The Peptidyl Prolyl Isomerase Rrd1 Regulates the Elongation of RNA Polymerase II during Transcriptional Stresses Jeremie Poschmann1, Simon Drouin2, Pierre-Etienne Jacques2, Karima El Fadili1, Michael Newmarch1, Fran c¸ois Robert2,3*, Dindial Ramotar1* 1 Department of Medicine, Maisonneuve-Rosemont Hospital, Research Center, University of Montreal, Montre´al, Que´bec, Canada, 2 Institut de recherches cliniques de Montre´al, Montre´al, Que´bec, Canada, 3 De´partement de Me´decine, Faculte´ de Me´decine, Universite´ de Montre´al, Montre´al, Que´bec, Canada Abstract Rapamycin is an anticancer agent and immunosuppressant that acts by inhibiting the TOR signaling pathway. In yeast, rapamycin mediates a profound transcriptional response for which the RRD1 gene is required. To further investigate this connection, we performed genome-wide location analysis of RNA polymerase II (RNAPII) and Rrd1 in response to rapamycin and found that Rrd1 colocalizes with RNAPII on actively transcribed genes and that both are recruited to rapamycin responsive genes. Strikingly, when Rrd1 is lacking, RNAPII remains inappropriately associated to ribosomal genes and fails to be recruited to rapamycin responsive genes. This occurs independently of TATA box binding protein recruitment but involves the modulation of the phosphorylation status of RNAPII CTD by Rrd1. Further, we demonstrate that Rrd1 is also involved in various other transcriptional stress responses besides rapamycin. We propose that Rrd1 is a novel transcription elongation factor that fine-tunes the transcriptional stress response of RNAPII. Citation: Poschmann J, Drouin S, Jacques P-E, El Fadili K, Newmarch M, et al. (2011) The Peptidyl Prolyl Isomerase Rrd1 Regulates the Elongation of RNA Polymerase II during Transcriptional Stresses. PLoS ONE 6(8): e23159. doi:10.1371/journal.pone.0023159 Editor: Fatah Kashanchi, George Mason University, United States of America Received May 11, 2011; Accepted July 7, 2011; Published August 24, 2011 Copyright: 2011 Poschmann et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: This work was supported by the grants CIHR-82891 and MOP-13152 from the Canadian Institutes of Health Research. The authors also thank the Canadian foundation for Innovation for the leader funding opportunity grant (202432-01) to D.R. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: francois.robert@ircm.qc.ca (FR); dramotar.hmr@ssss.gouv.qc.ca (DR) Introduction Rapamycin is an immunosuppressant and an anticancer molecule that acts through inhibition of the TOR (target of rapamycin) signaling pathway [1]. In the yeast Saccharomyces cerevisiae, TOR1 and TOR2 encode serine/threonine kinases that form the core of the rapamycin sensitive (TORC1) and the rapamycin insensitive (TORC2) complex, respectively [2,3]. TORC1 positively regulates anabolic processes by promoting mRNA translation and the transcription of ribosome biogenesis genes [2,3]. Upon nutrient starvation or rapamycin treatment, the TORC1 complex becomes inactivated, with the consequence of a severe reduction of anabolic processes, cell cycle progression and growth, as well as the induction of catabolic processes and stress responsive factors [2,3]. These drastic changes are driven by important alterations of gene transcription mediated, at least in part, by the translocation of transcription factors to the nucleus. Upon TORC1 inhibition, the ribosomal gene repressor, Crf1, translocates into the nucleus to inhibit ribosomal gene transcrip- tion [4]. Furthermore, the TORC1 regulator Tap42 activates PP2A and Sit4 phosphatases, which in turn dephosphorylate the transcription factors Rtg1/2 and Gln3 causing them to move into the nucleus and induce the expression of retrograde signalling genes (RTG) and nitrogen discrimination genes (NDG), respec- tively [2,5,6,7]. Once translocated into the nucleus, these transcription factors bind to specific DNA elements, alter the local chromatin state and recruit the general transcription machinery to mediate pre-initiation complex (PIC) assembly and transcription by RNAPII [8,9]. The exact mechanisms of these regulatory circuits are not fully understood but genome wide deletion screens in S.cerevisae have been a useful tool to identify novel factors that are required to mediate an efficient response to rapamycin [10,11,12,13]. One of these factors is the peptidyl prolyl isomerase Rrd1 (Resistant to rapamycin deletion 1). Rrd1D mutants exhibit multiple phenotypes including sensitivity to the carcinogen 4-nitroquinoline-1-oxide and UVA radiation, and, most prominently, extreme resistance to rapamycin [13]. Rrd1 is evolutionally conserved and shares 35% identity with its human homologue PTPA [14]. PTPA was first characterized to be an activator of the phospho-tyrosyl phospha- tase activity of PP2A phosphatases in vitro [15,16]. However, an in vivo role for this activity has not yet been described, and subsequent studies revealed that PTPA as well as Rrd1 are required for PP2A substrate specificity, complex formation and the reactivation of inactive PP2A complexes [17,18]. Both were later found to possess intrinsic peptidyl prolyl isomerase activity on a specific PP2A peptide [14]. Consistent with this function, we and others found that Rrd1 interacts with the yeast PP2A-like phosphatase Sit4 [19]. Sit4 and Rrd1 form a ternary complex with the Tor signaling mediator Tap42 [19]. As mentioned above, upon TORC1 inactivation Tap42 dissociates from Sit4-Rrd1, which then dephosphorylates and activates the transcription factor Gln3 PLoS ONE | www.plosone.org 1 August 2011 | Volume 6 | Issue 8 | e23159