Strains used in this work are listed in the Supplemental Material. Inactivation of Sen1 helicase activity in sen resulted in accumulation of RNA: Such a cooperative mechanism may facilitate Pcf11 recruitment at early stages of transcription. These data are presented graphically Fig. We also established that the NRD complex is intact in pcf cells based on protein levels of Nrd1, Nab3, and Sen1 as compared with wild-type cells Fig. Consistent with this, we observed that episomal expression of the Pcf11 CID from the GAL1 promoter only partially rescued pcf temperature-sensitive growth Fig. pcf11

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RNA and its application to immunodetection of hybrids.

Pcf11 orchestrates transcription termination pathways in yeast

Notably, we did not detect any change in hybrid accumulation between wild type and sen Fig. S8Aas previously reported Kim et al. For Western blot analysis, proteins were resolved in Tris-acetate gels Life Technologies and electrotransferred into nitrocellulose membrane. We overexpressed Pcf11 from a multicopy plasmid in wild-type cells Supplemental Fig.

Interactions of Sen1, Nrd1, and Nab3 with multiple phosphorylated forms of the Rpb1 C-terminal domain in Saccharomyces cerevisiae. Significantly, in pcfbut not in senNEL S accumulated in the first 10 min of inhibition and then subsequently decreased. We also thank A. DNA hybrids in the same regions in pcf Fig. E A model of NRD-dependent transcription termination. Promoter directionality is controlled by U1 snRNP and polyadenylation signals. Diagrams above the graphs depict loci and positions of amplicons.

This delayed Nrd1 recruitment profile in pcf was not due to a general termination defect over NRD-dependent genes. We thank members of N. Thiolutin inhibits transcription initiation but has less effect on elongation Tipper We also established that the NRD complex is intact in pcf cells based on protein levels of Nrd1, Nab3, and Sen1 as compared with wild-type cells Fig. In addition, annotations are classified as classical genetics or high-throughput e.

PCF11 / YDR228C Overview

To confirm this, we used ChIP to analyze Nrd1 and Pcf11 recruitment under varying expression levels of each factor. This experiment confirms that upon loss of Sen1 activity, Pol II pausing occurs downstream from the proximal regions of NRD terminators.

Pcf11 terminates the majority of ncRNA. NRD1 expression by growth in glucose-containing medium Fig. Consistently, thiolutin-dependent transcriptional shutoff in pcf was followed by an initially strong increase of ncRNA NELc and its subsequent delayed degradation.

Nucleic Acids Res Genetic Interactions Dosage Rescue: NRD may also regulate pvf11 of protein-coding genes through the promotion of premature transcription termination Steinmetz et al. J Immunol Methods Ser2-P marks are also needed for direct Sen1—Rpb1 interaction Chinchilla et al. PCF11 encodes a third protein component of yeast cleavage and polyadenylation factor I.

Regulators 1 Targets 0. These data are presented graphically Fig.