Genetic screen for suppression of transcriptional interference identifies a gain-of-function mutation in Pol2 termination factor Seb1.
The system of long noncoding RNA (lncRNA)-mediated transcriptional interference that represses fission yeast phosphate homoeostasis gene  pho1  provides a sensitive readout of genetic influences on cotranscriptional 3'-processing and termination and a tool for discovery of regulators of this phase of the Pol2 transcription cycle. Here, we conducted a genetic screen for relief of transcriptional interference that unveiled a mechanism by which Pol2 termination is enhanced via a gain-of-function mutation, G476S, in the RNA-binding domain of an essential termination factor, Seb1. The genetic and physical evidence for gain-of-function is compelling: 1)  seb1-G476S  de-represses  pho1  and  tgp1 , both of which are subject to lncRNA-mediated transcriptional interference; 2)  seb1-G476S  elicits precocious lncRNA transcription termination in response to lncRNA 5'-proximal poly(A) signals; 3)  seb1-G476S  derepression of  pho1  is effaced by loss-of-function mutations in cleavage and polyadenylation factor (CPF) subunits and termination factor Rhn1; 4) synthetic lethality of  seb1-G476S  with  pho1  derepressive mutants  rpb1-CTD-S7A  and  aps1 ∆ is rescued by CPF/Rhn1 loss-of-function alleles; and 5)  seb1-G476S  elicits an upstream shift in poly(A) site preference in several messenger RNA genes. A crystal structure of the Seb1-G476S RNA-binding domain indicates potential for gain of contacts from Ser476 to RNA nucleobases. To our knowledge, this is a unique instance of a gain-of-function phenotype in a eukaryal transcription termination protein.
Schwer B, Garg A, Jacewicz A, Shuman S