Function Analysis On Cellular Protein TIIMPSC And S24v2 Related To IFN-α-stimulating

Interferons(IFNs) possess the ability to defend against viral infection, regulate cell growth, and activateimmune proliferation. IFN-α, a typeâ… interferon, functions mainly in non-immune cells and inestablishment of antiviral state in cells. Studies performed to investigate the mechanism of antiviralactivity of IFN-αhave indicated that stimulation by IFN-αof cells is capable of inducing thetranscription of a significant number of genes and the synthesis of certain functional proteins incells. Investigation of such novel genes induced in cellular responses to IFN-αwill be useful for detailedstudy of the molecular mechanisms of IFN-αin cells.In the work described in this thesis, RNA differential display (RDD) technique was used to identifysome interesting cDNA gene fragments involved specifically in the response of human fibroblast toIFN-αtreatment in vitro. The results of RDD revealed that a few genes' expression were induced orinhibited in the response of human fibroblast to IFN-α. These genes included a typeâ…¡membraneprotein similar to CD69 (TIIMPSC, GenBank AB015628) with 697bp cds which is up-regulated byIFN-α, and an S24 variant 2(S24v2, which is a structural component of ribosome small subunit(GenBank NM.₀01026)with 593bp cds which is down-regulated by IFN-α. To analyze the functionof the coding proteins, two expression vector, pET-TIIMPSC and pGEX-S24v2 were constructed, andthe purified proteins were obtained as well. Subsequently, the anti-sera of above proteins were obtainedby immunizing the mice, and their specialty were verified by western blotting.Separately, the analysis on TIIMPSC focused on its character of trans-membrane protein.Immunofluorescence detection suggested that this protein is located on the surface of fibroblasts as aputative receptor. Cell proliferation assay revealed that activation of TIIMPSC improves fibroblastproliferation. Further, examination of signal transduction indicated that expression of this protein isup-regulated by IFN-αstimulation, and that it is involved in the regulation of fibroblast growth throughthe JAK-STAT signal pathway.The S24v2 is the component of the ribosome small subunit. In this case, IFN-αstimulation on humanfibroblast was shown to induced the inhibition of S24 variant 2 at mRNA level and protein level,implying a possible antiviral mechanism of IFN-αin human fibroblast. The delay of poliovirusreplication by IFN-αwas partially compensated for by S24v2 expressed in pcDNA vector transfected tocells, and the interference RNA of S24v2 was able to induce mimetically, in some extent, this poliovirus replication delay. These observations revealed that S24v2 could be involved in the antiviral effects ofIFN-αin human fibroblast.