TY - JOUR T1 - [Effect of SUMO-1 on mitochondria subcellular localization of alpha-synuclein and its degradation via ubiquitin-proteasome system]. AU - Chen,Tao, AU - Liao,Xiao-ping, AU - Wen,Guo-qiang, AU - Nong,Zhi-gang, AU - Ouyang,Feng, AU - Deng,Yi-dong, AU - Guo,Min, AU - Wu,Hui-ling, AU - Zhou,Peng, PY - 2010/6/10/entrez PY - 2010/6/10/pubmed PY - 2010/9/18/medline SP - 267 EP - 71 JF - Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics JO - Zhonghua Yi Xue Yi Chuan Xue Za Zhi VL - 27 IS - 3 N2 - OBJECTIVE: To investigate the effect of sumoylation of alpha-synuclein by SUMO-1 on the mitochondria subcellular localization of alpha-synuclein and its degradation via ubiquitin-proteasome system. METHODS: Primers of wild-type, A53T pathogenic mutant and K96R mutant of human alpha-synuclein were designed to amplify the corresponding cDNAs without stop codon. The cDNAs were cloned into pGEM T-easy vector, analyzed by using enzyme mapping and DNA sequencing, and subcloned into pEGFP-N1 vector. The recombinant plasmids of pEGFP-alpha-synuclein-WT, pEGFP-alpha-synuclein-A53T and pEGFP-alpha-synuclein-K96R were transfected into HEK293 cells by lipofectamine method. The expression of the alpha-synuclein protein was measured by immunofluorescence and confocal microscope. Then mitochondria staining as well as immunofluorescence were utilized to investigate the effect of wild-type, A53T mutant and sumoylation of alpha-synuclein on mitochondria subcellular localization of alpha-synuclein. The effect of sumoylation of alpha-synuclein on its degradation via the ubiquitin-proteasome system in the cells was assayed by Western-blot. RESULTS: The enzyme mapping suggested that the eukaryotic expression plasmids for human wild-type, A53T and K96R mutants of the alpha-synuclein gene were constructed successfully. By immunofluorescence and confocal microscope, it was observed that alpha-synuclein-WT and alpha-synuclein-A53T proteins aggregated in cytoplasm, and alpha-synuclein-K96R protein aggregation was decreased in cytoplasm of cultured cells. The alpha-synuclein proteins of wild-type, A53T and K96R mutants were co-localized with mitochondria. Western-blot analysis revealed that both wild-type and A53T mutant affected the amount of the ubiquitinated proteins. CONCLUSION: Neither overexpression of wild-type and A53T pathogenic mutant alpha-synuclein, nor sumoylation of alpha-synuclein, affected the subcellular localization in the mitochondria. However, overexpression of wild-type and A53T mutant alpha-synuclein affected the amount of the ubiquitinated proteins. SN - 1003-9406 UR - https://www.unboundmedicine.com/medline/citation/20533263/[Effect_of_SUMO_1_on_mitochondria_subcellular_localization_of_alpha_synuclein_and_its_degradation_via_ubiquitin_proteasome_system]_ DB - PRIME DP - Unbound Medicine ER -