Genomic organization and structure of the 5′‐flanking region of the TEX101 gene: Alternative promoter usage and splicing generate transcript variants with distinct 5′‐untranslated region

Abstract

A novel germ cell-specific antigen, TEX101 (TES101-reactive protein), was previously identified using a monoclonal antibody directed against mouse testicular cells. TEX101 is specifically located on the plasma membrane of germ cells, and its expression in gonadal organs is sexually dimorphic. To understand the fundamental mechanism directing gene expression, the genomic organization of TEX101 was studied. The gene consists of five translated exons (exons 2-6) and three 5'-untranslated exons (exon 1a, 1b, and 1c), respectively. TEX101 forms three major transcripts classified by usage of the three 5'-untranslated exons. One form of TEX101 mRNA is transcribed from exon 1c and spliced to the common acceptor site in exon 2. In the second form of the transcript, exon 1a is spliced to exon 1b and exon 2 in a sequential manner. Splicing from exon 1a to exon 2, arises the third form of transcript. Reverse Transcription (RT)-polymerase chain reaction (PCR) analysis demonstrated differential expression pattern of the TEX101 transcripts between testis and ovary. Whereas the expression of transcript-1 is constitutive in male and female gonads, the transcript-2 and -3 are detected only after starting of the spermatogenesis. Luciferase reporter assays using GC-2spd(ts) cells, a cell line from immortalized mouse testicular cells, showed that the 5'-flanking sequence of exon 1c has higher promoter activity than exon 1a. Deletion analysis of the chimeric structures indicated that sequences essential to gene expression are present on the 5'-flanking region between -3186 and +14, where the cluster of five CAAT boxes is located. Taken together, these findings should facilitate an understanding of the regulation of TEX101 expression during gametogenesis.