Study On Development/Spermatogenensis Related Genes In Human Testis

Human testis is a specific organ. There are two fundamental functions in human tetstis: androgen secretion and spermatogenesis. Spermatogenesis is a complex and very efficient process that begins with the division and differentiation of the spermatogenic stem cell within the seminiferous tubule of the testis. Because meiosis is different from mitosis, the expression of genes is different between testis and other organs. Also, spermatogenesis begins in puberty, so the expression of genes is different in different stages, for example in adult and fetal testis. Study of genes specifically expressed in the testis at different stages of development may reveal new genes related to the function of testis, especially spermatogenesis or find new function for reported genes. In our laboratory, human testis cDNA microarrays were used to identify the genes specifically expressed in adult or fetal testes and 1522 differentially expressed clones were found. Among these clones, 499 were unique genes. In this study, three of these genes (Rtn-T, NYD-TSP1 and NYD-SP6) were selected for functional study. Part 1. Study on Rtn-T geneReticulon (Rtn) is a gene family. All Rtn proteins show a 200-amino-acid residue region of sequence similarity at the C terminus of the protein. Related sequences have been recognized in human, bovine, mouse, rat, fly and worm genomes. It has been reported that Rtn is related to axon extention and apoptosis. The relationship between Rtn and testis function has not been reported. In this study, by using human testis cDNA microarray, a new Rtn like gene was found. It was named as Rtn-T for its specific expression in testis. The hybridization intensity of Rtn-T in adult and fetal testes was 180.64 and 50.15, respectively. The full length cDNA of Rtn-T was 3491 bp and encodes a 108 kDa protein of 986 amino acids. This Rtn-T has Genbank accession number AF333336. Blast search in the human genome datebase showed that Rtn-T consisted of 9 exons. Blast search of the contig map showed that Rtn-T was mapped to chromosome 2pl6. Analysis of the amino acid sequence revealed one signal sequence of 26 amino acids at theN-terminal and two long hydrophobic stretches of 37 and 35 amino acids at the C-terminal. Blast searches revealed that Rtn-T was highly homologous to six other genes. All of them belong to the Rtn family and come from humans. PCR showed that Rtn-T was particularly and strongly expressed in human testis. These results indicate that Rtn-T may be related to testis development, especially process of spermatogenesis. Whether Rtn-T influence apoptosis, and thus affects the process of spermatogenesis should be determined. Part 2. Study on NYD-TSP1Spermatogenensis involves the expression of many genes. The expression of these genes is promoted by testis-expressed transcription factors at the transcription level. One of the major classes of growth and development regulatory proteins consists of the basic helix-loop-helix (bHLH) motif-containing proteins. In this study, by using human testis cDNA microarray, a novel human testis specific gene, NYD-TSP1, was identified. NYD-TSP1 encodes a putative bHLH-Zip containing protein and is highly and specifically expressed in adult human testis. The level of hybridization intensity of NYD-TSP1 in adult and fetal testis was 134.27 and 22.53 respectively. The full length of NYD-TSP1 cDNA was 1848 bp and encoded a 49.4 KDa protein of 430 amino acids. The cDNA sequence of this clone was deposited with Genbank. The accession number was AF333098. NYD-TSP1 was also determined as Interim Gen Symbol (Unigene). The name is NYD-TSP1: testis-specific protein NYD-TSP1. UniGene number is Hs.98266. Predicted protein analysis showed that NYD-TSP1 contained two functional dormains: an N-terminal basic helix-loop-helix (bHLH) and a C-terminal Zip. Homologous analysis showed that the 430 amino acid sequences had sequence homology with Spz 1 protein of mouse. The positive rate of homology for full sequences was 58%. In bHLH and Zip motif the positive rate of homology was 59.