The Role of Zinc Finger Protein-1 Gene in Spermatogenesis of Male Mouse
Date Issued
2004
Date
2004
Author(s)
Tsai, Pei-Chun
DOI
zh-TW
Abstract
The purpose of this study was to clarify the molecular mechanisms of mouse zinc finger protein-1 (mZFG-1) gene potentially involved in the regulation of spermatogenesis within the mouse testis. To meet this purpose, an initial attempt was made to generate transgenic (Tg) mice characterized by conditional knock-down of the mZFG-1 gene. Of these studies, a transgene named anti-mZFG-1L/pIND/V5-His-TOPO, containing antisense transcript against to nucleotide sequences complementary to the long form of mZFG-1 cDNA, was constructed. Generation of Tg mice was conducted by co-injection transgenes of anti-mZFG-1L/pIND/V5-His-TOPO and pVgRXR, into the pronucleus of newly fertilized eggs. These have resulted in 10 out of 66 newborn mice to be transgenic and four out of these 10 Tg mice were confirmed, as evidences shown by PCR and Southern-blot analyses, to be harboring both of the transgenes said above. Each of the four double-transgenic founder mice appeared to be fertile with 16.7 ~ 100% of their transgenes being germline-transmitted to the F1 progenies obtained. Evidences from RT-PCR analyses further confirmed that, in comparison with those of the control mice, much less of endogenous mZFG-1 mRNA was found in the double-transgenic founder mice when they had been subjected to the ponasterone-A induction. While no gross anomalies was found in any somatic tissues from those ponasterone-A induced double-transgenic mice, much less number of mature spermatozoa were found in testis-specimens from the double-transgenic mice when comparison was made to those of control mice after ponasterone-A induction.
Moreover, the effect of mZFG-1 on spermatogenesis at molecular level was further verified using mouse Sertoli cell lines (TM4 cell lines) with the specific small interferon RNA (siRNA) strategy for knock-down the expression of endogenous mZFG-1 and the consequent expression profiles of its down-stream genes were investigated. Based on results from RT-PCR, two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and mass spectrometry (MS) analyses appeared that both Crisp-1 (Cysteine-rich secretory protein-1) and NPY (Neuropeptide Y) genes were down-regulated within Sertoli cells when they had been transfected with siRNA designed against to mZFG-1 gene.
Conclusions came to these present studies were that appropriate expression of mZFG-1 gene do play its physiological significances for ensuring normal spermatogenesis occurred within the testes and the potential effect of mZFG-1 gene on spermatogenesis may be mediated by modulation the expression of both Crisp-1 and NPY genes.
Subjects
鋅指蛋白
小鼠
生精作用
mouse
spermatogenesis
zinc finger
Type
thesis
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