Identification of target genes for spermatogenic cell-specific KRAB transcription factor ZFP819 in a male germ cell line

Sora Jin,Juri Jeong,Chunghee Cho,Jun Tae Kwon,Jaehwan Kim,Jihye Kim,Seong Hyeon Hong,Heejin Choi

doi:10.1186/s13578-016-0132-4

Abstract

BackgroundZfp819, a member of the Krüppel-associated box (KRAB) family, encodes a spermatogenic cell-specific transcription factor. Zfp819-overexpression induces apoptosis and inhibits proliferation in somatic cell lines.ResultsIn the present study, we examined the cellular effects of Zfp819 in a male germ cell line (GC-2 cells). Overexpression of Zfp819 demonstrated an increase in the number of apoptotic cells, leading to inhibition of proliferation in GC-2 cells. We further investigated genes regulated by ZFP819 using microarray analysis and chromatin-immunoprecipitation combined with microarray analysis (ChIP-chip) in GC-2 cells. We identified 118 downregulated genes in Zfp819-overexpressing GC-2 cells using microarray analysis. ChIP-chip assay revealed that 1011 promoter sites (corresponding to 262 genes) were specifically enriched in GC-2 cells transfected with Zfp819. Two genes (trinucleotide repeat containing 6b and annexin A11) were commonly found when we compared the data between microarray and ChIP-chip analyses. Consistent with these results, Zfp819 overexpression significantly reduced the transcript levels of the two genes by binding to their promoter regions. Tissue distribution analysis indicated that both genes were predominantly expressed in testis. It has been reported that these two genes function in apoptosis.ConclusionCollectively, our study provides inclusive information on germ cell-specific gene regulation by ZFP819, which is involved in apoptosis, to maintain the integrity of spermatogenesis.

Highlights

Zfp819, a member of the Krüppel-associated box (KRAB) family, encodes a spermatogenic cell-specific transcription factor
Repressive activity of Zfp819 in germ cell line (GC‐2 cell) We previously reported that the KRAB domain of Zfp819 has repressive activity using the GAL4-UAS system in somatic cell lines (HEK 293T and NIH3T3) [5]
To confirm the repressive activity of Zfp819 in the germ cell line as well, we performed a reporter assay using GAL4-DBD-Myc constructs fused to the KRAB domain sequences from Zfp819 and a luciferase expression construct containing an enhancer that bound to GAL4 (GAL4-UAS) (Fig. 1a)

Summary

Introduction

Zfp819, a member of the Krüppel-associated box (KRAB) family, encodes a spermatogenic cell-specific transcription factor. Spermatogenesis is a unique and highly elaborated process that requires a highly organized and tightly regulated network of genes, many of which are spermatogenic-cellspecific. The Krüppel-associated box and C2H2-type zinc finger motifs (KRAB-ZF) family, one of the largest transcription factor groups in vertebrates has a transcriptional repressive activity by binding to DNA in a sequence-specific manner [3]. The members of KRABZF family are generally expressed in various tissues and are involved in diverse processes including apoptosis, cell proliferation, and tumorigenesis [3]. It has been reported that a number of the KRAB-ZF genes on a certain chromosome are abundantly expressed in testis and fetal brain and may have important functions in these tissues [4].

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