Insights on how light regulates alternative polyadenylation in plants

MA. GUILLERMINA KUBACZKA; MICAELA A. GODOY HERZ; J. CHEN; BIN TIAN; ALBERTO KORNBLIHTT

Cold Sping Harbor

Congreso; Eukaryotic mRNA Processing, Cold Spring Harbor Laboratory; 2023

Cold Spring Harbor Laboratory

Light is an essential environmental cue for plants. We have previously shown that light generates a chloroplast retrograde signal that regulates nuclear alternative splicing (AS) of a subset of Arabidopsis thaliana transcripts, and that RNA Polymerase II (RNAPII) transcription elongation rate is involved in these alternative splicing decisions. We are now investigating the mechanism by which light and dark conditions regulate alternative polyadenylation (APA) and its differences and similarities with the control of AS. RNAPII has a characteristic C-terminal domain (CTD) in its largest subunit RPB1. This CTD is composed of numerous repetitions (37 in Arabidopsis) of the aminoacidic heptad YSPTSPS. Post translational modifications in this CTD play a key role in the transcription cycle. Here we show that Arabidopsis mutants defective of CPL1 (cpl1-6) and CPL2 (cpl2-2), two CTD-phosphatases specific of Ser5, abolish the light/dark effect on AS but not on APA. We found that CPL1 and CPL2 are important for the changes in AS in response to light. However, the same mutants show similar to wild type seedlings with respect to the light/dark effect on APA, suggesting that these two mRNA maturation steps show a different sensitivity to the lack of CPL1 and CPL2. Western blots revealed that the RNAPII signal from wild type seedlings exposed to light is stronger than that of seedlings kept in dark conditions. Moreover, for cpl2-2, but not for cpl1-6, total RNAPII abundance is not regulated by light, showing a strong signal in both light and dark conditions. Taken together, these data suggest a role for Ser5 phosphorylation in AS regulation, but not APA regulation, in response to light. More importantly, these results demonstrate a previously undescribed regulation of RNAPII abundance in response to light and a role of a CTD phosphatase in its stability. This raises new questions regarding RNAPII pool stability and degradation that help us understand how basal transcription machinery is sensitive to environmental cues and how this controls key steps in mRNA processing.