| Summary: | 碩士 === 中山醫學大學 === 生物醫學科學學系碩士班 === 95 === The homeobox gene products act as transcription factors during animal developmental process.These genes encode homeodomain proteins that play a foundamental role in cellular differentiations and determination of cell fate. We performed a degenerated oligonucleotide polymerase chain reaction (PCR) to screen a mouse embryonic stem (ES) cell cDNA library and isolated several novel homeobox-containing genes. One of these genes, which we detected expressions in ES cells and ovary named as Eso-3, encodes a homeodomain protein that has the highest amino acid sequence identity (58%) to Iroquois subclass homeodomain gene product. That three extra amino acid residues, proline-tyrosine-proline, exist in the loop between alpha helixⅠand helix Ⅱ of the homeodomain, suggests that Eso-3 is a new member of TALE class homeobox genes. By a combination of techniques including rapid amplification of cDNA ends and EST database searching, we found that Eso-3 has 3 different transcripts and 2 different open reading frames. From the results of reverse transcription (RT)-PCR, we identified that Eso-3 expressed in all tested mouse tissues and predominantly expressed in ovary and brain. Results of RT-PCR also revealed that Eso-3 expressions in ovaries started at embryonic stage day 12.5 and continuing to the adult stage. The Eso-3 expressions in ovary were gradually increased during ovary development. Results of in situ hybridization showed that Eso-3 expressions in adult ovary were detected in the somatic cells of follicles, but not in the theca cells. In testis, results of RT-PCR revealed that Eso-3 expressions were gradually decreased from embryonic stage day 12.5 to the adult stage. Results of in situ hybridization showed that Eso-3 expressions were detected in sertoli cells and some spermatocytes of adult testis. Based on our finding, we suggest that Eso-3 play a role during development of the reproduction system. Furthermore, we generated gene knockdown transgenic mice expressing Eso-3 small interference RNA (siEso-3) to inhibit Eso-3 protein expressions in mice for investigating the in vivo functions of Eso-3 protein. Reduced expressions of Eso-3 protein levels in brain of TG23 siEso-3 transgenic mouse were observed by western analysis. The statistic results showed a significant decrease of fertility and/or inheritance in siEso-3 transgenic mice than wild-type mice. The relationship between knockdown of Eso-3 expression and mouse fertility and/or inheritance will need further analyses.
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