Rrd1p, an RNA polymerase II-specific prolyl isomerase and activator of phosphoprotein phosphatase, promotes transcription independently of rapamycin response.
Bottom Line: Similarly, inducible, but rapamycin-responsive, non-GAL genes such as CTT1, STL1 and CUP1 are also regulated by Rrd1p.Consistently, transcription of the constitutively active genes is not changed in the Δrrd1 strain.Taken together, our results demonstrate a new function of Rrd1p in stimulation of initial rounds of transcription, but not steady-state/constitutive transcription, of both rapamycin-responsive and non-responsive genes independently of rapamycin treatment.
Affiliation: Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL 62901, USA.Show MeSH
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Mentions: Our above results at GAL1 reveal that Rrd1p promotes the PIC formation, association of elongating RNA polymerase II, and hence transcription following 90 min transcriptional induction. We next asked whether Rrd1p has any effect on GAL1 transcription when the steady-state is reached after a long induction in galactose-containing growth medium. To address this, we have continuously grown both the wild-type and Δrrd1 strains in galactose-containing growth medium up to an OD600 of 1.0 prior to crosslinking/harvesting, and then performed RT-PCR and ChIP analyses. We found that transcription of GAL1 in the Δrrd1 strain reached the wild-type level when the steady-state is reached after a long transcriptional induction (Figure 3A). Consistently, the level of RNA polymerase II at the GAL1 coding sequence in the Δrrd1 strain was almost same as that of the wild-type equivalent following long transcriptional induction (Figure 3B). Further, we performed the kinetic analysis for the association of RNA polymerase II with GAL1, and found that Rrd1p has significant stimulatory effects on RNA polymerase II association with GAL1 during initial stages of transcriptional induction, but not after long induction (Figure 3C and D). Thus, Rrd1p promotes the initial rounds of GAL1 transcription, but has no effect on steady-state transcription. Therefore, the growth defect of the Δrrd1 strain was not observed in the solid medium containing galactose (Figure 3E). Furthermore, we find that the role of Rrd1p in stimulation of RNA polymerase II association with GAL1 (and hence transcription) is correlated with facilitated nucleosomal disassembly as the eviction of histone H2B from GAL1 is impaired in the Δrrd1 strain following transcriptional induction (Figure 3F and G).
Affiliation: Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL 62901, USA.