ABSTRACT
Zfp318, a mouse gene with a Cys2/His2 zinc finger motif, is mainly expressed in germ cells in the testis. It encodes two alternative transcripts, which regulate androgen receptor-mediated transcriptional activation or repression by overexpression of them. However, the role of Zfp318 is still obscure in vivo, especially in spermatogenesis. To elucidate the role of Zfp318 during gamete production, we established a knockout mouse line. Zfp318-null male mice exhibited infertility, whereas Zfp318-null female mice displayed normal fertility. ZFP318 was expressed during multiple stages of spermatogenesis, from spermatocytes to round spermatids. The nuclei of secondary spermatocytes showed high levels of expression. Histological analysis and quantitative analysis of DNA content showed decreased numbers of both spermatids in the seminiferous tubules and mature spermatozoa in the epididymides of Zfp318-null mice. These results suggest that Zfp318 is expressed as a functional protein in testicular germ cells and plays an important role in meiosis during spermatogenesis.
Subject(s)
DNA-Binding Proteins/metabolism , Infertility, Male/genetics , Meiosis/genetics , Spermatogenesis/genetics , Zinc Fingers , Animals , DNA-Binding Proteins/genetics , Female , Infertility, Male/pathology , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Seminiferous Tubules/abnormalities , Seminiferous Tubules/metabolism , Spermatids/metabolism , Spermatids/pathology , Spermatocytes/metabolism , Spermatocytes/pathologyABSTRACT
Steroid hormones control the transcriptional activity of target genes mediated by intracellular nuclear receptors, and these transcriptional activities are modulated by the combination with coactivators and corepressors. We found in this study that testicular zinc finger protein (TZF) that was a nuclear protein with a zinc finger motif of the Cys2-His2 type was a novel corepressor of androgen receptor (AR). Fusion protein with green fluorescence protein GFP formed the specific foci in nuclei and TZF-dependent foci were located close to the splicing factor compartment. In addition, TZF was recruited into AR subnuclear foci after the treatment of dihydrotestosterone. Furthermore, we revealed that TZF bound to the activation function-1 (AF-1) domain (N-terminal transactivating domain) of AR protein. Transient over-expression of TZF in COS-7 cells or LNCaP human prostatic cancer cell resulted in decreased AR activity in a ligand-dependent fashion. Moreover, a transcriptional corepressor N-CoR additively decreased the transcriptional activity of AR with TZF. These findings suggest that TZF might be a novel corepressor of AR.
Subject(s)
Androgen Receptor Antagonists , Repressor Proteins/physiology , Zinc Fingers , Animals , COS Cells , Protein Binding , RNA Splicing , Receptors, Androgen/metabolism , Receptors, Androgen/physiology , Repressor Proteins/metabolism , Transcriptional Activation/physiologyABSTRACT
Testicular zinc finger protein (TZF) has a zinc finger motif of the Cys2-His2 type and its transcript is expressed predominantly in mouse spermatogenic cells. Using the fragment of TZF as a probe, we isolated the alternative splice variant form (TZF-L) from mouse testis cDNA library. Analysis of the open reading frame of each cDNA indicated that TZF and TZF-L were polypeptides of 942 and 2025 amino acid residues, respectively, and the N-terminal 902 amino acids of TZF-L were identical to those of TZF. The C-terminal region of TZF-L had more a zinc finger motif of the Cys2-His2 type and poly-Glu and poly-Pro regions. The mouse TZF/TZF-L gene spanned >20 kb and consisted of 11 exons. RT-PCR analysis of the expression level of mRNAs for mouse TZF and TZF-L showed that both transcripts are highly expressed in testis and moderately in kidney and ovary. Elevated expression of both transcripts during testicular development in mice was restricted to spermatocytes at the pachytene stage of meiotic prophase. Fusion proteins with GFP also demonstrated the nuclear localization of TZF and TZF-L. These experiments suggest that TZF and TZF-L may act to control the gene activity during spermatogenesis.