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

Abstract

Sulfotransferase 4A1 (SULT4A1) is a sulfotransferase-like protein that is highly conserved between species. In human tissues, there are two transcripts, one that produces a full-length protein and one that produces an unstable truncated protein. The second transcript, which includes a pseudo-exon between exons 6 and 7 (6p), is widely expressed, whereas the first is more restricted. Differentiation of neuronal cells results in the removal of the pseudo-exon and subsequent SULT4A1 protein expression. Recent studies with SULT4A1 knockout mice showed that the protein is essential for normal development and that its absence leads to a severe neurologic phenotype. Here, the regulation of SULT4A1 6p splicing was investigated during neuronal differentiation using SH-SY5Y cells, human induced pluripotent stem cells, and mouse embryonic tissue. In all three models, pseudo-exon 6p was removed during differentiation, resulting in stable SULT4A1 protein expression. Using a minigene splicing assay, a region upstream of pseudo-exon 6p was identified that is essential for correct splicing of SULT4A1 mRNA. Within this region, there were binding motifs for four RNA processing factors (MBNL-1, MBNL-2, CELF-1, and CELF-2). Time-dependent changes in SULT4A1 protein and MBNL/CELF protein during differentiation supported their role in correctly splicing the SULT4A1 mRNA. Furthermore, ectopic expression of each factor produced efficient splicing in the minigene assay as well as correct splicing of the endogenous SULT4A1 mRNA. These results show that SULT4A1 mRNA is a target for MBNL/CELFdependent splicing, which may be essential in producing stable, functional SULT4A1.

SULT4A1 Modulates Synaptic Development and Function by Promoting the Formation of PSD-95/NMDAR Complex

“…In mouse, Sult4a1 mRNA expression is low in fetal brains and remains nearly unchanged until postnatal day 30, after which a marked increase in expression has been observed (Alnouti et al, 2006). Consistent with this, SULT4A1 protein expression was found to increase during neuronal differentiation (Idris et al, 2019). Notably, Sult4a1-KO mice display severe neurological defects including tremor, rigidity and seizure and the survival of pups during postnatal development is strongly affected by loss of Sult4a1 (Garcia et al, 2018).…”

“…However, the functional role of SULT4A1 within neuronal development is largely unknown since no substrate nor biological functions have been identified yet. SULT4A1 mRNA and protein expression were found to increase during mouse neuron and brain development (Alnouti et al, 2006;Hashiguchi et al, 2018;Idris et al, 2019), supporting a role for SULT4A1 in neuronal maturation. Notably, we found that SULT4A1 is not only express in cytosol, mitochondrial and microsomal fractions (Garcia et al, 2018), but is also highly expressed in synaptosomal preparation, obtained from mouse cortical neurons (Fig.…”