Gene Expression and Functional Studies of Janus Kinase 3 (JAK3) in Ovarian Granulosa Cells

Abstract

Janus kinase 3 (JAK3) belongs to a family of membrane‐associated intracellular non‐receptor tyrosine kinase proteins that mediate signals initiated by cytokine and growth factor receptors through the JAK‐STAT pathway. In contrast with the ubiquitous expression of other family members (JAK1, JAK2, and TYK2), JAK3 is predominantly expressed in hematopoietic cells. We previously identified JAK3 as a differentially expressed gene in granulosa cells (GC) of bovine dominant or preovulatory follicles. The objectives of the current study were to characterize the spatio‐temporal expression pattern of JAK3 in bovine ovarian follicles and to elucidate JAK3 mode of action in GC. GC were obtained from small follicles (SF: 2‐4 mm), dominant follicles (DF) at day 5 of the estrous cycle, and ovulatory follicles (OF) 24 hours (h) following injection of hCG. RT‐PCR analyses showed greatest expression of JAK3 in GC of DF, while the lowest expression was in GC of OF (P < 0.0001). Temporal expression analysis of JAK3 in follicular walls (granulosa and theca cells) of OF at 0, 6, 12, 18 and 24 h after hCG injection showed 5‐ and 20‐fold reduction at 12 and 24 h, respectively, following hCG injection as compared to 0 h (P < 0.05). Results of a yeast‐two‐hybrid screening of DF‐cDNAs library with JAK3 showed activation of the GAL4‐responsive reporters suggesting physical interactions between JAK3 and specific proteins in GC, which were confirmed by co‐immunoprecipitation analyses.This study reports for the first time that JAK3 is differentially expressed in GC of preovulatory follicles supporting a physiologically relevant role of JAK3 in follicular development, and provides insights into the mode of action and function of JAK3 in GC. (Research supported by NSERC of Canada grant # 104199 to JGL)