Disruption of the enzymatic activities of the transcription factor TFIIH by the small molecules Triptolide (TPL) or THZ1 could be used against cancer. Here, we used the MCF10A-ErSrc oncogenesis model to compare the effect of TFIIH inhibitors between transformed cells and their progenitors. We report that tumour cells exhibited highly increased sensitivity to TPL or THZ1 and that the combination of both had a synergic effect. TPL affects the interaction between XPB and p52, causing a reduction in the levels of XPB, p52 and p8, but not other TFIIH subunits. RNA-Seq and RNAPII-ChIP-Seq experiments showed that although the levels of many transcripts were reduced, the levels of a significant number were increased after TPL treatment, with maintained or increased RNAPII promoter occupancy. A significant number of these genes encode for factors that have been related to tumour growth and metastasis, suggesting that transformed cells might rapidly develop resistance to TPL/THZ inhibitors. Some of these genes were also overexpressed in response to THZ1, of which depletion enhances the toxicity of TPL, and are possible new targets against cancer.
Transcription and splicing are intrinsically coupled. Transcription dynamics regulate splicing, and splicing feeds back to transcription initiation to jointly determine gene expression profiles. A recently described phenomenon called exon-mediated activation of transcription starts (EMATS) shows that splicing of internal exons can regulate transcription initiation and activate cryptic promoters. Here, we present the first complete catalog of human EMATS genes that have a weak alternative promoter located upstream and proximate to an efficiently spliced internal skipped exon. We found that EMATS genes are associated with Mendelian genetic diseases, specifically intellectual development disorders, cardiomyopathy, and immunodeficiency, and provide a list of EMATS genes with pathological variants. EMATS was originally described as a natural mechanism used during evolution to fine-tune gene expression through punctual genomic mutations that affect splicing. Here, we show that EMATS can be used to manipulate gene expression with therapeutic purposes. We constructed stable cell lines expressing a splicing reporter based on the alternative splicing of exon 7 of SMN2 gene under the regulation of different promoters. Using a small molecule (Risdiplam) and an antisense oligonucleotide (ASO) modeled after Spinraza, we promoted the inclusion of SMN2 exon 7 which triggered an increase in gene expression up to 40-folds by activating transcription initiation. We observed the strongest effects in reporters under the regulation of weak human promoters, where the highest drug doses dramatically increased exon inclusion. Overall, our findings present evidence to develop the first therapeutic strategy to use EMATS to activate gene expression using small molecules and ASOs that affect splicing.
Transcription and splicing are intrinsically coupled. Alternative splicing of internal exons can fine-tune gene expression through a recently described phenomenon called exon-mediated activation of transcription starts (EMATS). However, the association of this phenomenon with human diseases remains unknown. Here, we develop a strategy to activate gene expression through EMATS and demonstrate its potential for treatment of genetic diseases caused by loss of expression of essential genes. We first identified a catalog of human EMATS genes and provide a list of their pathological variants. To test if EMATS can be used to activate gene expression, we constructed stable cell lines expressing a splicing reporter based on the alternative splicing of motor neuron 2 (SMN2) gene. Using small molecules and antisense oligonucleotides (ASOs) currently used for treatment of spinal muscular atrophy, we demonstrated that increase of inclusion of alternative exons can trigger an activation of gene expression up to 45-fold by enhancing transcription in EMATS-like genes. We observed the strongest effects in genes under the regulation of weak human promoters located proximal to highly included skipped exons.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.