Sidharthan Neelima P, Butcher Neville J, Mitchell Deanne J, Minchin Rodney F
Laboratory for Molecular and Cellular Pharmacology, School of Biomedical Sciences, University of Queensland, Brisbane, Australia.
PLoS One. 2014 Jul 2;9(7):e101520. doi: 10.1371/journal.pone.0101520. eCollection 2014.
The cytosolic sulfotransferase SULT4A1 is highly conserved between mammalian species but its function remains unknown. Polymorphisms in the SULT4A1 gene have been linked to susceptibility to schizophrenia. There are 2 major SULT4A1 transcripts in humans, one that encodes full length protein (wild-type) and one that encodes a truncated protein (variant). Here, we investigated the expression of SULT4A1 in human tissues by RT-PCR and found the wild-type mRNA to be expressed mainly in the brain, gastrointestinal tract and prostate while the splice variant was more widely expressed. In human cell-lines, the wild-type transcript was found in neuronal cells, but the variant transcript was expressed in nearly all other lines examined. Western blot analysis only identified SULT4A1 protein in cells that expressed the wild-type mRNA. No variant protein was detected in cells that expressed the variant mRNA. Ectopically expressed full length SULT4A1 protein was stable while the truncated protein was not, having a half-life of approximately 3 hr. SULT4A1 was also shown to homodimerize, consistent with other SULTs that contain the consensus dimerization motif. Mutation of the dimerization motif resulted in a monomeric form of SULT4A1 that was rapidly degraded by polyubiquitination on the lysine located within the dimerization motif. These results show that SULT4A1 is widely expressed in human tissues, but mostly as a splice variant that produces a rapidly degraded protein. Dimerization protects the protein from degradation. Since many other cytosolic sulfotransferases possess the conserved lysine within the dimerization motif, homodimerization may serve, in part, to stabilize these enzymes in vivo.
胞质磺基转移酶SULT4A1在哺乳动物物种间高度保守,但其功能尚不清楚。SULT4A1基因的多态性与精神分裂症易感性相关。人类中有两种主要的SULT4A1转录本,一种编码全长蛋白(野生型),另一种编码截短蛋白(变体)。在此,我们通过逆转录聚合酶链反应(RT-PCR)研究了SULT4A1在人体组织中的表达,发现野生型mRNA主要在脑、胃肠道和前列腺中表达,而剪接变体的表达更为广泛。在人类细胞系中,野生型转录本在神经元细胞中被发现,但变体转录本在几乎所有检测的其他细胞系中均有表达。蛋白质印迹分析仅在表达野生型mRNA的细胞中鉴定出SULT4A1蛋白。在表达变体mRNA的细胞中未检测到变体蛋白。异位表达的全长SULT4A1蛋白稳定,而截短蛋白不稳定,半衰期约为3小时。SULT4A1还显示能形成同源二聚体,这与其他含有共有二聚化基序的磺基转移酶一致。二聚化基序的突变导致SULT4A1的单体形式,该单体形式在二聚化基序内的赖氨酸上通过多聚泛素化迅速降解。这些结果表明,SULT4A1在人体组织中广泛表达,但大多以产生快速降解蛋白的剪接变体形式存在。二聚化可保护蛋白质不被降解。由于许多其他胞质磺基转移酶在二聚化基序内具有保守的赖氨酸,同源二聚化可能部分用于在体内稳定这些酶。