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The THP1-SAC3-SUS1-CDC31 complex works in transcription elongation-mRNA export preventing RNA-mediated genome instability

Centro Andaluz de Biologia Molecular y Medicina Regenerativa, Universidad de Sevilla-CSIC, 41092 Sevilla, Spain.
Molecular biology of the cell (Impact Factor: 5.98). 08/2008; 19(10):4310-8. DOI: 10.1091/mbc.E08-04-0355
Source: PubMed

ABSTRACT The eukaryotic THO/TREX complex, involved in mRNP biogenesis, plays a key role in the maintenance of genome integrity in yeast. mRNA export factors such as Thp1-Sac3 also affect genome integrity, but their mutations have other phenotypes different from those of THO/TREX. Sus1 is a novel component of SAGA transcription factor that also associates with Thp1-Sac3, but little is known about its effect on genome instability and transcription. Here we show that Thp1, Sac3, and Sus1 form a functional unit with a role in mRNP biogenesis and maintenance of genome integrity that is independent of SAGA. Importantly, the effects of ribozyme-containing transcription units, RNase H, and the action of human activation-induced cytidine deaminase on transcription and genome instability are consistent with the possibility that R-loops are formed in Thp1-Sac3-Sus1-Cdc31 as in THO mutants. Our data reveal that Thp1-Sac3-Sus1-Cdc31, together with THO/TREX, define a specific pathway connecting transcription elongation with export via an RNA-dependent dynamic process that provides a feedback mechanism for the control of transcription and the preservation of genetic integrity of transcribed DNA regions.

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    • "The N-terminal RRM, LRR, and NTF2L domains are thought to recognize RNA, whereas the NTF2L domain binds Mtr2 and, together with the UBA domain, provides lowaffinity binding sites for nucleoporin FG repeats (Kang et al., 1999; Liker et al., 2000; Fribourg et al., 2001; Bachi et al., 2000; Grant et al., 2002, 2003; Aibara et al., 2015). Transcription export complex 2 (TREX-2) contributes to both mRNA nuclear export and its integration with the nuclear steps of the gene expression pathway (Rodriguez-Navarro et al., 2004; Gonzá lez-Aguilera et al., 2008). TREX-2 is based on a Sac3 scaffold (Figure 1A) to which Thp1, Sem1, Sus1, and Cdc31 bind, and is localized primarily at NPCs through interactions with proteins located on the nucleoplasmic face, including Nup1 (Fischer et al., 2002, 2004; Faza et al., 2009; Jani et al., 2009, 2014; Ellisdon and Stewart, 2012). "
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    ABSTRACT: The TREX-2 complex integrates mRNA nuclear export into the gene expression pathway and is based on a Sac3 scaffold to which Thp1, Sem1, Sus1, and Cdc31 bind. TREX-2 also binds the mRNA nuclear export factor, Mex67:Mtr2, through the Sac3 N-terminal region (Sac3N). Here, we characterize Chaetomium thermophilum TREX-2, show that the in vitro reconstituted complex has an annular structure, and define the structural basis for interactions between Sac3, Sus1, Cdc31, and Mex67:Mtr2. Crystal structures show that the binding of C. thermophilum Sac3N to the Mex67 NTF2-like domain (Mex67(NTF2L)) is mediated primarily through phenylalanine residues present in a series of repeating sequence motifs that resemble those seen in many nucleoporins, and Mlp1 also binds Mex67:Mtr2 using a similar motif. Deletion of Sac3N generated growth and mRNA export defects in Saccharomyces cerevisiae, and we propose TREX-2 and Mlp1 function to facilitate export by concentrating mature messenger ribonucleoparticles at the nuclear pore entrance. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.
    Structure 06/2015; 23(7). DOI:10.1016/j.str.2015.05.002 · 6.79 Impact Factor
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    • "The action of Nup1-bound TREX2 and chromatin-bound SAGA in retaining GAL1 at the NPC ensures that repression by Ssn6 and Tup1 is suppressed because of proximity to Ulp1. The proximity of GAL1 and other highly regulated actively transcribing genes to the NPC may facilitate the nuclear export of transcripts as well as facilitating transcriptional memory and promoting gene stability (1,3,4,7,8,9,10,11). "
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    ABSTRACT: The conserved Sac3:Thp1:Sem1:Sus1:Cdc31 (TREX2) complex binds to nuclear pore complexes (NPCs) and, in addition to integrating mRNA nuclear export with preceding steps in the gene expression pathway, facilitates re-positioning of highly regulated actively transcribing genes (such as GAL1) to NPCs. Although TREX2 is thought to bind NPC protein Nup1, defining the precise role of this interaction has been frustrated by the complex pleiotropic phenotype exhibited by nup1Δ strains. To provide a structural framework for understanding the binding of TREX2 to NPCs and its function in the gene expression pathway, we have determined the structure of the Nup1:TREX2 interaction interface and used this information to engineer a Sac3 variant that impairs NPC binding while not compromising TREX2 assembly. This variant inhibited the NPC association of both de-repressed and activated GAL1 and also produced mRNA export and growth defects. These results indicate that the TREX2:Nup1 interaction facilitates the efficient nuclear export of bulk mRNA together with the re-positioning of GAL1 to NPCs that is required for transcriptional control that is mediated by removal of SUMO from repressors by NPC-bound Ulp1.
    Nucleic Acids Research 04/2014; 42(10). DOI:10.1093/nar/gku252 · 9.11 Impact Factor
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    • "Our results provide new evidence for the link between mRNA biogenesis and genome instability. Mutants affecting various steps of transcription, from initiation to termination, and RNA processing have been shown to lead to an increase of γH2A foci, YAC instability or hyperrecombination in yeast and human cells [14], [17], [19], [20], [31], [43], [44]. In addition, our screening identified other non-RNA related factors whose deletion could have an indirect impact on genome instability: APC9, encoding a subunit of the Anaphase-Promoting Complex/Cyclosome (APC/C), and TOS3, considered as the functional orthologous of LKB1, a mammalian kinase associated with Peutz-Jeghers cancer-susceptibility syndrome (Figure 1). "
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    ABSTRACT: To get further insight into the factors involved in the maintenance of genome integrity we performed a screening of Saccharomyces cerevisiae deletion strains inducing hyperrecombination. We have identified trf4, a gene encoding a non-canonical polyA-polymerase involved in RNA surveillance, as a factor that prevents recombination between DNA repeats. We show that trf4Δ confers a transcription-associated recombination phenotype that is mediated by the nascent mRNA. In addition, trf4Δ also leads to an increase in the mutation frequency. Both genetic instability phenotypes can be suppressed by overexpression of RNase H and are exacerbated by overexpression of the human cytidine deaminase AID. These results suggest that in the absence of Trf4 R-loops accumulate co-transcriptionally increasing the recombination and mutation frequencies. Altogether our data indicate that Trf4 is necessary for both mRNA surveillance and maintenance of genome integrity, serving as a link between RNA and DNA metabolism in S. cerevisiae.
    PLoS ONE 06/2013; 8(6):e65541. DOI:10.1371/journal.pone.0065541 · 3.23 Impact Factor
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