Expression of the Orphan Cytosolic Sulfotransferase SULT4A1 and Its Major Splice Variant in Human Tissues and Cells: Dimerization, Degradation and Polyubiquitination

Abstract

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.

The MBNL/CELF Splicing Factors Regulate Cytosolic Sulfotransferase 4A1 Protein Expression during Cell Differentiation

“…Two different transcripts have been reported, one that includes a pseudo-exon between exons 6 and 7 (6p), termed the variant transcript, and one where 6p has been removed, termed the wild-type (WT) transcript. The variant transcript introduces a premature stop codon resulting in an unstable, truncated protein (Sidharthan et al, 2014). In human tissues, SULT4A1 protein expression correlates well with the WT transcript, consistent with the notion that the variant transcript does not produce measurable protein levels (Sidharthan et al, 2014).…”

“…The variant transcript introduces a premature stop codon resulting in an unstable, truncated protein (Sidharthan et al, 2014). In human tissues, SULT4A1 protein expression correlates well with the WT transcript, consistent with the notion that the variant transcript does not produce measurable protein levels (Sidharthan et al, 2014). Thus, alternative splicing appears to be an important mechanism for regulating SULT4A1 expression.…”

“…In both neuroblastoma SH-SY5Y and SK-N-MC cells, the switch from the variant to the WT transcript is seen during cellular differentiation (Sidharthan et al, 2014). This alternative splicing is also observed over time in cultured neuron-glia mixed cells (Hashiguchi et al, 2018).…”

“…This important work suggests that SULT4A1 is fundamental to normal neuronal development. Second, two reports independently showed that SULT4A1 protein expression is dependent on alternative splicing of its transcript (Sidharthan et al, 2014;Hashiguchi et al, 2018). Whereas the SULT4A1 protein is mostly confined to neuronal tissue, its mRNA is more widely expressed (Falany et al, 2000;Sidharthan et al, 2014).…”

“…Second, two reports independently showed that SULT4A1 protein expression is dependent on alternative splicing of its transcript (Sidharthan et al, 2014;Hashiguchi et al, 2018). Whereas the SULT4A1 protein is mostly confined to neuronal tissue, its mRNA is more widely expressed (Falany et al, 2000;Sidharthan et al, 2014). Two different transcripts have been reported, one that includes a pseudo-exon between exons 6 and 7 (6p), termed the variant transcript, and one where 6p has been removed, termed the wild-type (WT) transcript.…”