Changes in the transcriptome of bovine ovarian cortex during follicle activation in vitro

Abstract

The signals that regulate activation, a key transition in ovarian follicular development, are still not well understood, especially in nonrodent species. To gain insight into the regulation of this transition in cattle, we combined a microarray approach with an in vitro system in which ovarian cortical pieces cultured in control medium are enriched for primordial follicles, whereas pieces cultured with insulin are enriched for primary follicles. Total RNA was extracted from cultured cortical pieces, and then transcripts were identified and analyzed using the Affymetrix Bovine Genome GeneChip array. Around 65% of the transcripts in the bovine GeneChip were detected in cultured cortical pieces. Comparison between pieces cultured with or without insulin generated 158 differentially expressed transcripts. Compared with controls, 90 transcripts were upregulated and 68 were downregulated by insulin. These transcripts are involved in many biological processes and functions, but most are associated with cellular growth or cell cycle/cell death. The transcript encoding ubiquitin-conjugating enzyme E2C (UBE2C) was significantly upregulated during follicle activation, and Ingenuity Pathways Analysis revealed that UBE2C can interact with the tumor suppressor phosphatase and tensin homolog (PTEN). Both PTEN mRNA and protein were lower in cortical pieces cultured with insulin than in controls. In addition, FOXO3a, a downstream effector of PTEN signaling, underwent nuclear-cytoplasmic shuttling during primordial to primary follicle development in bovine fetal ovaries, further suggesting the involvement of the PTEN pathway in follicle activation in cattle. Genes and pathways identified in this study provide interesting candidates for further investigation of mechanisms underlying follicle activation.