Journal Article

Pol II phosphorylation regulates a switch between transcriptional and splicing condensates

Shrinivas, Krishna, Manteiga, John C., Sharp, Phillip A., Henninger, Jonathan E., Young, Richard A., Hannett, Nancy M., Guo, Yang Eric, Lee, Tong Ihn, Spille, Jan-Hendrik, Rimel, Jenna K., Abraham, Brian J., Dall’Agnese, Alessandra, Decker, Tim-Michael, Zamudio, Alicia V., Afeyan, Lena K., Sabari, Benjamin R., Boija, Ann, Cisse, Ibrahim I., Taatjes, Dylan J., Fant, Charli B.

2019

Overview

The synthesis of pre-mRNA by RNA polymerase II (Pol II) involves the formation of a transcription initiation complex, and a transition to an elongation complex 1 – 4 . The large subunit of Pol II contains an intrinsically disordered C-terminal domain that is phosphorylated by cyclin-dependent kinases during the transition from initiation to elongation, thus influencing the interaction of the C-terminal domain with different components of the initiation or the RNA-splicing apparatus 5 , 6 . Recent observations suggest that this model provides only a partial picture of the effects of phosphorylation of the C-terminal domain 7 – 12 . Both the transcription-initiation machinery and the splicing machinery can form phase-separated condensates that contain large numbers of component molecules: hundreds of molecules of Pol II and mediator are concentrated in condensates at super-enhancers 7 , 8 , and large numbers of splicing factors are concentrated in nuclear speckles, some of which occur at highly active transcription sites 9 – 12 . Here we investigate whether the phosphorylation of the Pol II C-terminal domain regulates the incorporation of Pol II into phase-separated condensates that are associated with transcription initiation and splicing. We find that the hypophosphorylated C-terminal domain of Pol II is incorporated into mediator condensates and that phosphorylation by regulatory cyclin-dependent kinases reduces this incorporation. We also find that the hyperphosphorylated C-terminal domain is preferentially incorporated into condensates that are formed by splicing factors. These results suggest that phosphorylation of the Pol II C-terminal domain drives an exchange from condensates that are involved in transcription initiation to those that are involved in RNA processing, and implicates phosphorylation as a mechanism that regulates condensate preference. RNA polymerase II with a hypophosphorylated C-terminal domain preferentially incorporates into mediator condensates, and with a hyperphosphorylated C-terminal domain into splicing-factor condensates, revealing phosphorylation as a regulatory mechanism in condensate preference.

• Publisher: Nature Publishing Group UK,Nature Publishing Group
• Subject: 13/100 / 13/106 / 13/109 / 14/1 / 14/19 / 14/32 / 14/34 / 14/35 / 14/63 / 38 / 38/23 / 38/5 / 38/77 / 42 / 45 / 631/208/200 / 631/337/572 / 631/80/386 / 82 / 82/29 / 82/80 / 82/83 / 96 / Analysis / Animals / Cell Line / Chemical synthesis / Condensates / Cyclin-dependent kinases / DNA-directed RNA polymerase / Elongation / Enhancer Elements, Genetic - genetics / Enhancers / Gene expression / Gene Expression Regulation - genetics / Genetic transcription / Genomics / Humanities and Social Sciences / Humans / Incorporation / Initiation complex / Kinases / Letter / Mediator Complex - chemistry / Mediator Complex - genetics / Mediator Complex - metabolism / Methods / Mice / Microscopy / multidisciplinary / Phosphorylation / Protein Domains / RNA polymerase / RNA polymerase II / RNA Polymerase II - chemistry / RNA Polymerase II - genetics / RNA Polymerase II - metabolism / RNA polymerases / RNA processing / RNA Splicing / RNA Splicing Factors - chemistry / RNA Splicing Factors - genetics / RNA Splicing Factors - metabolism / Science / Science (multidisciplinary) / Splicing / Splicing factors / Stem cells / Transcription initiation / Transcription, Genetic