From structure to systems: high-resolution, quantitative genetic analysis of RNA polymerase II.
RNA polymerase II (RNAPII) lies at the core of dynamic control of gene expression. Using 53 RNAPII point mutants, we generated a point mutant epistatic miniarray profile (pE-MAP) comprising ∼60,000 quantitative genetic interactions in Saccharomyces cerevisiae. This analysis enabled functional assignment of RNAPII subdomains and uncovered connections between individual regions and other protein complexes. Using splicing microarrays and mutants that alter elongation rates in vitro, we found an inverse relationship between RNAPII speed and in vivo splicing efficiency. Furthermore, the pE-MAP classified fast and slow mutants that favor upstream and downstream start site selection, respectively. The striking coordination of polymerization rate with transcription initiation and splicing suggests that transcription rate is tuned to regulate multiple gene expression steps. The pE-MAP approach provides a powerful strategy to understand other multifunctional machines at amino acid resolution.
Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA.
Hannes
H
Braberg
Huiyan
H
Jin
Erica A
EA
Moehle
Yujia A
YA
Chan
Shuyi
S
Wang
Michael
M
Shales
Joris J
JJ
Benschop
John H
JH
Morris
Chenxi
C
Qiu
Fuqu
F
Hu
Leung K
LK
Tang
James S
JS
Fraser
Frank C P
FC
Holstege
Philip
P
Hieter
Christine
C
Guthrie
Craig D
CD
Kaplan
Nevan J
NJ
Krogan
D000483:Alleles; D004843:Epistasis, Genetic; D055106:Genome-Wide Association Study; D017354:Point Mutation; D012319:RNA Polymerase II / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D012326:RNA Splicing; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D024363:Transcription Initiation Site; D014158:Transcription, Genetic; D059467:Transcriptome
D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D012319:RNA Polymerase II