The Highly Prolific Phenotype of Lacaune Sheep Is Associated with an Ectopic Expression of the B4GALNT2 Gene within the Ovary

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.

Genes con efecto mayor sobre la fertilidad de ovejas. Revisión

“…The high prolificacy of Lacaune sheep was explained by overexpression of B4GALNT2 in the ovary leading to atypical glycosylation of inhibin (54) .…”

“…La elevada fertilidad de las ovejas Lacaune se explica por a sobreexpresión de B4GALNT2 en el ovario llevando a una glicosilación atípica de la inhibina (54) .…”

The high prolificacy of D’man sheep is associated with the segregation of the FecLL mutation in the B4GALNT2 gene

“…Even if quite imprecise due to the relatively low number of animals studied, we have determined a high frequency (0.65) and a rather strong effect (+0.25 lamb/lambing) of the FecL L allele in the Tunisian D'man sheep, highlighting the presence of a first major gene involved in the genetic determinism of LS variability within this breed known for its high prolificacy. The FecL L mutation was firstly discovered in Lacaune breed (Drouilhet et al, ) and more recently in Noire du Velay (Chantepie et al, ), thus raising the question of the origin of this mutation. It is more likely that the FecL L mutation has an ancestral origin which occurred before the establishment of Lacaune, Noire du Velay and D'man breeds.…”

“…The precise mechanism of action of the intronic FecL L on the B4GALNT2 gene regulation and over expression within the granulosa cells is still unknown. Several hypotheses such as the impairment of transcription factor or miRNA binding and mRNA alternative splicing were tested in vitro or in silico but without any success (Drouilhet et al, ). The use of high‐throughput sequencing technologies dedicated to chromatin accessibility and detection of all kind of RNA (small or long non‐coding RNA, mRNA) in the presence or absence of the mutation could contribute to understand the mechanism in the future.…”

“…At the molecular level, the FecL L mutation leads to a huge increase in the B4GALNT2 expression and glycosylation activity within the ovarian granulosa cells targeting the inhibin/activin system. This glycosylation mechanism was proposed as the physiological determinant of the regulation of ovulation rate and LS in the Lacaune breed (Drouilhet et al, ).…”