The Transcription Elongation Factor Spt5 Influences Transcription by RNA Polymerase I Positively and Negatively

Abstract

Spt5p is a universally conserved transcription factor that plays multiple roles in eukaryotic transcription elongation. Spt5p forms a heterodimer with Spt4p and collaborates with other transcription factors to pause or promote RNA polymerase II transcription elongation. We have shown previously that Spt4p and Spt5p also influence synthesis of ribosomal RNA by RNA polymerase (Pol) I; however, previous studies only characterized defects in Pol I transcription induced by deletion of SPT4. Here we describe two new, partially active mutations in SPT5 and use these mutant strains to characterize the effect of Spt5p on Pol I transcription. Genetic interactions between spt5 and rpa49⌬ mutations together with measurements of ribosomal RNA synthesis rates, rDNA copy number, and Pol I occupancy of the rDNA demonstrate that Spt5p plays both positive and negative roles in transcription by Pol I. Electron microscopic analysis of mutant and WT strains confirms these observations and supports the model that Spt4/5 may contribute to pausing of RNA polymerase I early during transcription elongation but promotes transcription elongation downstream of the pause(s). These findings bolster the model that Spt5p and related homologues serve diverse critical roles in the control of transcription.

The Composition of the Arabidopsis RNA Polymerase II Transcript Elongation Complex Reveals the Interplay between Elongation and mRNA Processing Factors

“…A few studies yielded the finding that TEFs (i.e., SPT5, SPT6, and FACT) known to cooperate with RNAPII also play a role in RNAPI (and RNAPIII)-mediated transcription (Anderson et al, 2011;Birch et al, 2009;Engel et al, 2015). Our proteomics experiments did not provide conclusive evidence for the interaction of the seven analyzed TEFs with RNAPI/ RNAPIII, as no RNAPIII-specific subunits copurified with the TEFs and three RNAPI subunits were only isolated along with SPT4 (Supplemental Data Set 5).…”

“…Certain TEFs, including SPT5 and SPT6, were reported to play a role in RNAPI-catalyzed transcription in yeast (Anderson et al, 2011;Engel et al, 2015). Additionally, there is evidence that mammalian FACT is involved in chromatin transcription by RNAPI and RNAPIII (Birch et al, 2009).…”

Biochemical Analysis of Yeast Suppressor of Ty 4/5 (Spt4/5) Reveals the Importance of Nucleic Acid Interactions in the Prevention of RNA Polymerase II Arrest

“…The ability of the SPT5(302-308A) mutant to support viability seems at odds with its inability to prevent arrest in vitro. However, most bona fide elongation factors are non-essential in yeast; SPT5 is the exception because it participates in a number of essential functions such as splicing, capping, and ribosomal RNA production (5,34,38,70,71). The elongation promoting activity of Spt4/5 may not be an essential function for cell survival under idealized growth conditions, but it imparts a competitive advantage to the organism or is more important during stress.…”

Involvement of PIP2 in RNA Polymerase I transcription

“…In the early stages of rRNA processing, Fib, being a component of ribonucleoprotein complex called box C/D small nucleolar RNP, binds to precursor rRNA in the DFC region and functions in site-specific 29-Omethylation of rRNA (Hernandez-Verdun, 1991). Mutations affecting Pol I elongation also have an effect on precursor rRNA cleavage by the Spt4-Spt5 complex in yeast linking both machineries (Anderson et al, 2011;Schneider et al, 2006;Schneider et al, 2007). Fib is recruited to nucleoli upon transcription initiation in telophase, and its presence at this point of the cell cycle was shown to be essential for cell survival (Dundr et al, 1997;Kopp et al, 2007).…”