genetic evidence that lower circulating

7/24/2019 Genetic Evidence That Lower Circulating

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GENETIC evidence that lower circulating

FSH levels lengthen menstrual cycle,

increase age at menopause and impact

female reproductive health

atherine S! "uth,"o#in N! $eaumont, %essicaTyrrell, Samuel E! %ones,

&arcus '! Tu(e, Hanieh )aghoot(ar, 'ndrew "!*ood,"achel &! Freathy, &ichael N!*eedon, Timothy &! Frayling, and

'nna &urray+

Genetics of Comple Traits, -niversity of Eeter &edical School, "I./ .evel 0, "oyal

/evon and Eeter Hospital,

$arrac( "oad, Eeter E12 3/*, -

+Correspondence address! E4mail5 a!murray6eeter!ac!u(

Su#mitted on Septem#er 78, 2973: resu#mitted on ;cto#er 2

summaryanswer5 The T allele of the FSH$ promoter polymorphism ?rs79@03A0@: c!4277G!TBresults in longer menstrual cycles and

later menopause and, while having detrimental effects on fertility, is protective against

endometriosis!

what is (nown already5 The FSH$ promoter polymorphism ?rs79@03A0@: c!4277G!TB affects

levels of FSH$ transcription and, as a

result, circulating levels of FSH! FSH is re=uired for normal fertility and genetic variants at

the FSH$ locus are associated with age at menopause and

polycystic ovary syndrome ?C;SB!

study design, siDe, duration5 *e used cross4sectional data from the - $io#an( to loo( at

associations #etween the FSH$

promoter polymorphism and reproductive traits, and performed a genome4wide association

study ?G*'SB for length of menstrual cycle!

participantsmaterials, setting, methods5 *e included white $ritish individuals aged 9A8

years in 299A2979, in the

&ay 2973 release of genetic data from - $io#an(! *e tested the FSH4lowering T allele of

the FSH$ promoter polymorphism ?rs79@03A0@:

c!4277G!TB for associations with 28, mainly female, reproductive phenotypes in up to A0 039

women and 3A A9@ men! *e conducted a

G*'S in 830 individuals to identify genetic variants associated with length of menstrual

cycle!

main results and the role of chance5 The FSH4lowering T allele of the FSH$ promoterpolymorphism ?rs79@03A0@: &'F

9!7AB was associated with longer menstrual cycles 9!7A S/ ?c! 7 dayB per minor allele: 83

confidence interval ?CIB 9!729!29:

J A K 7927AL, later age at menopause ?9!70 years per minor allele: 83 CI 9!99!22: J

3!< K 7920B, greater female nulliparity odds

ratio ?;"B J 7!9A: 83 CI 7!927!77: J !@ K 7920L and lower ris( of endometriosis ?;"

J 9!


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limitations, reasons for caution5 The data included might #e affected #y recall #ias! Cycle

length was not availa#le for 23 of

women still cycling ?7 did not answer, A did not (now and for 7@ cycle length was

recorded as MirregularB! *omen with a cycle length

recorded were aged over 9 and were approaching menopause: however, we did not find

evidence that this affected the results! &any of thegroups with illnesses had relatively small sample siDes and so the study may have #een under4

powered to detect an effect!

wider implications of the findings5*e found a strong novel association #etween a genetic

variant that lowers FSH levels and

longer menstrual cycles, at a locus previously ro#ustly associated with age at menopause! The

variant was also associated with nulliparity and

endometriosis ris(! These findings should now #e verified in a second independent group of

patients!*e conclude that lifetime differences in

circulating levels of FSH #etween individuals can influence menstrual cycle length and arange of reproductive outcomes, including menopause

timing, infertility, endometriosis and C;S!

study fundingcompeting interest?sB5None!

trial registration num#er5Not applica#le!

ey words5 FSH # su#unit menstrual cycle menopause endometriosis fertility

Introduction

FSH is a (ey pituitary hormone, which stimulates maturation of oocytes

and is a #iomar(er of ovarian reserve! FSH is a heterodimer comprised a

hormone4specific #4chain ?FSH4#B associated with an a4chain shared #y

other mem#ers of the glycoprotein hormone family ?NagirnaOa et al!,

2979B! The anterior pituitary produces FSH, with transcription of FSH$

#eing the rate4limiting step for FSH production! FSH stimulates target

cells #y #inding to the FSH receptor ?FSH"B, a G4protein4coupled receptor

?Fan and Hendric(son, 2993B, promoting follicle maturation and estrogen

production in women, and Sertoli cell proliferation and

spermatogenesis in men ?NagirnaOa et al!, 2979B!

"are mutations in the FSH$ gene cause truncation of the FSH4#

protein and result in hypogonadism and primary amenorrhoea in

females ?.ayman et al!, 788


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homeodomain transcription factor #inding ?$enson et al!, 2970B! The

T allele of rs79@03A0@ ?c!4277G!TB is associated with lower FSH

levels in men and women, and with higher .H and lower testicular

volume, sperm count, FSH.H ratio, inhi#in $ and testosterone in

men, and has #een found at a higher prevalence in infertile men ?Grigorova

et al!, 299@, 2979, 2977: Tuttelmann et al!, 2972: .a &arca et al!,2970: Schuring et al!, 2970: Simoni and Casarini, 297: "uth et al!,

2973B! Genetic association studies have identified signals at the FSH$

locus associated with age at menopause ?Stol( et al!, 2972: /ay et al!,

2973B, polycystic ovary syndrome ?C;SB ?Hayes et al!, 2973B and

levels of .H ?Hayes et al!, 2973: "uth et al!, 2973B!

-sing the uni=ue resource of the - $io#an( ?'llen et al!, 297B,

we show that a common genetic variant (nown to alter FSH levels

impacts a wide range of traits important to female reproductive

health, including fertility, endometriosis and menstrual cycle length! In

the first genome4wide association study ?G*'SB for menstrual cycle

length, we identified the FSH$ locus as the only signal associatedwith this trait!

&aterials and &ethods

Source of data

The - $io#an( includes data for 390 023 people aged 9A8 years recruited

in 299A2979 from across the - ?'llen et al!, 297B!*e analysed data from

the&ay 2973 interim release of imputed genetic data from- $io#an(,which

contains


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loss ?!7 caseB!

*e defined two infertility4related #inary phenotypes: never pregnant

?femalesB and never fathered a child ?malesB! *e analysed female medical

conditions as #inary outcomes, comparing people reporting a condition

?caseB with those who did not ?controlB! &edical conditions included

dysmenorrhoea, endometriosis, fi#roids, irregular menstrual cycles, menopausalsymptoms, menorrhagia, ovarian cysts, C;S, uterine polyps,

vaginaluterine prolapse and #reast, endometrial and ovarian cancer! 's

more general indicators of gynaecological health, we included the medical

interventions #ilateral oophorectomy or hysterectomy in our analysis!

articipants

In our analysis, we included individuals who #oth self4identified as white

$ritish and were confirmed as ancestrally Caucasian #y - $io#an( from

genetic information ?n J 72@ 2AAB! *e calculated principal components

?CsB for inclusion as covariates in our analyses using FlashC' ?'#raham

and Inouye, 297B! Cs were calculated in 729 2@A unrelated participants

?as identified #y - $io#an(B #ased on 83 303 independent, directly genotyped

SNs ?pairwise r2 , 9!7B! These SNs had a minor allele fre=uency

?&'FB 2!3 and missing4ness ,7!3 across all participants in the &ay

2973 interim release of genetic data, and had a Hardy*ein#erg e=uili#rium

?H*EB ! 7 K 792A within the white $ritish participants!

Testing for associations of theFSH$promoter

polymorphism with reproductive phenotypes

*e tested the FSH4lowering T allele of the FSH$ promoter polymorphism

?rs79@03A0@: c!4277G!TB for associations with reproductive phenotypes

?up to A0 039 women and 3A A9@ menB! SN rs79@03A0@ was wellimputed in the data ?imputation =uality 9!883: H*E J 9!7A: missing

rate J 9!0B! 'll analyses were carried out in males or females as appropriate

?#ased on self4defined seB using Stata ?v70B ?StataCorp ., College

Station, T1, -S'B!

For continuous phenotypes, we transformed the phenotype #y adOusting

for recruitment centre, age at recruitment and the first five Cs prior

to inverse4normaliDation! *e performed linear regression of transformed

phenotype on imputed minor4allele dosages at SN rs79@03A0@ with genotyping

chip as a covariate!*e carried out a sensitivity analysis of the effect of

different transformations, e!g! inverse normaliDing the trait prior to calculating

the residuals:however, this did not materially affect our results! Since the dataon length of menstrual cycle included a wide range of values ?Supplementary

data, Figs S7 and S2B,wecarried out analyses on cycles from27 to 03 days and

in women aged ,3 and 3 years at recruitment!*e validated our results

for length of menstrual cycle #y carrying out analyses in two randomly chosen,

e=ually siDed groups! For age at menopause and age at menarche,we also ran

analysis using the phenotype definition from the "eproGen Consortium

G*'S ?www!reprogen!orgB ?untransformed age at menopause #etween

9 and A9 years not adOusted for age, untransformed age at menarcheB to

allow comparisons with pu#lished data ?Stol( et al!, 2972: erry et al!,

297a,#: /ay et al!, 2973B!

For #inary outcomes, we performed logistic regression of the phenotypeon minor4allele dosages at SN rs79@03A0@ including the first five Cs, recruitment


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centre, age at recruitment and genotyping chip as covariates!

G*'S of length of menstrual cycle

*e conducted aG*'S to identify genetic variants associated with length of

menstrual cycle ?n J 830B using the $;.T4.&& algorithm ?descri#ed in

.oh et al!, 2973B from the freely availa#le $;.T4.&& software pac(age

version 2!2, https5data!#roadinstitute!orgal(esgroup$;.T4.&&?7< /ecem#er 2973, date last accessedBL to account for relatedness and

population structure! This allowed us to include related individuals who

were ecluded from the association analysis of the FSH$ promoter polymorphism

?Supplementary data, Ta#le SIB!*e transformed length of menstrual

cycle #y adOusting for recruitment centre and age at recruitment

prior to inverse4normaliDation, and performed association testing while

adOusting for genotype chip! *e filtered results on imputation =uality !9!,

H*E ! 7 K 7923, and &'F !9!7, resulting in P7A!@ million variants

that were tested! 's the - $io#an( G*'S included more variants than a

standard G*'S and we did not have a replication sample availa#le, we

chose a threshold of , 3 K 7928, #ased on a $onferroni correction forthe num#er of variants tested, rather than the conventional , 3 K 792@!

"esults

' common allele in the FSH$ gene, (nown to

lower FSH levels, is associated with longer

length of menstrual cycle

The FSH4lowering T allele of the FSH$ promoter polymorphism

?rs79@03A0@: &'F 9!7AB was associated with longer menstrual cycles

9!7A S/ ?P7 dayB per minor allele: 83 confidence interval ?CIB

9!729!29: J A K 7927AL! ;f the reproductive traits tested ?Ta#les I

and IIB, length of menstrual cycle was the most strongly associated with

rs79@03A0@ ?Fig! 7 and Ta#le IIIB! The SN was also associated with

cycle length when we dichotomiDed data into women reporting a cycle

length of R29 days compared with those reporting an average length

of 2@ days odds ratio ?;"B J 9!


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per minor allele: 83 CI 9!939!77: J 7!8 K 792@B!

The FSH$ allele associated with longer

cycle length is associated with later

menopause

The FSH4lowering Tallele of rs79@03A0@ was associated with later age at

menopause for those in the - $io#an( 9!70 years per minor allele?"eproGen definitionB: 83 CI 9!99!22: J 3!< K 7920L! There was

no association #etween rs79@03A0@ and menopause age when we

dichotomiDed the phenotype into early menopause compared with

later menopause ?Ta#le IIIB! The FSH$ locus is (nown to #e associated

with timing of menopause5 in aG*'S conducted #y the "eproGen consortium,

the signal at this locus ?rs722879B increases age at menopause

#y 9!20 years ?83 CI 9!7A9!28: J 7!3 K 79277B ?Stol(

et al!, 2972B! .ater menopause has #een shown to #e associated with

later age at last #irth ?'yatollahi et al!, 2990: /orOgochoo et al!, 299@B

and rs79@03A0@ was also associated with later age at last #irth 9!92

S/ ?P9!7 yearsB per T allele: 83 CI 9!999!9: J !2 K 7922L!.onger cycle length is not a general feature of

alleles associated with later age at menopause

*e net tested the role of all 3A genetic variants associated with age

at menopause! In addition to the age at menopause signal at the FSH$

locus ?rs722879B, only one of the other 33 pu#lished age at menopause

signals was nominally associated with cycle length ? ! 9!93B5

rs79


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having the medical interventions #ilateral oophorectomy ?;" J 7!72:

83 CI 7!9A7!78: J 7! K 792B and hysterectomy ?;" J 7!70:

83 CI 7!9A7!29: J 7!9 K 792B, which are used as treatments for

a range of gynaecological conditions including endometriosis!

The common FSH$ variant, associated with

FSH levels, is not associated with reproductivetraits more generally

There was no consistent evidence that the FSH$ variant ?rs79@03A0@B

was associated with age at menarche! There was a 9!904year increase

in age at menarche ?"eproGen definitionB per T allele of rs79@03A0@

?83 CI 9!979!93: J 7! K 7922B and the #inary phenotype of

early menarchewas associated at ! 9!93 ?Ta#le IIIB! None of 722 pu#lished

G*'S signals for menarche ?erry et al!, 297a,#B were associated

with length of menstrual cycle at , 9!99@!

The FSH$ promoter polymorphism ?rs79@03A0@B was not associated

with other reproductive illnesses or conditions at , 9!93 ?Ta#le IIIB,

ecept for menopausal symptoms ?;" J 9!A2: 83 CI 9!79!80: J 9!92B ?Fig! 7B! No associations were found with dysmenorrhoea,

fi#roids, irregular menstrual cycles, menorrhagia, multiple pregnancy

loss, ovarian cysts, C;S, uterine polyps or vaginaluterine prolapse,

or with female #reast, ovarian or endometrial cancer!

/iscussion

In the first G*'S of menstrual cycle length, we found a strong association

#etween an FSH lowering, li(ely functional, variant in the FSH$ promoter

and longer cycles ?Hoogendoorn et al!, 2990: Grigorova et al!,

299@, 2979: Tuttelmann et al!, 2972: $enson et al!, 2970: .a &arca

et al!, 2970: Simoni and Casarini, 297: "uth et al!, 2973B! This locus

has #een previously ro#ustly associated with age at menopause in the

"eproGen consortium G*'S of menopause timing ?Stol( et al!, 2972:

/ay et al!, 2973B and the allele associated with longer cycle length is

also associated with later age at menopause!*e did not o#serve associations

for the maOority of age at menopauseG*'S signals with length

of menstrual cycle, including the four signals with effects of over one4third

of a year per allele on menopause timing, implying that the association is

specific to FSH$5 either FSH4# has independent effects on #oth cycle

length and menopause or changes in cycle length are causally influencing

menopause timing!;ur results are consistent with the o#served epidemiological relationship

#etween longer menstrual cycles and later age at menopause

?*helan et al!, 7889: acDmare(, 299


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pill influences menopause timing ?van Noord et al!, 788


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genetic evidence that lower circulating

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