The highly prolific phenotype of Lacaune sheep is associated with an ectopic expression of the B4GALNT2 gene within the ovary.

Laurence Drouilhet,Grégoire Harichaux,Loys Bodin,Philippe Bardou,Jérome Lluch,Julien Sarry,Catherine Viguié,Stéphane Fabre,Florent Woloszyn,Camille Mansanet,Philippe Mulsant,Danielle Monniaux,Kamila Tabet

doi:10.1371/journal.pgen.1003809

Abstract

Prolific sheep have proven to be a valuable model to identify genes and mutations implicated in female fertility. In the Lacaune sheep breed, large variation in litter size is genetically determined by the segregation of a fecundity major gene influencing ovulation rate, named FecL and its prolific allele FecLL. Our previous work localized FecL on sheep chromosome 11 within a locus of 1.1 Mb encompassing 20 genes. With the aim to identify the FecL gene, we developed a high throughput sequencing strategy of long-range PCR fragments spanning the locus of FecLL carrier and non-carrier ewes. Resulting informative markers defined a new 194.6 kb minimal interval. The reduced FecL locus contained only two genes, insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) and beta-1,4-N-acetyl-galactosaminyl transferase 2 (B4GALNT2), and we identified two SNP in complete linkage disequilibrium with FecLL. B4GALNT2 appeared as the best positional and expressional candidate for FecL, since it showed an ectopic expression in the ovarian follicles of FecLL/FecLL ewes at mRNA and protein levels. In FecLL carrier ewes only, B4GALNT2 transferase activity was localized in granulosa cells and specifically glycosylated proteins were detected in granulosa cell extracts and follicular fluids. The identification of these glycoproteins by mass spectrometry revealed at least 10 proteins, including inhibin alpha and betaA subunits, as potential targets of B4GALNT2 activity. Specific ovarian protein glycosylation by B4GALNT2 is proposed as a new mechanism of ovulation rate regulation in sheep, and could contribute to open new fields of investigation to understand female infertility pathogenesis.

Highlights

Women, cattle, goats and ewes have generally one or two offspring, whereas other mammals, such as sows, rodents and dogs are prolific and produce more than three offspring
Cattle, goats and ewes have generally one or two offspring, whereas other mammals, such as sows, rodents and dogs are prolific and produce more than three offspring. It relies on the number of ovulations at each estrus cycle i.e. the ovulation rate (OR), for which the underlying genetic mechanism was puzzling until the identification of fecundity genes in sheep, bone morphogenetic protein-15 (BMP15), growth and differentiation factor-9 (GDF9) and Bone Morphogenetic Protein (BMP) receptor-1B (BMPR1B) [1]
Genetic characterization of the FecL locus The fine mapping strategy combining marker development through high-throughput sequencing and genotyping of selected recombinant animals allowed the delimitation of the FecL locus on OAR11 between single nucleotide polymorphism (SNP) markers g.36910171T.C and g.37107627G.C

Summary

Introduction

Cattle, goats and ewes have generally one or two offspring, whereas other mammals, such as sows, rodents and dogs are prolific and produce more than three offspring. It relies on the number of ovulations at each estrus cycle i.e. the ovulation rate (OR), for which the underlying genetic mechanism was puzzling until the identification of fecundity genes in sheep, bone morphogenetic protein-15 (BMP15), growth and differentiation factor-9 (GDF9) and BMP receptor-1B (BMPR1B) [1]. FecX is known as BMP15 and in the Lacaune breed, the mutant allele (FecXL) associated with high prolificacy was identified as a pCys321Tyr substitution altering the BMP15 protein function [3]. Heterozygous FecXL mutation is associated with a twofold increase in OR, Author Summary

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Conclusion