MEF2 and NR2F2 cooperate to regulate Akr1c14 gene expression in mouse MA‐10 Leydig cells

Abstract

SUMMARYLeydig cells are essential for male reproductive development and health throughout life. Production of androgens [testosterone, dihydrotestosterone (DHT)] as well as intermediate steroids [progesterone, dihydroprogesterone (DHP)] is tightly regulated. In the mouse, the 3a-hydroxysteroid dehydrogenase enzyme (3a-HSD, AKR1C14) catalyses the interconversion of DHP and DHT into less potent steroids. Despite its importance, nothing is currently known regarding the regulation of Akr1c14 expression in Leydig cells. Recently, the transcription factors MEF2 and NR2F2 were identified in the mouse testis including in Leydig cells where they were found to regulate expression of genes involved in steroidogenesis. Analyses of transcriptomic data from MEF2-or NR2F2-deficient MA-10 Leydig cells revealed a significant decrease in Akr1c14 mRNA levels. Using qPCR, we confirmed that Akr1c14 mRNA levels were decreased in MEF2-and in NR2F2-deficient conditions. Conversely, overexpression of MEF2A or/and NR2F2 in MA-10 Leydig cells led to an increase in endogenous Akr1c14 mRNA levels. Recruitment of MEF2 and NR2F2 to the Akr1c14 promoter was confirmed by ChIP while DNA precipitation assays revealed direct binding of MEF2 but not NR2F2 to this region. In functional promoter studies, NR2F2 was found to activate the Akr1c14 promoter while MEF2A on its own had no effect. Combination of both NR2F2 and MEF2A led to a cooperative activation of the Akr1c14 promoter and this required intact MEF2 and NR2F2 elements. Finally, coimmunoprecipitation experiments showed that MEF2 and NR2F2 are present in the same protein complex. In conclusion, our results identify a novel cooperation between MEF2 factors and NR2F2 in the expression of the Akr1c14 gene involved in the regulation of DHP/DHT levels.

Two distinct pathways of pregranulosa cell differentiation support follicle formation in the mouse ovary

“…4 B ). These included the ketogenic gene Hmgcs2 ( 70 ), the hydroxysteroid dehydrogenase gene Ark1c14 ( 71 ), and multiple others ( Fig. 4 B ).…”

“…The rate of Drosophila ovarian follicle development can be strongly modulated by nutrition, as reflected in insulin signaling (76), and mammalian follicular growth is additionally influenced by activin/inhibin and steroid signaling (77,78). BPG cells consistently express more of the androgen-degrading enzyme 3α-hydroxysteroid dehydrogenase encoded by Akr1c14 (71), which may help to sustain female development and promote direct development of wave 1 follicles. Akr1c14 is expressed 8 to 20 times higher in BPG cells than in EPG cells from at least E12.5 to E18.5.…”

Transcription Factors in the Regulation of Leydig Cell Gene Expression and Function

“…It cooperates with SF1 on the Star and Insl3 promoters ( 24 , 25 ) and with SP1 on the Amhr2 promoter ( 26 ). The Akr1c14 gene, which codes for the 3α-HSD enzyme that catalyzes the interconversion of dihydrotestosterone (DHT) into 5α-androstane-3α,17β-diol (3α-diol), is activated by COUP-TFII in cooperation with MEF2 ( 27 ). COUP-TFII also activates the expression of Gsta3 and Inha , genes involved in the inactivation of reactive oxygen species and in the homeostasis of the hypothalamic-pituitary-gonadal axis, respectively ( 28 ).…”

Gene expression profiling of cultured mouse testis fragments treated with ethinylestradiol

“…This in vitro mouse testis organ culture system was limited to 20 days of culture because of germ cells degeneration in the seminiferous tubules. When histological evaluation of testis fragments is not performed using enough seminiferous tubules with germ cell differentiation or the criteria of histological evaluation is not consistent, these gene expression Zhou et al, 2008;Tan et al, 2005;Schrimpf et al, 2014;Hogarth et al, 2015;Wu et al, 2017;Wang et al, 2015a;2015b;Sperm flagellum Dnah7a↓;Luzp4↑;Spef2↓ Neesen et al, 1997;Sironen et al, 2010Sironen et al, , 2011 Ge et al, 1999;Di-Luoffo et al, 2016 Estrogen responsive genes/transcripts Rad9a ↑; Arnt2 ↑; Akr1c14↑; Greb1 ↑ Sun et al, 2007;Kretzschimar et al, 2010;Karlsson et al, profiles may be useful in determining testicular toxicity in vitro. Plcz1 and Tssk4 genes can be used for examining the effects of testicular toxicity to germ cell differentiation; Cyp26b1 gene will be used for testicular cell survival; and Il16 gene will be used for immune response.…”