Regulation And Function Of Neural Salient Serine/Arginine Rich Protein1 (NSSR1) In Neural Differentiation

The main focus of the present study is to investigate the regulation and function ofalternative pre-mRNA splicing (APS) factors in neural differentiation. In order to analyzethe regulation of APS factors in neural differentiation, we established the methods toisolate, culture and induce the differentiation of mouse primary neural stem cells (NSCs).Then, we examined the expression of several known APS factors (Tra2β, NSSR1,hnRNP-R, Zis and authentic SR proteins) in NSCs. The results show: 1). No differencewas observed for the expression of hnRNP-R between the NSCs derived from variousresources, such as either from the cortex (cNSC) or spinal cord (spNSC). 2). Theexpression of Zis is not altered by differentiation of NSC. However, Zis proteins in cspNPC show higher molecular weights than in NSC, possibly due to the differences in cthe isoforms or phosphorylation status between two tissues. 3). The differentiationdown-regulates the expression of Tra2β. In addition, Tra2β shows a lower expression inspNSC compared to in NSC 4). In contrast to the expression of Tra2β, NSSR1 is cup-regulated following the NPC differentiation. In addition, 1H4 antibodies detected nosignals for any phosphorylated authentic SR protein in NSC, while in the differentiatedmature neural cells there are two bands detected with molecular weights of 22-30 kDaand 43-50 kDa. These bands were also detected in mouse uterus tissue and are similar toproteins detected in the neuronal differentiated P19 cells. It is possible that they mightrepresent the newly characterized hypophosphorylated SR proteins of NSSR1 andNSSR2. 4复旦大学博士学位论文NSSR1 在神经分化中的表达调控及功能研究 英文摘要 NSSR1 is the only SR protein that is silently expressed in nervous system. Theup-regulation of NSSR1 expression following the maturing of neural cells indicates thefunctional importance of NSSR1 during neural differentiation. Therefore, in thefollowing studies, we focused on the regulation and function of NSSR1 during neuraldevelopment or differentiation. Previous studies have shown that NSSR1 transcripts are saliently expressed inneural and testis tissues in mice, while widely expressed in various tissues in humans. Itseems that the expression of NSSR1 varies depending on the species. In the present study,we systemically examined the expression of NSSR1 mRNA and proteins in mouse andhuman tissues. The results using RT-PCR analysis demonstrated that NSSR1 mRNA iswidely expressed in various tissues in both species. To examine the expression of NSSR1proteins, we prepared the antibodies specific to NSSR1 proteins. Western-blot analysisshowed that NSSR1 proteins are expressed only in neural tissues and testis either in bothspecies. The data convince that NSSR1 is a neural specific SR protein either in human orin mice. To begin to study the potential function of NSSR1, we then appliedimmunohistochemical analysis to examine the distribution of NSSR1 proteins in mousebrain and found that NSSR1 proteins are distributed in the whole brain, with the highestexpression in granule cells of the hippocampus, neurons in the cerebrum and Purkinjecells in the cerebellum. In addition, ICC (Immunocytochemistry) analysis in culturedmouse glial cell showed that the distribution of NSSR1 proteins was cell cycle dependent.NSSR1 proteins are mainly distributed in the cytosol around the nuclei in the splittingcells, while restricted in the nuclei in non-splitting cells. It suggests that distributionpattern of NSSR1 proteins be regulated by cell cycles. 5复旦大学博士学位论文NSSR1 在神经分化中的表达调控及?...