Abstract
Calretinin, a Ca2+-binding protein, participates in many cellular events. Our previous studies found the high expression of calretinin in testicular Leydig cells. In this study, (MLTC-1 cells were infected with LV-calb2, R2C cells with LV-siRNA-calb2. The primary mouse Leydig cells were also used to confirm those data from cell lines. Testosterone level was significantly higher in the MLTC-1 cells with over-expressed calretinin than in the control, while progesterone was lower in the R2C cells in which down-regulated calretinin. The expressions of StAR changed in synchrony with hormones. Cytoplasmic Ca2+ level was significantly increased when calretinin was over-expressed. When MLTC-1 cells were infected with LV-calb2 and then stimulated using Clopiazonic, a Ca2+-releasing agent, testosterone was significantly increased. Interestingly, the expression levels of PLC, p-PKCµ (PKD), p-MARCKS and CREB, were significantly increased in the MLTC-1 cells with over-expressed calretinin, while PLC, p-PKD, p-MARCKS, MARCKS and CREB were decreased in the R2C cells with down-regulated calretinin. We also observed the increased expression of calretinin up-regulated testosterone production and the expressions of StAR and PLC in primary mouse Leydig cells. So, calretinin as a Ca2+-binding protein participates in the regulation of steroidogenesis via the PLC-Ca2+-PKC pathway in Leydig cells.
Highlights
The main functions of testis are spermatogenesis and androgen production
Our preliminary studies showed that adult rats expressed highest calretinin in the cytoplasm of Leydig cells when compared with preadolescent or old rats, which is in accord with the androgen level, suggesting that calretinin may prompt the steroidogenesis by Ca2+-related pathway in Leydig cells[17]
The main raw material of androgen synthesis is cholesterol, which from either the ester of plasma lipoproteins, or directly from the extracellular cholesterol into Leydig cells cytoplasm through scavenger receptor (SR-B1)
Summary
The main functions of testis are spermatogenesis and androgen production. Androgenesis is completed in testicular Leydig cells. The protein kinase A (PKA) is activated by cAMP, phosphorylating those steroidogenetic enzymes or activating transcription factors to promote the expression of steroidogenetic enzymes, which enhances the synthesis of testosterone. This is the so-called classic LH-PKA signal pathway[3]. The cytoplasmic calcium ions are increased together with DAG, and activate protein kinase C (PKC), by which phosphorylates steroidogenetic enzymes or activates transcription factors to promote the expression of steroidogenetic enzymes, such as the transcription factors CREB and STAR, leading to the promotion of testosterone synthesis[4]. Our preliminary studies showed that adult rats expressed highest calretinin in the cytoplasm of Leydig cells when compared with preadolescent or old rats, which is in accord with the androgen level, suggesting that calretinin may prompt the steroidogenesis by Ca2+-related pathway in Leydig cells[17]
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