Post-transcriptional regulation of CLMP mRNA is controlled by tristetraprolin in response to TNFα via c-Jun N-terminal kinase signalling

Abstract

During spermatogenesis, extensive restructuring of blood–testis barrier takes place to facilitate the migration of preleptotene/leptotene spermatocytes from the basal to the adluminal compartment in the seminiferous epithelium. However, the biochemical mechanisms involved in this event remain elusive. Recent studies have shown that pro-inflammatory cytokine TNFα (tumour necrosis factor α) plays a crucial role in this event by inhibiting the expression of tight junction proteins in Sertoli cells. In the present study, we sought to examine the detailed mechanism on how TNFα affects the expression of CLMP (coxsackie- and adenovirus-receptor-like membrane protein), a newly identified tight junction transmembrane protein, in the testis. Addition of TNFα (10 ng/ml) to Sertoli cell culture (TM4 cells) significantly reduced the steady-state CLMP mRNA and protein levels. In an mRNA stability assay, it was shown that the rate of CLMP mRNA degradation was significantly increased when cells treated with TNFα were compared with vehicle. Blockage of the JNK (c-Jun N-terminal kinase) signalling pathway by SP600125 significantly abolished the TNFα-mediated destabilization of CLMP mRNA. Activation of the JNK signalling pathway by TNFα up-regulated the expression of an RNA-binding protein, TTP (tristetraprolin). TTP was present in the RNA–protein complex in the RNA EMSA (electrophoretic mobility shift assay) and decreased the CLMP 3′-UTR (untranslated region)-dependent luciferase activity. Taken together, these results suggest that the TNFα-mediated mRNA degradation of the CLMP gene is controlled by TTP through the JNK signalling cascade.