Updating the rna polymerase ctd
We also exploited our collection of CTD mutants to query whether and how perturbations of CTD primary structure affect CTD serine phosphorylation patterns in vivo, as gauged by Rpb1 reactivity with phospho-specific antibodies.We found that Ser2 phosphorylation does not rely on Ser5, Pro6, Ser7, or Thr4, whereas Ser5 phosphorylation does not depend on Ser2, Thr4, or Ser7.These issues notwithstanding, the informational rules that govern the CTD code on a cellular and organismal level can be probed genetically by manipulating the composition and structure of the Rpb1 CTD (4 CTD usually is cited as consisting of 29 heptad repeats (12).The four most proximal repeats deviate in size and/or sequence from the consensus heptad, and the informational content of these deviant repeats is uncertain.
Four of the seven coding “letters” of the fission yeast CTD (Tyr1, Pro3, Ser5, Pro6) are essential in vivo, but the grammatical rules of the code are obscure.
This result shows that capping enzyme recruitment is a chief function of the Ser5-PO mark in vivo.
Here we interrogate the grammar and punctuation of the CTD code, focusing on the following questions.
Stiller and Cook (15) put this idea to a clever genetic test in budding yeast, by inserting alanine spacers between every CTD heptad in four different registers: between Ser7 and Tyr1, between Pro6 and Ser7, between Tyr1 and Ser2, and between Pro3 and Thr4.
This alanine spacer mutation was lethal in all four cases.