
parents to offspring. It didn’t account for the
differencesthatareobservedbetweenindividuals,
andwhysomeofthesetendto“runinfamilies”.
Heritable, distinctive features find their origin in
the “genetic polymorphism” that characterizes
outbred populations (in contrast to inbred strains
in which all individuals are genetically identical or
monomorphic). In human, the genome of two
randomlyselectedgametes(forinstancethesperm
cellandoocytethatgavebirthtoyou),differevery
6
∼1,000 nucleotides, for a total of ∼3x10 differences. The number of difference per
nucleotide between two randomly selected
genomes(oraverageheterozygositypernucleotide
siteforarandomlyselectedindividual)iscalledthe
nucleotidediversity(π).Inmostdomesticanimals,
the nucleotide diversity is slightly lower than in
human,oftheorderof∼1/2,000to∼1/3,000.This
difference is related to the population size (see
6
Chapter III). The ∼3x10 differences between two
randomly selected genomes correspond mainly to
SingleNucleotidePolymorphismsor“SNPs”.These
are genome positions for which more than one
alleleisobservedinthepopulation.MostSNPsare
saidtobebiallelic:onlytwoallelesareobservedat
appreciable frequency in the population. Biallelic
SNPs are often characterized by their Minor Allele
Frequencyor“MAF”,i.e.thepopulationfrequency
of the rarest allele, which ranges from 0 to 0.5.
MostSNPscorrespondtonucleotidesubstitutions,
andthesecaneitherbetransitionsortransversions.
ItisimportantwhendescribingSNPstoagreeona
referencestrand,whethertheCrickortheWatson
strand. Having done this, transitions are either a
cytosine<->thymine
or
adenine<->guanine
substitution.Transversionscorrespondtothefour
possible
purine<->pyrimidine
substitutions.
Although there are twice as many possible
transversions as transitions, transitions are more
thantwicemorecommonthantransversions.
If one were to randomly sample a second pair of
human genomes, their nucleotide diversity would
likewisebe∼1/1,000.However,thecorresponding
6
∼3x10 differenceswouldonlyoverlapinpartwith
thoseofthefirstpair.ThetotalnumberofSNPsin
thehumanpopulationisestimatedat>12million.
The corresponding MAF are characterized by an
exponential distribution: there are many SNP with
lowMAFandfewerwithhighMAF.Thelatterare
called “common SNPs”. As a matter of fact,
approximately halve of the differences between
two randomly selected human genomes
correspondtoSNPswithMAF<1%.Somepeople
refer to such rare SNPs as “mutations”, but we
prefer to reserve this term for the process that
generatespolymorphisms.
Chromosomes sampled in the population are
characterizedbyspecificcombinationsofallelesfor
strings of adjacent SNPs, called “haplotypes”.
Hence,ifoneconsiderstwoadjacentSNPs,oneC<>T and the other A<->C, the four possible
combinations or haplotypes are C-A, T-A, C-G and
T-G. The human genome and that of domestic
animals is subdivided in “haplotype blocks”, i.e.
segments measuring of the order of 20-Kb (hence
comparable to the size of a typical gene although
there is no coincidence between the limits of
haplotypeblocksandgenes).Haplotypeblacksare
typically characterized by 5-10 haplotypes
accounting for a large majority of chromosomes
encountered in the population. Haplotype blocks
typically encompass tens to hundreds of SNPs,
which could – in theory – assort in thousands of
haplotype combinations. Five to ten haplotypes
per block is thus very limited. This is because
alleles at neighboring SNPs don’t assort
independently. They tend to preferably associate
in specific allelic combinations, a phenomenon
called “linkage disequilibrium” (see Chapter III).
Thelimitsbetweenhaplotypeblockscoincidewith
the position of recombination hotspots where
crossing-overs preferentially occur. Across
recombination hotspots, haplotypes from adjacent
blocks tend to assort independently, i.e. to be in
“linkageequilibrium”.
Thelevelofgeneticpolymorphism(measuredbyπ)
that is observed in a population at a given time,
reflects a balance between (i) the generation of
new variation by the process of mutation, (ii) the
loss of variants as a result of “genetic drift” and
selection. As mentioned before, every newly
formed zygote carries ∼100 de novo mutations.
Thecorrespondingnewvariantiscalledthederived
allele,byoppositiontotheoriginalancestralallele.
The vast majority of the millions of de novo
Chapter2:GenesinpedigreesPage8/14
mutationthatarehenceinjectedinthepopulation
ateverygenerationwillremainveryrare,restricted
to specific families. But the population frequency
of a minority of them will – with time –
progressively increase, to sometimes even
completely supplant the ancestral allele, i.e. reach
fixation.Randomdrift,i.e.therandomsamplingof
alleles from the parents to produce the next
generation,issufficienttoexplainthelossofmost
de novo mutation, the ascent of a lucky few,
and the resulting transient polymorphism that is
observed in outbred populations. However, the
faithofsomedenovomutationsisalsodetermined
by their phenotypic effects, and the resulting
selectiveadvantageor–moreoften–disadvantage
that the confer to the individuals that inherited
them(seeChapterIII).
Substitutions (transitions and transversions) are
nottheonlykindofgeneticpolymorphisms.There
arealso(i)deletionsandinsertionsofoneormore
nucleotides (“indels”), (ii) variable numbers of
tandem repetitions (“VNTR” or “SSR”) of small
motifs defining microsatellites or larger motifs
defining mini- and regular satellites, (iii) copy
number variations (“CNVs”), and (iv) chromosomal
polymorphisms. We will describe those in more
detaillaterinthischapter.
Functional effects of genetic polymorphisms:
codingandregulatoryvariants.Inmammals,most
polymorphisms that affect one (SNPs) or a small
number of nucleotides are probably selectively
“neutral”ornearlyso.Indeed,mostofthemfallin
poorly conserved genome segments located in
intergenic regions or introns, as these represent
the largest fraction of the genome. The faith of
such neutral polymorphisms is primarily
determinedbygeneticdrift.
However, some genetic polymorphisms do have
functional consequences. These include, in
particular, “coding SNPs” (“cSNPs”) which are
changing the amino-acid sequence of a protein,
and “regulatory SNPs” (“rSNPs”) which are
affectinggeneregulation.
cSNPs include (i) missense variants causing aminoacid substitutions which can be conservative (the
two alternative amino-acids belong to the same
category: neutral, polar, positively charged,
negatively charged) or non-conservative (the two
amino-acids are of a different category), (ii)
nonsensevariantswhichchangeatripletcodingfor
an amino-acid in one of the three stop codons
(“stop-gained”), (iii) frame-shift variants which by
incorporatingordeletingbasepairscauseashiftin
the reading frame resulting in the addition of a
nonsensicalpeptideatthecarboxy-terminalendof
a truncated protein. Nonsense and frame-shift
variantsoftenrevealstopcodonsinnon-lastexons.
This usually causes degradation of the
corresponding transcripts perceived as abnormal
and triggering “nonsense mediated RNA decay”
(NMRD).SNPsinORFcanbesynonymous,i.e.not
changing the amino-acid sequence. Most
synonymousSNPsaffectthethirdbaseofacodon.
Deletions or insertions of three or a multiple of
three nucleotides in the open reading frame may
alter the amino-acid sequence without causing a
frame-shift.
An allelic series of cSNPs in the myostatin (MSTN)
geneunderliesthe“double-muscling”phenotypein
cattle. Double-muscled animals exhibit a severe
and generalized muscular hypertrophy. Doublemusclingisencounteredinseveralbreedsincluding
Asturiana, Belgian-Blue, Maine-Anjou, Parthenaise,
Charolais, Piedmontaise, etc. In all these breeds,
double-musclingisduetomutationsthatcauseloss
of MSTN function. MSTN is an autoregulatory
peptide hormone of the TGFβ family that is
secretedbyskeletalmuscleandinhibitsitsgrowth.
AtleastsixMSTNloss-of-functionmutationshave
been identified in cattle so far including a frameshifting 11-bp deletion in the third exon
(p.D273RfsX13=c.818-828delATGAACACTCC), the
substitution of the fifth of nine highly conserved
carboxyterminal cysteines (p.C313Y) mediating an
intramolecular disulfide bridge stabilizing the
structure of the bioactive domain, three stop
codons resulting from single nucleotide
substitutions (p.Q204X, p.E226X, p.E291X), and a
stop codon resulting from a complex insertiondeletion
event
(p.F140X=c.419426delTTAAATTTinsAAGCATACAA). In addition, a
missense mutation (p.F94L) is segregating at high
frequencyinLimousinsandislikelytocontributeto
thepronouncedmusculardevelopment(evenifnot
double-muscledperse)ofthisbreed.
cSNPs may also alter the amino-acid sequence by
perturbingsplicing.Such“splice-sitevariants”may
destroy splice donor or acceptor sites, or/and
activate cryptic splice sites. Splice-site variants
cause exon skipping or exonification of intronic
sequences, and this usually results in gross
alterations of the amino-acid sequence. An
interesting splice-site variant was recently
describedintheMSTNgeneofBlonded’Aquitaine
cattle.Blonded’AquitaineareaFrenchbeefbreed
characterized by a pronounced muscular
hypertrophy reminiscent of double-muscling.
Initially, however, no obvious loss-of-function
mutationswerefoundwhensequencingtheORFof
the MSTN gene in this breed. Recently, French
researchersfoundthattheMSTNmRNAofBlonde
d’Aquitaine animals has a ...-bp insertion at a
position coinciding exactly with the boundary
between exons 1 and 2. They demonstrated that
theinsertionisduetothe“exonification”ofapiece
of intron 1, which is due to a point mutation that
activates a cryptic splice acceptor site. The
illegitimateexon...
Chapter2:GenesinpedigreesPage9/14
Belgian-Bluesprovideuswithanotherexampleofa
splice-site variant. Between 200x and 200x,
breeders noticed an increase in the number of
calves aged ∼6 months exhibiting signs of stunted
growth or growth retardation. Using positional
cloning (see Chapter IV), an A->G SNP (c.124-2A>G) affecting the splice-acceptor site (position -2)
ofthefirstintronoftheRNF11genewasidentified
as being the cause of ∼40% of cases. The splicesitevariantcauseseitherskippingofexon2(which
introduces a frame-shift in the amino-acid
sequence resulting in NMRD or at best a severely
truncated protein), or the activation of a cryptic
splice site seven base pairs downstream of the
original one. This causes a 7-bp deletion in the
mRNA, resulting in a frame-shift that likewise
results in NMRD and a severely truncated protein.
RNF11 is a component of the A20 complex that
down-regulates NF-κB-mediated inflammatory
response. Concomitantly, most animals that are
homozygous for the c.124-2A->G mutation, were
subsequently shown to die at a very young age
from excessive and uncontrolled inflammatory
responses.
Regulatory SNPs or rSNPs affect cis-acting gene
regulatoryelementscontrollingeithertranscription
or a post-transcriptional process. A nice example
of the first type of rSNP in farm animals is a G->A
transitioninasilencerelementlocatedinthethird
intron of the porcine IGF2 gene. The wild-type
version of this element normally binds the ZBED6
trans-acting factor in post-natal striated skeletal
muscle. The mutation abrogates this interaction
leadingtooverexpressionoftheIGF2growthfactor
and a concomitant ∼3% increase in muscle mass
(∼3kgs of additional muscle per 100Kg carcass).
This mutation is present in many European pig
breeds in which its frequency has increased as a
result of selection for meatiness. It is noteworthy
that it was found on a haplotype that differs at
∼1/100 nucleotides from the other haplotypes
foundinEuropeanpigs,anucleotidediversitythat
is an order of magnitude larger than expected
withinspecies.Themutationactuallyoccurredon
a haplotype that is found at high frequency (with
and without the mutation) in Chinese pig breeds.
European and Chinese haplotypes derive from
different sub-species of wild boars, which were
independently domesticated in Europe and China
∼10,000 years ago. The European and Asian wildboarsub-speciesderivedfromacommonancestor
much earlier, probably millions of years ago, and
this explains the high degree of divergence of the
corresponding haplotypes. Chinese pigs were
imported in the 19th century to improve the
leanness of the then extremely fatty European
breeds.Chinesebreedremainofinteresttodayfor
the pig breeding industry because of their
exceptionalfertility.
TexelsheepprovideaniceexampleofarSNPthat
operates at the post-transcriptional level. Texel
sheep are quite famous for their pronounced
muscular development. It is now known that the
meatinessofTexelsheepisdueinpart(∼25%)toa
G->A transition in the 3’UTR of the ovine MSTN
gene. This substitution creates an illegitimate
target site for two microRNAs that are highly
expressedinskeletalmuscle:miR1andmiR206.As
a result, MSTN mRNA with the mutation are
destabilizedandtheirtranslationinhibited,causing
a reduction in circulating MSTN protein thereby
increasingmusclemass.
Monogenic traits. We have seen how genes are
transmitted from parents to offspring via haploid
gametes, as well as the fact that genes are
encountered in the population in distinct forms or
alleleswhichmaydifferfunctionally.Thesefactors
explain to a large extent why individuals differ in
their “phenotype” and how these phenotypes
segregateinfamilies.Forafirstclassofphenotypes,
the differences between individuals are fully
explained by allelic variation at one locus. Such
phenotypes are said to be monogenic. Most
geneticdefectsbehavelargelyasmonogenictraits.
A second class of phenotypes, referred to as
oligogenic, are determined by allelic variation at a
small number of loci (typically two to three). Coat
color variation in domestic animals is typically
oligogenic. Finally, most phenotypes are
determinedbyalargenumberofgenes(referredto
as “polygenes”), as well as by non-genetic
environmental and even stochastic effects. Such
traits are referred to as “complex” or
“multifactorial”. Production traits in livestock are
typically complex, as are most of the common
diseases afflicting human, including cancer, blood
pressure,diabetes,etc.
[Boxx:pedigreedrawingandallelicnomenclature]
The simplest category of monogenic traits are the
Mendelian traits. They are characterized by two
phenotypicclasses(f.i.affectedbyageneticdefect
versus non-affected) and segregate according to
one of five basic Mendelian inheritance patterns:
autosomal recessive, autosomal dominant, X- (or
Z)-linked recessive, X- (or Z-) linked dominant, and
Y-(ofW-)linked.
As their name implies, autosomal recessive traits
are due to recessive alleles at autosomal (or
exceptionally pseudo-autosomal) loci. We will
refertoarecessivealleleinlowercase(sayd),and
use the “+” sign to label the corresponding “wildtype” allele. The defining feature of a recessive
alleleisthatitsphenotypiceffectismaskedin+/d
heterozygous or carrier individuals, which are
“wild-type” in appearance. The trait is only
apparentind/dhomozygousindividuals.
The segregation of an autosomal recessive trait in
an outbred pedigree is characterized by the
following distinctive features: (i) affected
Chapter2:GenesinpedigreesPage
10/14
individuals are usually born to unaffected but
carrier parents, (ii) individuals of both sexes are
equally affected, (iii) after birth of an affected
offspringfromunaffectedparents,¼ofsubsequent
full-sibs are affected, and (iv) an enrichment of
parental consanguinity. Why does parental
consanguinity increase the incidence of offspring
affected with an autosomal recessive condition?
All individuals carry of the order of ∼5 deleterious
recessives. For any given gene, however, the
frequencyofdeleteriousallelesinthepopulationis
-3
verylow,oftheorderof10 .Thustheprobability
that an offspring, from a mating between
individuals that are not closely related, would be
-3
affectedbyoneoffiveconditionsis∼5x½x10 ≈
0.0025. However, the probability for an offspring
oftwocousinstobeaffectedforatleastoneofthe
∼10 diseases carried by the two shared grand6 10
parents is ∼1-(1-(½) ) ≈ 0.15. In addition, this
consanguinous offspring could be affected by a
recessive condition not inherited from shared
grand-parents.
Why are some alleles recessive? Recessive alleles
are typically “loss-of-function” mutations in genes
forwhichonefunctionalcopyissufficienttoensure
correct cellular/organismal operation (“haplosufficiency”).Nonsenseandframe-shiftmutations,
forinstance,usually“knockthegeneout”(creating
so-callednullalleles),butifthecopyinheritedfrom
the other parent is normal, the heterozygous
individualwillbehealthy.
[Theoryofrecessivityforenzymes]
The long list of genetic defects typically involves
such recessive, loss-of-function mutations. The
OMIM database (On-line Mendelian Inheritance in
Man; www....) reports x such human defects.
Causal genes and mutations have been identified
for ... of these. The OMIA database (On-line
Mendelian Inheritance in Animals; ...) reports x
such animal defects. Causal genes and mutations
havebeenidentifiedfor...ofthese.
Autosomal dominant traits are due to dominant
alleles at a (pseudo-) autosomal locus. We will
typically refer to a dominant allele in upper case
(say D), and use the “+” sign to label the
corresponding “wild-type” allele. The defining
feature of a dominant allele is that its phenotypic
effect is apparent in +/D heterozygous or carrier
individuals, as well as in D/D individuals. Only
homozygous+/+individualshavenormal,wild-type
appearance. In outbred populations, autosomal
dominanttraitarecharacterizedby(i)thefactthat
affected individuals have at least one affected
parent, (ii) that they equally affect individuals of
bothsexes,(iii)thattheyareequallytransmittedby
both sexes, and (iii) one in two affected amongst
offspringofaffectedparents(ifrarecondition).
Alleles may be dominant for several molecular
reasons. Mutations which result in a “gain-offunction” are typically dominant. An interesting
example of such a gain-of-function is the so-called
Pittsburghalleleinthehumanproteaseinhibitor1
gene (PI). PI encodes α1-antitrypsin, a protease
whichnormallytargets(therebyinhibiting)elastase.
PI null mutations underlie recessively inherited
emphysema due to the exacerbated elastase
activity.ThePIPittsburghallele,observedina14yearoldboy,hasaGtoTtransversionthatcauses
a missense methionine to arginine substitution at
position 358 (p.Met358Arg). As a result, the
mutant α1-antitrypsin loses its affinity for elastase
but targets coagulation factors, thereby causing a
dominantly inherited bleeding disorder. Other
examples of alleles that act dominantly as a result
of a gain-of-function are mutations that cause
membrane-bound
receptors
to
become
constitutively active. +++ Some “loss-of-function”
mutations are dominant. Amongst these are
“dominant negative” mutations in genes encoding
subunitsofmultimericproteins.Dominantnegative
mutations abrogate the normal function of the
subunit but not its ability to integrate in the
multimeric complex. When integrated, the
mutated subunit acts as a poison, inactivating the
complex. Imagine a complex like hemoglobin,
comprising two alpha and two beta-subunits. An
individual carrying a dominant negative α-globin
allele would only have 25% of its hemoglobin
tetramersdevoidofdefectivesubunits,whichmay
causedisease.Examples:+++
Denovomutations.
X-(orZ-)linkedrecessivetraitsareduetorecessive
alleles of genes mapping to the X- (or Z-) specific
portion of the gonosomes (i.e. not the pseudoautosomal region). While both sexes can be
affectedinprinciple,individualswiththecondition
are nearly exclusively heterogametic (for rare
conditions). Indeed, hemizygous heterogametic
individualsonlyrequiretheirsinglegenecopytobe
mutant to be affected, while homogametic
individualsneedtwomutantcopiestoexpressthe
condition. Assuming that the frequency of the
recessivealleleisp,thefrequencyofthecondition
2
is ∼p in heterogametic individuals yet ∼p in
homogametic ones. For rare conditions, the
resulting 1/p ratio is thus very large. Affected
heterogametic individuals inherit the mutation
from their usually asymptomatic carrier
homogametic parent, which will often have
affected heterogametic relatives. Affected
homogametic individuals can result from the
mating between an affected heterogametic and
carrierhomogameticparent.
Ofnote,ifthegeneissubjecttoX-inactivation(asis
thecaseforthemajorityofgenesontheX-specific
region),∼halvethecellsofa+/dcarrierXXfemale
will have inactivated the + allele and therefore be
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deficient for the corresponding gene. If the gene
productissharedbetweencells,thismayhavelittle
if any consequence for the operation of the cell.
However, if the encoded function is cellautonomous,∼halvethecellswillbedysfunctional.
In such cases, mosaic XX females will have a
mixture of healthy and deficient cells. This is for
instance observed in women carrying loss-offunction mutations in the ... gene (causing
anhidroticectodermaldysplasiaind/-menandd/d
women), which exhibit dermal patches devoid of
sweat glands adjacent to normally operating skin
sectors.
An example of a Z-linked recessive trait in animals
is“Z-linkeddwarfism”inpoultry....
X- (or Z-) linked dominant traits are due to
dominantallelesofgenesmappingtotheX-(orZ-)
specific portion of the gonosomes. In outbred
populations both sexes are affected, yet
homogametic individuals are more often affected
thanheterogameticones.Indeed,ifthefrequency
of the dominant allele in the population is p,
heterogametic individuals have a probability ∼p of
being affected, while the probability for
homogametic individuals is ∼2p(1-p). Hence, the
ratio is 2(1-p) ∼ 2 for rare conditions. For rare
conditions, offspring of affected homogametic
individuals have a ½ probability to be affected,
irrespective of their sex. On the contrary, all
homogametic offspring of affected heterogametic
parents will be affected while none of the
heterogametic offspring will. As a result of Xinactivation+/Dfemalesareoftenmoremildlyand
morevariablyaffectedthanmales.
An example of a Z-linked dominant trait is “slow
feathering”inpoultry....
Y-(W-)linkedtraits,involvinggenesmappingtothe
Y-(W-)specificportionofthegonososmes,areonly
expressed in the heterogametic sex and
transmittedfromheterogameticparentstoalltheir
heterogametic offspring. The only known example
ofaY-linkedtraitis...
For some supposedly Mendelian traits, the
proportionofaffectedindividualsmaydeviatefrom
expectation. This may for instance be caused by
incompletepenetrance.Thepenetranceisdefined
astheproportionofindividualsofagivengenotype
that expresses the trait. For a regular autosomal
recessive trait the penetrance of genotypes +/+,
+/dandd/d(where+isthewild-typealleleandda
recessive disease causing allele) is 0, 0 and 1. For
some traits (including genetic defects) not all d/d
individuals are affected. Such traits are said to be
characterized by incomplete penetrance.
Incomplete penetrance results from the fact that
individuals with same genotype, f.i. d/d may have
have been exposed to distinct environments or
differ at other genes influencing disease outcome.
Penetrance may differ between sexes or evolve
with age, defining distinct liability classes. As an
example, the penetrance of the autosomal
dominant Huntington’s chorea in human is ..., ...
and1for+/Dindividualsagedrespectively20,40
and70yearsofage.
Deviations from expected Mendelian proportions
also occur in the case of lethality of one of
genotype. One such example is the tail-less
phenotype of so-called Manx cats, caused by the
dominant ... allele. Contradicting Mendel’s law,
thematingoftwo+/...Manxcatsyields2/3rather
than ¾ affected cats. This is due to the fact that
the¼.../...dieinuterofromseveredevelopmental
anomalies.
Genes underlying Mendelian traits may be
characterized by allelic heterogeneity, i.e. there
maybemultipledistinctmutationsinthegene.In
some instances, the different mutations will cause
the same phenotype. One such example is
“double-muscling” in cattle involving an allelic
series of loss-of-mutations in the MSTN gene (cfr.
above). In other instances, distinct mutations in
the same gene may cause different phenotypes.
Example...
compoundheterozygotes
Mendelian traits are typically binary, i.e.
characterized by two distinct phenotypic classes.
Somemonogenictraits,however,arecharacterized
by more than two phenotypic classes. This is for
instance the case when the phenotype of
heterozygotes differs from that of either
homozygotes, which can be due either to semidominant or co-dominant alleles. Semi-dominant
alleles cause the heterozygotes to have a
phenotype, measured on a continuous scale,
intermediate between the two heterozygotes. As
an example, the muscular development of cattle
carrying one copy of MSTN loss-of-function alleles
(causing
full-blown
double-muscling
in
homozygotes),isslightlyhigher(xresidualstandard
deviations) than that of homozygous wild-type
animals. MSTN loss-of-function alleles are
therefore said to be semi-dominant or also partial
recessives.
Alleles are said to be codominant when
heterozygotes express the phenotypes associated
withbothalleles.Examplesofcodominancecanbe
foundinthebloodgroups.ThehumanABOblood
group system for instance, is the manifestation of
three alleles of a gene (I) encoding a glycosyl
A
transferase. The I allele catalyzes the addition of
N-acetylgalactosamine(=Aantigen)toaprecursor
carbohydrate present on the surface of red blood
B
andothersomaticcells.TheI allele,differingfrom
theIAalleleatfouramino-acidpositions,catalyzes
the addition of galactose (=B antigen) to the the
O
sameprecursor.TheI alleleisanullalleleofthe
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transferase. The added N-acetylgalactosamine
groups can be detected using anti-A antibody,
while the galactose residues are evidenced using
AB
anti-B antibody. Red blood cells from I I heterozygous individuals simultaneously express
AA
boththeAandBantigenontheirsurface,fromI I AO
BB
andI I individualsonlytheAantigen,fromI I and
BO
OO
I I individuals only the B antigen, and from I I individualsneithertheAnortheBantigen.[MHC]
Another example of codominance is the roan
phenotypeincattle.+++
Oligogenictraits
When the phenotypic differences between
individualsreflectallelicvariationatmorethanone
yet a small number of loci, the trait is said to be
oligogenic. Oligogenic traits can be classified
according to the number of phenotypic classes
(two, three or four). Oligogenic traits with two
phenotypic classes involve complementation,
suppressionandduplication,withthreephenotypic
classes epistasis, and with four phenotypic classes
modifiergenes.
Complementation.Thesametraitmayreflectallelic
variation at distinct loci in distinct families or
populations. Such traits are said to be
characterized by “locus heterogeneity”. As an
example,inheriteddeafnessinhumanmaybedue
torecessiveloss-of-functionmutationsinatleastx
genes. This explains why deaf couples often have
children with perfectly normal hearing. If the
deafnessoftheparentsresultsfromhomozygosity
for null alleles at two distinct loci, all children will
be double carriers but have a functional allele at
each locus therefore have normal hearing. The
outcome whereby the mating between mutant
parents yields 100% wild-type offspring is referred
to as complementation. Complementation tests
play an essential role in the genetic dissection of
metabolicpathwaysinmodelorganisms.Imagine
thataphenotype..explainwithexample].
Suppression. Alleles at some loci (say B) may
suppress the effect of mutants alleles at another
locus (say A). The suppressive alleles may be
dominant or recessive. Assume an F2 generation
segregatingattwolociAandB(F1genotypeAaBb).
Assume also that the mutant phenotype depends
ontherecessiveaallele.IntheF2generation,the
proportionofmutantindividualswillbe3/16(aaB)incaseofrecessivesuppression,and1/16(aabb)
incaseofdominantsuppression.Examples...
Duplication. Some organisms contain more than
two copies of all or par gene. This is particularly
common in polyploid species, whose genome has
undergone duplication or more in an ancestral
species.
Epistasis. Imagine a locus with two alleles A and a
underlyingabinarytrait,animalshavingphenotype
“A” (genotype A-) or “a” (genotype aa). A distinct
locus (say B, with alleles B and b) is said to be
epistatic(litterally“standingupon”)withrespectto
locus A, if individuals with genotype B- (in which
casetheepistasisissaidtobedominant)orbb(in
which case the epistasis is said to be recessive)
exhibit a distinct, third phenotype (say “B” or “b”)
maskingeitherofthetwophenotypesdetermined
by the A locus. In a F2 generation obtained by
intercrossing double heterozygous F1 parents
(genotypeAaBb),thethreephenotypes(A,aandB
or b) are expected in proportions 3:1:12
corresponding to genotypes A-bb:aabb:--B-
(dominant epistasis), or 9:3:4 corresponding to
genotypes A-B-:aaB-:--bb (recessive epistasis). A
well known example of recessive epistasis in
domesticanimalsistheinteractionbetweenthe“X”
and“E(xtension)”lociinlabradorretrievers.The“X”
locus is characterized by the ... and ... alleles,
explainingtheoccurenceofblack(genotype...)and
brownorchocolate(genotype...)labradors.TheX
locuscorrespondstothe...gene....The“E”locus
isalsocharacterizedbytwoalleles:...and...Dogs
with ... genotype will either be black or chocolate
depending on their genotype at the ... locus.
However labradors of ... genotypes will be yellow
or golden irrespective of their genotype at the ..
locus.TheElocuscorrespondstothe...gene....
Modifiergenes.
Sex
Polygenicinheritance(heritability)
Parentalimprinting(callipyge)
Mitochondrial:faithofpaternalmitochondria
Epigeneticinheritance
Chromosomal anomalies (aberrant euploidies,
aneuploidies,
isodisomies;
translocations,
inversions,duplications)
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