中国生物工程杂志

Calsenilin/KChIP3/DREAM, which was originally identified as a presenilin and calcium binding protein expressed at higher levels in nerve system, plays multiple funtional roles as a transcriptional repressor in the nucleus. It binds to the downstream regulatory element (DRE) of prodynorphin or c-fos and its transcriptional repression is modulated by intracellular calcium release. As a voltage-gated potassium (Kv) channels interaction proteins, there are four KchIPs isoform, KChIP1 expressed in variant tissues, KChIP2 specific expressed in heart, KChIP3 and KChIP4 high expressed in brain. They are high homogoly sequence in C- terminus but difference in N- terminus. Because of lacking of the antibody for specific recognize the N-terminus i of calsenilin, it is not clear that the funcrional differents among the KChIP3 family. In other hand, Calsenilin is phosphorylated by CKI（casein kinase I）, PKC, PKA. The multiple phosphorylaton sites containing Ser62, Ser63, and Ser65. Importantly, phosphorylation of the Ser63 oppose its degradation by caspase-3. Calsenilin also called KChIP3 and DREAM appears to process multiple functions in variant tissues by interaction with different moulecules and regulates gene expression, such as depressor the prodynorphin , c-fos , CREB, GnRH , expression in response to calcium influx; regulation of cytokine expression in T-cell proliferation. On the other hand, calsenilin as a activator promote the Vitamin D and retinoic acid response. Thus, calsenilin/ KChIP3/ DREAM as a dual transcriptional regulator on different target molecules. To understaning the functional mechanism of calsenilin/KChIP3/DREAM in aging and oxidative stress response, the functional role in the phosphorylation of calsenilin and relationship between transcriptional repression and phosphorylation of calsenilin in nerve system, the monoclonal antibody gainst human calsenilin was generated by tranditional methods: the human calsenilin was cloned into pGEX-4T-2 vector and expressed by IPTG induction. The purified GST-calsenilin fusion protein was used to immunize mice. PEG-1500 reagent was used to fuse cells. Confocal micrograph results indicate that the monoclonal antibody against calsenilin workes well for Immuno- histochemical analysis, so the overexpressed calsenilin was able to observed in ER/ Golgi complex of H4 neuroglioma cells. Western blot analysis confirmed the monoclonal antibody against human calsenilin reacted on overexpressed protein calsenilin. Ho