Expression analysis of gene differentially expressed in the developping ovary

In recent years, several factors required for follicle assembly and/or early growth of newly formed primordial follicles have been characterized, but it is likely that additional factors remain to be identified. We have used cDNA arrays to compare gene expression in the neonatal mouse ovary 48 h after birth (when primordial follicles are being assembled), and at 96 h (when early follicular growth is taking place), to that of ovaries collected less than 24 h after birth (when follicles have not yet been formed). Segregating genes according to their pattern of expression revealed the presence of one cluster of 24 genes whose expression consistently increased at 48 and 96 h. The top increaser in this cluster encodes a ~1.5 Kb mRNA containing an open reading frame of 1,401 bp that encodes a protein of 466 amino acids. The predicted 52.3 kDa protein, which we have termed Eop1 (for Early ovarian protein-1) is a novel member of the F-box only (Fbxo) protein family. Eop1 has a cytoplasmic localization that includes the endoplasmic reticulum. Eop1 mRNA expression is oocyte-specific; Eop1 mRNA is first detected on gestational day 18, decreasing thereafter to minimal levels on the day of birth. Eop1 mRNA prevalence increases again 48 and 96 h after birth, coinciding with the time of follicular assembly and the initiation of early follicular growth. The specific expression of Eop1 in oocytes and its developmental pattern of expression suggest a role for this novel gene in the regulation of oocyte physiology. Keywords: Time-course Ovaries from C57Bl/6J were obtained from pups at <24h and 48h after birth. In this strain of mice, assembly of ovarian follicles is initiated 48h after birth, and growth of the newly formed follicles starts shortly thereafter. To analyze differentially expressed mRNA profiles at the time of follicular formation and early follicular growth, we employed a two-dye spotted cDNA microarray containing 8,400 gene probes generated from a mouse NIA 15K gene set (GPL5369), printed in duplicate on glass slides by the Gene Microarray Shared Resource Facility of the Oregon Health & Science University. RNA samples from 2 (2-1, 2-2, 2-3) and 4 day-old mice (4-1, 4-2, 4-3) were compared to RNA samples from 0 day-old mice (0-1, 0-2, 0-3, 0-4, 0-8). Each comparison pair was hybridized twice on the same slide (4 experimental replicates) and three RNA sample were used for each age category (3 biological replicates).