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Tribbles Homolog 2 (TRIB2) plays a potent role in ovarian granulosa cells proliferation and function
Author(s) -
Warma Aly,
Lussier Jacques G.,
Ndiaye Kalidou
Publication year - 2019
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2019.33.1_supplement.476.33
Subject(s) - follicular phase , estrous cycle , corpus luteum , biology , medicine , endocrinology , theca , messenger rna , granulosa cell , ovarian follicle , ovary , andrology , follicular fluid , oocyte , microbiology and biotechnology , gene , embryo , biochemistry
Tribbles homolog (TRIB) 1, 2 and 3 represent atypical members of the serine/threonine kinase superfamily and are homologs of Drosophila tribbles. TRIB2 mRNA is rapidly induced by mitogens, has a short half‐life, and is expressed in a cell‐specific manner. We previously identified TRIB2 as a differentially expressed gene in granulosa cells of bovine preovulatory follicles. This study aimed to further investigate TRIB2 mRNA and protein regulation, to identify its binding partners, and study its function in granulosa cells of bovine dominant follicles. Granulosa cells (GC) were obtained from follicles at different developmental stages: small follicles (SF: 2–4 mm), dominant follicles (DF) at day 5 of the estrous cycle (day 0 = day of the oestrous), ovulatory follicles 24 hours following injection of an ovulatory dose of hCG (OF), and corpus luteum (CL) at day 5 of the oestrous cycle. In addition to this in vivo model, an in vitro model of cultured GC was used for functional studies using the CRISPR‐Cas9 approach. RT‐qPCR analyses showed greatest expression of TRIB2 in GC of DF, while the weakest expression was in OF and CL (P < 0.0001). Temporal expression of TRIB2 mRNA was further studied in follicular walls (granulosa and theca cells) obtained from ovulatory follicles recovered at 0, 6, 12, 18 and 24 hours after hCG injection. There was a significant reduction of TRIB2 steady‐state mRNA levels in follicular walls, starting at 6 hours through 24 hours post‐hCG as compared to 0 hour (P < 0.001). Additionally, we have generated specific anti‐TRIB2 polyclonal antibodies that confirmed, in western blot analyses, TRIB2 downregulation by hCG at the protein level. In vitro studies showed that FSH stimulates TRIB2 expression (P < 0.05) while inhibition of TRIB2 using CRISPR‐Cas9 resulted in significantly reduced GC proliferation (P < 0.05). These results provide strong evidence that TRIB2 is a potent regulator of GC proliferation. Support or Funding Information Research supported by NSERC of Canada grant #RGPIN‐2018‐04516 to KN. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .