The Nuclear Ribonucleoprotein SmD1 Interplays with Splicing, RNA Quality Control, and Posttranscriptional Gene Silencing in Arabidopsis

RNA quality control (RQC) eliminates aberrant RNAs based on their atypical structure, whereas posttranscriptional gene silencing (PTGS) eliminates both aberrant and functional RNAs through the sequence-specific action of short interfering RNAs (siRNAs). The Arabidopsis thaliana mutant smd1b was identified in a genetic screen for PTGS deficiency, revealing the involvement of SmD1, a component of the Smith (Sm) complex, in PTGS. The smd1a and smd1b single mutants are viable, but the smd1a smd1b double mutant is embryo-lethal, indicating that SmD1 function is essential. SmD1b resides in nucleoli and nucleoplasmic speckles, colocalizing with the splicing-related factor SR34. Consistent with this, the smd1b mutant exhibits intron retention at certain endogenous mRNAs. SmD1 binds to RNAs transcribed from silenced transgenes but not nonsilenced ones, indicating a direct role in PTGS. Yet, mutations in the RQC factors UPFRAMESHIFT3, EXORIBONUCLEASE2 (XRN2), XRN3, and XRN4 restore PTGS in smd1b, indicating that SmD1 is not essential for but rather facilitates PTGS. Moreover, the smd1b mtr4 double mutant is embryo-lethal, suggesting that SmD1 is essential for mRNA TRANSPORT REGULATOR4-dependent RQC. These results indicate that SmD1 interplays with splicing, RQC, and PTGS. We propose that SmD1 facilitates PTGS by protecting transgene-derived aberrant RNAs from degradation by RQC in the nucleus, allowing sufficient amounts to enter cytoplasmic siRNA bodies to activate PTGS

The epigenetic landscape of renal cancer

This is an accepted manuscript of an article published by Nature in Nature Reviews: Nephrology on 28/11/2016, available online: https://doi.org/10.1038/nrneph.2016.168 The accepted version of the publication may differ from the final published version.The majority of kidney cancers are associated with mutations in the von Hippel-Lindau gene and a small proportion are associated with infrequent mutations in other well characterized tumour-suppressor genes. In the past 15 years, efforts to uncover other key genes involved in renal cancer have identified many genes that are dysregulated or silenced via epigenetic mechanisms, mainly through methylation of promoter CpG islands or dysregulation of specific microRNAs. In addition, the advent of next-generation sequencing has led to the identification of several novel genes that are mutated in renal cancer, such as PBRM1, BAP1 and SETD2, which are all involved in histone modification and nucleosome and chromatin remodelling. In this Review, we discuss how altered DNA methylation, microRNA dysregulation and mutations in histone-modifying enzymes disrupt cellular pathways in renal cancers

ETUDE DES PARTENAIRES DE LA PROTEINE MPS2 IMPLIQUEE DANS LE MECANISME DE DUPLICATION DU SPB, LE CENTRE ORGANISATEUR DES MICROTUBULES CHEZ LA LEVURE SACCHAROMYCES CEREVISIAE

LA PROTEINE DE LA MEMBRANE NUCLEAIRE MPS2 (MONOPOLAR SPINDLE PROTEIN) EST IMPLIQUEE DANS LA DUPLICATION DU CENTRE ORGANISATEUR DES MICROTUBULES (LE SPB, SPINDLE POLE BODY) CHEZ LA LEVURE SACCHAROMYCES CEREVISIAE. LA RECHERCHE DE PARTENAIRES PAR LA METHODE DU DOUBLE-HYBRIDE A REVELE DES INTERACTIONS AVEC DEUX PROTEINES DU SPB, BBP1 ET SPC24, AINSI QU'AVEC LA PROTEINE NON CARACTERISEE YNL107W. LE MUTANT BBP1-1 EST DECRIT COMME SUBISSANT UNE MAUVAISE DUPLICATION DU SPB ET EST IMPLIQUE AVEC MPS2 DANS LE MECANISME D'ACCROCHAGE DU SPB NEOSYNTHETISE A LA MEMBRANE NUCLEAIRE. CES DEUX PROTEINES CO-REGULENT LES ETAPES TARDIVES DE LA DUPLICATION DU SPB. L'ETUDE DE CES PROTEINES A MONTRE QUE LA LOCALISATION DE SPC24 AU SPB DEPEND DE MPS2, CONTRAIREMENT A BBP1 QUI EST TOUJOURS PRESENTE DANS LES MEMES CONDITIONS. LE MUTANT SPC24-11 SUBIT UN DEFAUT DE SEGREGATION DES CHROMATIDES SURS LORS DE L'ANAPHASE ET N'INTERVIENT PAS DANS LE MECANISME DE DUPLICATION DU SPB. NOS DONNEES CONFIRMENT LES PUBLICATIONS RECENTES LOCALISANT SPC24 DANS LE KINETOCHORE AU SEIN D'UN COMPLEXE FORME DES PROTEINES SPC25/ NUF2 ET NDC80. NOTRE HYPOTHESE EST QUE L'INTERACTION ENTRE MPS2 ET SPC24 EST UTILE AU MECANISME DE CLUSTERING DES CENTROMERES AUTOUR DU SPB LORS DE L'INTERPHASE. LE GENE CODANT POUR LA PROTEINE YNL107 DE FONCTION INCONNUE EST ISSU DU SEQUENCAGE SYSTEMATIQUE DU GENOME DE LA LEVURE S.CEREVISIAE. LA DELETION DU GENE YNL107 EST VIABLE, MAIS LES MUTANTS SONT HYPER-SENSIBLES AU BENOMYL (UNE DROGUE DESTABILISANT LES MICROTUBULES) ET AUX IRRADIATIONS UV. LES MUTANTS SONT LETAUX SYNTHETIQUES AVEC LES MUTANTS SPC24 ET BBP1 THERMOSENSIBLES. CES OBSERVATIONS MONTRENT QUE LA PROTEINE MPS2 JOUE UN ROLE CENTRAL POUR L'INSERTION DU SPB DANS LA MEMBRANE NUCLEAIRE, ET OUVRENT DES PERSPECTIVES NOUVELLES SUR SON IMPLICATION DANS D'AUTRES MECANISMES CELLULAIRES.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

sgs1: a neomorphic nac52 allele impairing post-transcriptional gene silencing through SGS3 downregulation

Post-transcriptional gene silencing (PTGS) is a defense mechanism that targets invading nucleic acids from endogenous (transposons) or exogenous (pathogens, transgenes) sources. Genetic screens based on the reactivation of silenced transgenes have long been used to identify cellular components and regulators of PTGS. Here we show that the first isolated PTGS-deficient mutant, sgs1, is impaired in the transcription factor NAC52. This mutant exhibits striking similarities to a mutant impaired in the H3K4me3 demethylase JMJ14 isolated from the same genetic screen. These similarities include increased transgene promoter DNA methylation, reduced H3K4me3 and H3K36me3 levels, reduced PolII occupancy and reduced transgene mRNA accumulation. It is likely that increased DNA methylation is the cause of reduced transcription because the effect of jmj14 and sgs1 on transgene transcription is suppressed by drm2, a mutation that compromises de novo DNA methylation, suggesting that the JMJ14-NAC52 module promotes transgene transcription by preventing DNA methylation. Remarkably, sgs1 has a stronger effect than jmj14 and nac52 null alleles on PTGS systems requiring siRNA amplification, and this is due to reduced SGS3 mRNA levels in sgs1. Given that the sgs1 mutation changes a conserved amino acid of the NAC proteins involved in homodimerization, we propose that sgs1 corresponds to a neomorphic nac52 allele encoding a mutant protein that lacks wild-type NAC52 activity but promotes SGS3 downregulation. Together, these results indicate that impairment of PTGS in sgs1 is due to its dual effect on transgene transcription and SGS3 transcription, thus compromising siRNA amplification

Invasion of the Arabidopsis Genome by the Tobacco Retrotransposon Tnt1 Is Controlled by Reversible Transcriptional Gene Silencing1[W]

Long terminal repeat (LTR) retrotransposons are generally silent in plant genomes. However, they often constitute a large proportion of repeated sequences in plants. This suggests that their silencing is set up after a certain copy number is reached and/or that it can be released in some circumstances. We introduced the tobacco (Nicotiana tabacum) LTR retrotransposon Tnt1 into Arabidopsis (Arabidopsis thaliana), thus mimicking the horizontal transfer of a retrotransposon into a new host species and allowing us to study the regulatory mechanisms controlling its amplification. Tnt1 is transcriptionally silenced in Arabidopsis in a copy number-dependent manner. This silencing is associated with 24-nucleotide short-interfering RNAs targeting the promoter localized in the LTR region and with the non-CG site methylation of these sequences. Consequently, the silencing of Tnt1 is not released in methyltransferase1 mutants, in contrast to decrease in DNA methylation1 or polymerase IVa mutants. Stable reversion of Tnt1 silencing is obtained when the number of Tnt1 elements is reduced to two by genetic segregation. Our results support a model in which Tnt1 silencing in Arabidopsis occurs via an RNA-directed DNA methylation process. We further show that silencing can be partially overcome by some stresses

Spc24 interacts with Mps2 and is required for chromosome segregation, but is not implicated in spindle pole body duplication

International audienceMps2 (monopolar spindle protein) is a coiled-coil protein found at the spindle pole body (SPB) and at the nuclear envelope that is required for insertion of the SPB into the nuclear envelope. We identified three proteins that interact with Mps2 in a two-hybrid screen: Bbp1, Ynl107w and Spc24. All three proteins contain coiled-coil motifs that appear to be required for their interaction with Mps2. In this work, we verified the Mps2-Spc24 interaction by co-immunoprecipitation in vivo and by the in vitro interaction of recombinant proteins. Previous two-hybrid screens with Spc24 as bait had identified Spc25 and Ndc80 as putative interacting partners, and we verified these interactions in vivo by purification of TAP-tagged derivatives of Spc24 and Ndc80. Finally, we found that spc24 thermosensitive mutants had a chromosome segregation defect, but no apparent defect in SPB duplication. These results are consistent with recently published data showing that Spc24, Spc25 and Ndc80 are peripheral kinetochore com-ponents required for chromosome segregation. The Mps2-Spc24 interaction may contribute to the localization of Spc24 and other kinetochore components to the inner plaque of the SPB

Yaf9, a Novel NuA4 Histone Acetyltransferase Subunit, Is Required for the Cellular Response to Spindle Stress in Yeast

Yaf9 is one of three proteins in budding yeast containing a YEATS domain. We show that Yaf9 is part of a large complex and that it coprecipitates with three known subunits of the NuA4 histone acetyltransferase. Although Esa1, the catalytic subunit of NuA4, is essential for viability, we found that yaf9Δ mutants are viable but hypersensitive to microtubule depolymerizing agents and synthetically lethal with two different mutants of the mitotic apparatus. Microtubules depolymerized more readily in the yaf9Δ mutant compared to the wild type in the presence of nocodazole, and recovery of microtubule polymerization and cell division from limiting concentrations of nocodazole was inhibited. Two other NuA4 mutants (esa1-1851 and yng2Δ) and nonacetylatable histone H4 mutants were also sensitive to benomyl. Furthermore, wild-type budding yeast were more resistant to benomyl when grown in the presence of trichostatin A, a histone deacetylase inhibitor. These results strongly suggest that acetylation of histone H4 by NuA4 is required for the cellular resistance to spindle stress

The nuclear ribonucleoprotein smd1 interplays with splicing, rna quality control, and posttranscriptional gene silencing in arabidopsis

International audienceRNA quality control (RQC) eliminates aberrant RNAs based on their atypical structure, whereas posttranscriptional gene silencing (PTGS) eliminates both aberrant and functional RNAs through the sequence-specific action of short interfering RNAs (siRNAs). The Arabidopsis thaliana mutant smd1b was identified in a genetic screen for PTGS deficiency, revealing the involvement of SmD1, a component of the Smith (Sm) complex, in PTGS. The smd1a and smd1b single mutants are viable, but the smd1a smd1b double mutant is embryo-lethal, indicating that SmD1 function is essential. SmD1b resides in nucleoli and nucleoplasmic speckles, colocalizing with the splicing-related factor SR34. Consistent with this, the smd1b mutant exhibits intron retention at certain endogenous mRNAs. SmD1 binds to RNAs transcribed from silenced transgenes but not nonsilenced ones, indicating a direct role in PTGS. Yet, mutations in the RQC factors UPFRAMESHIFT3, EXORIBONUCLEASE2 (XRN2), XRN3, and XRN4 restore PTGS in smd1b, indicating that SmD1 is not essential for but rather facilitates PTGS. Moreover, the smd1b mtr4 double mutant is embryo-lethal, suggesting that SmD1 is essential for mRNA TRANSPORT REGULATOR4-dependent RQC. These results indicate that SmD1 interplays with splicing, RQC, and PTGS. We propose that SmD1 facilitates PTGS by protecting transgene-derived aberrant RNAs from degradation by RQC in the nucleus, allowing sufficient amounts to enter cytoplasmic siRNA bodies to activate PTGS