GSK-3 is an RNA polymerase II phospho-CTD kinase

Abstract

We have previously found that UV-induced DNA damage causes hyperphosphorylation of the carboxy terminal domain (CTD) of RNA polymerase II (RNAPII), inhibition of transcriptional elongation and changes in alternative splicing (AS) due to kinetic coupling between transcription and splicing. In an unbiased search for protein kinases involved in the AS response to DNA damage, we have identified glycogen synthase kinase 3 (GSK-3) as an unforeseen participant. Unlike Cdk9 inhibition, GSK-3 inhibition only prevents CTD hyperphosphorylation triggered by UV but not basal phosphorylation. This effect is not due to differential degradation of the phospho-CTD isoforms and can be reproduced, at the AS level, by overexpression of a kinase-dead GSK-3 dominant negative mutant. GSK-3 inhibition abrogates both the reduction in RNAPII elongation and changes in AS elicited by UV. We show that GSK-3 phosphorylates the CTD in vitro, but preferentially when the substrate is previously phosphorylated, consistently with the requirement of a priming phosphorylation reported for GSK-3 efficacy. In line with a role for GSK-3 in the response to DNA damage, GSK-3 inhibition prevents UV-induced apoptosis. In summary, we uncover a novel role for a widely studied kinase in key steps of eukaryotic transcription and pre-mRNA processing.

DNA damage-induced transcription stress triggers the genome-wide degradation of promoter-bound Pol II

“…4d ). Together these results not only corroborate that the degradation of promoter-bound Pol II is regulated in trans and implicate GSK3 in the signal transduction from TBL-stalled Pol II to promoter-bound Pol II, but also complement the recently described role of GSK3 in the transcriptional response to DNA damage 67 .

Fig.

…”

“…Besides its early-identified role in metabolism, GSK3 has a large number of additional phosphorylation targets involved in different signaling pathways, and thereby regulates a variety of biological processes, including gene expression, cell survival, and cell proliferation 64 , 69 . In line with a role for GSK3 in the transcription stress response, it was recently identified to phosphorylate Pol II upon DNA damage and thereby regulating alternative splicing 67 . While it is tempting to speculate that the GSK3-mediated phosphorylation of RPB1 itself could result in a phosphodegron-based degradation of Pol II, we could not observe any effect for βTRCP or FBXW7 (data not shown), proteins that can target GSK3-phosphorylated proteins for degradation 70 , 71 .…”

The Phantom Mark: Enigmatic roles of phospho‐Threonine 4 modification of the C‐terminal domain of RNA polymerase II

“…While largely unexplored, investigating the entire kinome under different signaling conditions may very well reveal new kinases with pThr4 activity. Indeed, novel CTD kinases that act on residues other than Thr4 have been discovered when cells were subject to distinct signals and cell cycle cues (Chasman et al, 2014; Di Vona et al, 2015; Kõivomäigi et al, 2021; Nieto Moreno et al, 2020; Singh et al, 2017; Tee et al, 2014; Yurko et al, 2017). While many of these signaling kinases have other cellular substrates, their ability to directly phosphorylate the CTD was unexpected and suggests a direct role in modulating RNA polymerase function.…”

Development of a 2,4-Diaminothiazole Series for the Treatment of Human African Trypanosomiasis Highlights the Importance of Static–Cidal Screening of Analogues

“…Prepared following general procedure A, 140 mg, 38%; δ H (500 MHz, CDCl 3 ): 7.26 (d, J = 7.9 Hz, 2H, ArH), 7.19−7.17 (23). Prepared following general procedure A, 110 mg, 30%; δ H (500 MHz, CDCl (26). Prepared following general procedure A, 179 mg, 49%; δ H (500 MHz, CDCl 3 ): 7.69 (dd, J = 6.7 and 2.1 Hz, 2H, ArH), 7.09 (d, J = 8.9 Hz, 2H, ArH), 6.50 (t, J = 73.6 Hz) (fluorine splitting, OCHF 2 ), 6.00 (bs, 1H, NH), 3.21 (bs, 1H, CH), 1.97 (dd, J = 12.6 and 3.2 Hz, 2H, CH), 1.70 (dt, J = 13.6 and 3.9 Hz, 2H, CH), 1.56 (dt, J = 12.9 and 3.9 Hz, 1H, CH), 1.30 (ddd, J = 25.0, 11.…”