Trio, a novel high fecundity allele: I. Transcriptome analysis of granulosa cells from carriers and noncarriers of a major gene for bovine ovulation rate†

Mamat H Kamalludin, Alvaro Garcia-Guerra, Milo C Wiltbank, Brian W Kirkpatrick

A major gene for bovine ovulation rate has been mapped to a 1.2 Mb region of chromosome 10. Screening of coding regions of positional candidate genes within this region failed to reveal a causative polymorphism, leading to the hypothesis that the phenotype results from differences in candidate gene expression rather than alteration of gene structure. This study tested differences in expression of positional candidate genes in granulosa cells between carriers and non-carriers of the high fecundity allele, as well as characterizing differences in the transcriptomic profile between genotypes. Five carriers and five non-carriers, female descendants of "Trio," a carrier of the high fecundity allele were initially used in an RNA-seq analysis of gene expression. Four of ten samples were contaminated with theca cells, so that six samples were used in the final analysis (three of each genotype). Of 14,973 genes expressed, 143 were differentially expressed (false discovery rate p < 0.05) in carriers versus non-carriers. Among the positional candidate genes, SMAD6 was 6.6 fold over expressed in the carriers compared to non-carriers (p < 5 × 10-5). This result was replicated in an independent group of 12 females (7 carriers, 5 non-carriers) using quantitative real-time PCR; SMAD6 was 9.3 fold overexpressed in carriers versus non-carriers (p = 1.17 × 10-6). Association of overexpression of SMAD6, an inhibitor of the BMP/SMAD signaling pathway, with high ovulation rate corresponds well with disabling mutations in ligands (BMP15, GDF9) and a receptor (BMPR1B) of this pathway that cause increased ovulation rate in sheep.