Structure, function and transcriptional regulation of JY-1: A novel gene specific to bovine oocytes/embryos

The oocyte plays key roles in regulation of female fertility. A growing body of evidence suggests that genes expressed in the oocyte are important for normal folliculogenesis, oocyte function and early embryogenesis and thus required for subsequent establishment of pregnancy. However, the identities of the oocyte expressed genes regulating above processes and required for fertility are limited, especially in monotocous mammals and farm animal species. During initial analysis of expressed sequence tags (ESTs) from a bovine oocyte cDNA library, an abundant novel oocyte expressed transcript (designated as JY-1) was identified. Additional cloning and sequence analysis indicated that the JY-1 sequence is totally novel and encodes for a secreted protein of unknown function. Expression of JY-1 mRNA is ovary specific, intraovarian expression of JY-1 mRNA and protein is restricted exclusively to the oocyte and immunoreactive JY-1 protein of ∼11,000 M, is detectable within lysates of bovine oocytes. Furthermore, JY-1 mRNA is temporally regulated during the window of meiotic maturation through embryonic genome activation and embryonic JY-1 mRNA is oocyte derived. Based on its oocyte specific expression and temporal and spatial expression pattern in embryos, I hypothesize that the JY-1 gene is conserved in other species and may have an important function during bovine folliculogenesis and early embryogenesis. To test this hypothesis, I will (a) determine the gene structure of bovine JY-1 and investigate presence of orthologous genes to JY-1 in other species (b) determine the requirement of JY-1 for early embryonic development and c) determine the ability of JY-1 to modulate FSH induced regulation of granulosa cell function. Genomic Southern blot analysis detected putative JY-1 genes in bovine, sheep, pig and human genomic DNA. Genomic library screening and mining of sequence from available genome databases revealed that the JY-1 gene is located on bovine chromosome 29, approximately 16 kb in length, and encoded by 3 exons (25, 92 and 1400 by in length) separated by two introns (12.8 and 1.5 kb in length). JY-1 like sequences were identified in DNA fragments on human chromosome 11 and syntenic regions in other species (chimpanzee chromosome 11, dog chromosome 21, mouse chromosome 7 and rat chromosome 1). Expression of a putative human mRNA ortholog of JY-1 was detected in human ovary. Extensive sequence analysis of the putative human mRNA ortholog of JY-1 and the conserved JY-1-like sequences in the genome of other vertebrate species did not reveal an open reading frame. Disruption of JY-1 mRNA and protein in early embryos by siRNA mediated gene knockdown perturbed embryo development to the blastocyst stage. Treatment of granulosa cells with recombinant JY-1 (rJY-1) stimulated progesterone synthesis in a dose dependent manner, but decreased the FSH stimulated increase in granulosa cell number and estradiol concentrations. Based on these results, I conclude that JY-1 is a novel protein encoding maternal effect gene restricted to the genome of cattle and potentially other ruminant species with important roles in regulation of folliculogenesis and early embryogenesis.