An F-Box Protein, Mdm30, Interacts with TREX Subunit Sub2 To Regulate Cellular Abundance Cotranscriptionally in Orchestrating mRNA Export Independently of Splicing and Mitochondrial Function

Abstract

Although an F-box protein, Mdm30, is found to regulate ubiquitylation of the Sub2 component of TREX (transcription-export) complex for proteasomal degradation in stimulation of mRNA export, it remains unknown whether such ubiquitin-proteasome system (UPS) regulation of Sub2 occurs cotranscriptionally via its interaction with Mdm30. Further, it is unclear whether impaired UPS regulation of Sub2 in the absence of Mdm30 alters mRNA export via splicing defects of export factors and/or mitochondrial dynamics/function, since Sub2 controls mRNA splicing and Mdm30 regulates mitochondrial aggregation. Here, we show that Mdm30 interacts with Sub2, and temporary shutdown of Mdm30 enhances Sub2’s abundance and impairs mRNA export. Likewise, Sub2’s abundance is increased following transcriptional inhibition. These results support Mdm30’s direct role in regulation of Sub2’s cellular abundance in a transcription-dependent manner. Consistently, the chromatin-bound Sub2 level is increased in the absence of Mdm30. Further, we find that Mdm30 does not facilitate splicing of export factors. Moreover, Mdm30 does not have a dramatic effect on mitochondrial respiration/function, and mRNA export occurs in the absence of Fzo1, which is required for mitochondrial dynamics/respiration. Collective results reveal that Mdm30 interacts with Sub2 for proteasomal degradation in a transcription-dependent manner to promote mRNA export independently of splicing or mitochondrial function, thus advancing our understanding of mRNA export.

Proteasomal Regulation of Mammalian SPT16 in Controlling Transcription

“…Impairment of proteasomal degradation of human SPT16 would increase its level/ abundance. Such an increase of human SPT16 abundance might affect transcription, as our previous studies in yeast (37,38) demonstrated that increased abundance of Spt16 in the absence of proteasomal degradation alters transcription. To test this, we analyzed the effect of the high abundance/expression of SPT16 on transcription in HEK293 cells.…”

“…Toward addressing these questions, we carried out experiments in yeast (Saccharomyces cerevisiae) and found that the Spt16 subunit of FACT undergoes ubiquitylation by an E3 ubiquitin ligase, San1, and degradation by the 26S proteasome, hence deciphering a new proteasomal regulation of FACT (37). Furthermore, we found that increased abundance of Spt16 in the absence of proteasomal degradation alters transcription in yeast (37,38). Such a mechanism might be present in humans to regulate FACT and alter the transcription of the genes associated with cancer.…”