Inhibition of CDK7, CDK9 or CDK12 affects phosphorylation of human SPT5.

In HEK293 cells, inhibiting CDK7, CDK9, or CDK12 reduces phosphorylation of endogenous SPT5, with CDK9 inhibition showing the most marked effect.

• Why it matters: SPT5 phosphorylation regulates transcription elongation and checkpoint release, but the specific kinase contributions and how they affect PAF1C-linked phosphorylation sites were unclear.
• What they did: The study used in vitro phosphorylation of SPT5 C-terminus (GST pull-down) to confirm RTF1 interaction with residues 750–850 only when phosphorylated, and generated phospho-S666 and phospho-T806 antibodies.
• The result: Western blots and ChIP showed that all three kinases contribute to SPT5 phosphorylation, and CDK9 inhibition most strongly reduced PAF1C subunit association with SPT5 and phosphorylation of chromatin-associated SPT5.