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Open AccessArticle

CDK12 Activity-Dependent Phosphorylation Events in Human Cells

1
Department of Biochemistry, Duke Med. Ctr., Durham, NC 27710, USA
2
Proteomics Core Facility, Duke Med. Ctr., Durham, NC 27710, USA
*
Author to whom correspondence should be addressed.
Biomolecules 2019, 9(10), 634; https://doi.org/10.3390/biom9100634
Received: 25 August 2019 / Revised: 16 October 2019 / Accepted: 17 October 2019 / Published: 22 October 2019
(This article belongs to the Section Molecular Biology)
We asked whether the C-terminal repeat domain (CTD) kinase, CDK12/CyclinK, phosphorylates substrates in addition to the CTD of RPB1, using our CDK12analog-sensitive HeLa cell line to investigate CDK12 activity-dependent phosphorylation events in human cells. Characterizing the phospho-proteome before and after selective inhibition of CDK12 activity by the analog 1-NM-PP1, we identified 5,644 distinct phospho-peptides, among which were 50 whose average relative amount decreased more than 2-fold after 30 min of inhibition (none of these derived from RPB1). Half of the phospho-peptides actually showed >3-fold decreases, and a dozen showed decreases of 5-fold or more. As might be expected, the 40 proteins that gave rise to the 50 affected phospho-peptides mostly function in processes that have been linked to CDK12, such as transcription and RNA processing. However, the results also suggest roles for CDK12 in other events, notably mRNA nuclear export, cell differentiation and mitosis. While a number of the more-affected sites resemble the CTD in amino acid sequence and are likely direct CDK12 substrates, other highly-affected sites are not CTD-like, and their decreased phosphorylation may be a secondary (downstream) effect of CDK12 inhibition. View Full-Text
Keywords: analog-sensitive CTD kinase; transcription; CTD; RNA polymerase II; RNA processing; mRNA nuclear export; nuclear pore; mESC undifferentiated state; XPC; TPR; 1-NM-PP1 analog-sensitive CTD kinase; transcription; CTD; RNA polymerase II; RNA processing; mRNA nuclear export; nuclear pore; mESC undifferentiated state; XPC; TPR; 1-NM-PP1
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Bartkowiak, B.; Yan, C.M.; Soderblom, E.J.; Greenleaf, A.L. CDK12 Activity-Dependent Phosphorylation Events in Human Cells. Biomolecules 2019, 9, 634.

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