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Kathleen Boris-lawrie - One of the best experts on this subject based on the ideXlab platform.
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Evidence thAt Lin28 stimulAtes trAnslAtion by recruiting RNA HelicAse A to polysomes.
Nucleic acids research, 2011Co-Authors: Jianyu Jin, Chen Feng, Wei Jing, Xinxiang Lei, Shuping Peng, Kathleen Boris-lawrie, Yingqun HuangAbstract:The stem cell protein Lin28 functions to inhibit the biogenesis of A group of miRNAs but Also stimulAtes the expression of A subset of mRNAs At the post-trAnscriptionAl level, the underlying mechAnism of which is not yet understood. Here we report the chArActerizAtion of the moleculAr interplAy between Lin28 And RNA HelicAse A (RHA) known to plAy An importAnt role in remodeling ribonucleoprotein pArticles during trAnslAtion. We show thAt reducing Lin28 expression results in decreAsed RHA AssociAtion with polysomes while increAsing Lin28 expression leAds to elevAted RHA AssociAtion. Further, the cArboxyl terminus of Lin28 is necessAry for interAction with both the Amino And cArboxyl termini of RHA. ImportAntly, A cArboxyl terminAl deletion mutAnt of Lin28 thAt retAins RNA-binding Activity fAils to interAct with RHA And exhibits dominAnt-negAtive effects on Lin28-dependent stimulAtion of trAnslAtion. TAken together, these results leAd us to suggest thAt Lin28 mAy stimulAte trAnslAtion by Actively recruiting RHA to polysomes.
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FeAtures of Double-strAnded RNA-binding DomAins of RNA HelicAse A Are NecessAry for Selective Recognition And TrAnslAtion of Complex mRNAs
The Journal of biological chemistry, 2010Co-Authors: Arnaz Ranji, Nikolozi Shkriabai, Mamuka Kvaratskhelia, Karin Musier-forsyth, Kathleen Boris-lawrieAbstract:The DExH protein RNA HelicAse A (RHA) plAys numerous roles in cell physiology, And post-trAnscriptionAl ActivAtion of gene expression is A mAjor role Among them. RHA selectively ActivAtes trAnslAtion of complex cellulAr And retrovirAl mRNAs. Although RHA requires interAction with structurAl feAtures of the 5'-UTR of these tArget mRNAs, the moleculAr bAsis of their trAnslAtion ActivAtion by RHA is poorly understood. RHA contAins A conserved ATPAse-dependent HelicAse core thAt is flAnked by two α-β-β-β-α double-strAnded RNA-binding domAins At the N terminus And repeAted Arginine-glycine residues At the C terminus. The individuAl recombinAnt N-terminAl, centrAl HelicAse, And C-terminAl domAins were evAluAted for their Ability to specificAlly interAct with cognAte RNAs by in vitro biochemicAl meAsurements And mRNA trAnslAtion AssAys in cells. The results demonstrAte thAt N-terminAl residues confer selective interAction with retrovirAl And junD tArget RNAs. Conserved lysine residues in the distAl α-helix of the double-strAnded RNA-binding domAins Are necessAry to engAge structurAl feAtures of retrovirAl And junD 5'-UTRs. Exogenous expression of the N terminus coprecipitAtes junD mRNA And inhibits the trAnslAtion Activity of endogenous RHA. The results indicAte thAt the moleculAr bAsis for the ActivAtion of trAnslAtion by RHA is recognition of tArget mRNA by the N-terminAl domAin thAt tethers the ATP-dependent HelicAse for reArrAngement of the complex 5'-UTR.
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RNA HelicAse A modulAtes trAnslAtion of HIV-1 And infectivity of progeny virions
Nucleic acids research, 2009Co-Authors: Cheryl Bolinger, Amit Sharma, Deepali Singh, Kathleen Boris-lawrieAbstract:Retroviruses rely on host RNA-binding proteins to modulAte vArious steps in their replicAtion. Previously severAl AnimAl retroviruses were determined to mediAte Dhx9/RNA HelicAse A (RHA) interAction with A 5′ terminAl post-trAnscriptionAl control element (PCE) for efficient trAnslAtion. Herein PCE reporter AssAys determined HTLV-1 And HIV-1 RU5 confer orientAtion-dependent PCE Activity. The effect of Dhx9/RHA down-regulAtion And rescue with siRNA-resistAnt RHA on expression of HIV-1NL4–3 provirus determined thAt RHA is necessAry for efficient HIV-1 RNA trAnslAtion And requires ATPAse-dependent HelicAse function. QuAntitAtive AnAlysis determined HIV-1 RNA steAdy-stAte And cytoplAsmic AccumulAtion were not reduced; rAther the trAnslAtionAl Activity of virAl RNA wAs reduced. Western blotting determined thAt RHA-deficient virions Assemble with Lys-tRNA synthetAse, exhibit processed reverse trAnscriptAse And contAin similAr level of virAl RNA, but they Are poorly infectious on primAry lymphocytes And HeLA cells. The results demonstrAte RHA is An importAnt host fActor within the virus-producer cell And within the virAl pArticle. The identificAtion of RHA-dependent PCE Activity in cellulAr junD RNA And in six of seven generA of RetroviridAe suggests conservAtion of this trAnslAtionAl control mechAnism Among vertebrAtes, And convergent evolution of RetroviridAe to utilize this host mechAnism.
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MechAnisms employed by retroviruses to exploit host fActors for trAnslAtionAl control of A complicAted proteome
Retrovirology, 2009Co-Authors: Cheryl Bolinger, Kathleen Boris-lawrieAbstract:Retroviruses hAve evolved multiple strAtegies to direct the synthesis of A complex proteome from A single primAry trAnscript. Their mechAnisms Are modulAted by A breAdth of virus-host interActions, which Are of significAnt fundAmentAl interest becAuse they ultimAtely Affect the efficiency of virus replicAtion And diseAse pAthogenesis. Motifs locAted within the untrAnslAted region (UTR) of the retrovirAl RNA hAve estAblished roles in trAnscriptionAl trAns-ActivAtion, RNA pAckAging, And genome reverse trAnscription; And A growing literAture hAs reveAled A necessAry role of the UTR in modulAting the efficiency of virAl protein synthesis. ExAmples include A 5' UTR post-trAnscriptionAl control element (PCE), present in At leAst eight retroviruses, thAt interActs with cellulAr RNA HelicAse A to fAcilitAte cAp-dependent polyribosome AssociAtion; And 3' UTR constitutive trAnsport element (CTE) of MAson-Pfizer monkey virus thAt interActs with TAp/NXF1 And SR protein 9G8 to fAcilitAte RNA export And trAnslAtionAl utilizAtion. By contrAst, nucleAr protein hnRNP E1 negAtively modulAtes HIV-1 GAg, Env, And Rev protein synthesis. AlteRNAtive initiAtion strAtegies by ribosomAl frAmeshifting And leAky scAnning enAble polycistronic trAnslAtion of the cAp-dependent virAl trAnscript. Other studies posit cAp-independent trAnslAtion initiAtion by inteRNAl ribosome entry At structurAl feAtures of the 5' UTR of selected retroviruses. The retrovirAl ArmAmentArium Also commAnds mechAnisms to counter cellulAr post-trAnscriptionAl innAte defenses, including protein kinAse R, 2',5'-oligoAdenylAte synthetAse And the smAll RNA pAthwAy. This review will discuss recent And historicAlly-recognized insights into retrovirus trAnslAtionAl control. The expAnding knowledge of retrovirAl post-trAnscriptionAl control is vitAl to understAnding the biology of the retrovirAl proteome. In A broAd perspective, eAch new insight offers A prospective tArget for AntivirAl therApy And strAtegic improvement of gene trAnsfer vectors.
Flossie Wongstaal - One of the best experts on this subject based on the ideXlab platform.
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compArisons of RNAi ApproAches for vAlidAtion of humAn RNA HelicAse A As An essentiAl fActor in hepAtitis c virus replicAtion
Journal of Virological Methods, 2008Co-Authors: Qiuchen S He, Hengli Tang, Flossie Wongstaal, Jing Zhang, Ky Truong, Demin ZhouAbstract:A mAjor issue of current virology concerns the chArActerizAtion of cellulAr proteins thAt operAte As functionAl components of the virAl multiplicAtion process. RNAi is A powerful tool to elucidAte gene functions. In this study three RNAi ApproAches (trAnsient trAnsfection, stAble trAnsduction And inducible RNAi) were Assessed to vAlidAte humAn RNA HelicAse A (RHA) As An essentiAl fActor in hepAtitis C virus (HCV) replicAtion. It indicAted thAt RHA trAnsient knockdown by synthetic siRNA hAd no effect on HCV replicAtion, while RHA stAble knockdown viA lentivector trAnsduction cAused cell lethAlity. The involvement of RHA in HCV replicAtion wAs verified by An RNAi inducible system thAt, on the one hAnd, mAintAined long-term gene silencing, but on the other hAnd, AlleviAted siRNA toxicity during the essentiAl gene silencing. A 21-dAy follow-up of the response of HCV replicAtion to the presence And Absence of RNAi indicAted thAt RHA is A cellulAr fActor involved in the HCV replicAtion process.
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RNA HelicAse A interActs with nucleAr fActor κb p65 And functions As A trAnscriptionAl coActivAtor
FEBS Journal, 2004Co-Authors: Toshifumi Tetsuka, Flossie Wongstaal, Hiroaki Uranishi, Takaomi Sanda, Kaori Asamitsu, Jiangping Yang, Takashi OkamotoAbstract:RNA HelicAse A (RHA), A member of DNA And RNA HelicAse fAmily contAining ATPAse Activity, is involved in mAny steps of gene expression such As trAnscription And mRNA export. RHA hAs been reported to bind directly to the trAnscriptionAl coActivAtor, CREB-binding protein, And the tumor suppressor protein, BRCA1, And links them to RNA PolymerAse II holoenzyme complex. Using yeAst two-hybrid screening, we hAve identified RHA As An interActing molecule of the p65 subunit of nucleAr fActor kAppAB (NF-kAppAB). The interAction between p65 And RHA wAs confirmed by glutAthione-S trAnsferAse pull-down AssAy in vitro, And by co-immunoprecipitAtion AssAy in vivo. In trAnsient trAnsfection AssAys, RHA enhAnced NF-kAppAB dependent reporter gene expression induced by p65, tumor necrosis fActor-AlphA, or NF-kAppAB inducing kinAse. The mutAnt form of RHA lAcking ATP-binding Activity inhibited NF-kAppAB dependent reporter gene expression induced by these ActivAtors. Moreover, depletion of RHA using short interfering RNA reduced the NF-kAppAB dependent trAnsActivAtion. These dAtA suggest thAt RHA is An essentiAl component of the trAnsActivAtion complex by mediAting the trAnscriptionAl Activity of NF-kAppAB.
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Arginine methylAtion of RNA HelicAse A determines its subcellulAr locAlizAtion
Journal of Biological Chemistry, 2004Co-Authors: Wendell A Smith, Flossie Wongstaal, Brandon T Schurter, Michael DavidAbstract:RNA HelicAse A (RHA) undergoes nucleAr trAnslocAtion viA A clAssicAl import mechAnism utilizing kAryopherin betA. The nucleAr trAnsport domAin (NTD) of RHA is known to be necessAry And sufficient for its bi-directionAl nucleAr trAfficking. We report here thAt Arginine methylAtion is A novel requirement for NTD-mediAted nucleAr import. NucleAr trAnslocAtion of glutAthione S-trAnsferAse (GST)-NTD fusion proteins is AbrogAted by Arginine-methylAtion inhibitors. However, in vitro Arginine-methylAtion of GST-NTD prior to injection Allows the fusion protein to locAlize to the nucleus in the presence of methylAtion inhibitors. RemovAl of the Arginine-rich C-terminAl region negAtes the effects of the methylAtion inhibitors on NTD import, suggesting thAt methylAtion of the NTD C terminus the relieves the cytoplAsmic retention of RHA. The NTD physicAlly interActs with PRMT1, the mAjor protein Arginine methyltrAnsferAse. These findings provide evidence for A novel Arginine methylAtion-dependent regulAtory pAthwAy controlling the nucleAr import of RHA.
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A novel shuttle protein binds to RNA HelicAse A And ActivAtes the retrovirAl constitutive trAnsport element
Journal of Biological Chemistry, 2000Co-Authors: Christopher Westberg, Jianping Yang, Hengli Tang, Thipparthi R Reddy, Flossie WongstaalAbstract:The constitutive trAnsport element (CTE) of type D retroviruses mediAtes the nucleAr export of unspliced virAl trAnscripts. We previously showed thAt RNA HelicAse A functionAlly interActs with CTE And contAins A bidirectionAl nucleAr trAnsport domAin At the cArboxyl terminus. Here we report the identificAtion of A novel humAn protein, HelicAse A-binding protein 95 (HAP95), which specificAlly binds to the cArboxyl terminus of RNA HelicAse A. HAP95 is pArtiAlly homologous to AKAP95, A member of the A kinAse-Anchoring protein fAmily, but lAcks the protein kinAse A binding domAin chArActeristic of this fAmily. HAP95 is A nucleAr protein At steAdy stAte but shuttles between the nucleus And cytoplAsm. Overexpression of HAP95 significAntly increAses CTE-dependent gene expression but hAs no effect on generAl gene expression or thAt mediAted by the Rev/Rev response element of humAn immunodeficiency virus type 1.
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the cArboxyl terminus of RNA HelicAse A contAins A bidirectionAl nucleAr trAnsport domAin
Molecular and Cellular Biology, 1999Co-Authors: Hengli Tang, David Mcdonald, Tamara Middlesworth, Thomas J Hope, Flossie WongstaalAbstract:HumAn RNA HelicAse A wAs recently identified to be A shuttle protein which interActs with the constitutive trAnsport element (CTE) of type D retroviruses. Here we show thAt A domAin of 110 Amino Acids At the cArboxyl terminus of HelicAse A is both necessAry And sufficient for nucleAr locAlizAtion As well As rApid nucleAr export of glutAthione S-trAnsferAse fusion proteins. The import And export Activities of this domAin overlAp but Are sepArAble by point mutAtions. This bidirectionAl nucleAr trAnsport domAin (NTD) hAs no obvious sequence homology to previously identified nucleAr import or export signAls. However, the RAn-dependent nucleAr import of NTD wAs efficiently competed by excess Amounts of the nucleAr locAlizAtion signAl (NLS) peptide from simiAn virus 40 lArge T Antigen, suggesting thAt import is mediAted by the clAssicAl NLS pAthwAy. The nucleAr export pAthwAy Accessed by NTD is insensitive to leptomycin B And thus is distinct from the leucine-rich nucleAr export signAl pAthwAy mediAted by CRM1.
Cheryl Bolinger - One of the best experts on this subject based on the ideXlab platform.
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virion AssociAted host derived dhx9 RNA HelicAse A enhAnces the processivity of hiv 1 reverse trAnscriptAse on genomic RNA
Journal of Biological Chemistry, 2019Co-Authors: Samantha Brady, Cheryl Bolinger, Gatikrushna Singh, Zhenwei Song, Ioana Boeras, Kexin Weng, Bria Trent, William Clay Brown, Kamalendra Singh, Kathleen BorislawrieAbstract:DHX9/RNA HelicAse A (RHA) is A host RNA HelicAse thAt pArticipAtes in mAny criticAl steps of the HIV-1 life cycle. It co-Assembles with the virAl RNA genome into the cApsid core. Virions deficient in RHA Are less infectious As A result of reduced reverse trAnscription efficiency, demonstrAting thAt the virion-AssociAted RHA promotes reverse trAnscription before the virion gAins Access to the new host's RHA. Here, we quAntified reverse-trAnscription intermediAtes in HIV-1-infected T cells to clArify the mechAnism by which RHA enhAnces HIV-1 reverse trAnscription efficiency. Consistently, purified recombinAnt humAn RHA promoted reverse trAnscription efficiency under in vitro conditions thAt mimic the eArly reverse trAnscription steps prior to cApsid core uncoAting. We did not observe RHA-mediAted structurAl remodeling of the tRNALys3-virAl RNA-AnneAled complex. RHA did not enhAnce the DNA synthesis rAte until incorporAtion of the first few nucleotides, suggesting thAt RHA pArticipAtes primArily in the elongAtion phAse of reverse trAnscription. Pre-steAdy-stAte And steAdy-stAte kinetic studies reveAled thAt RHA hAs little impAct on the kinetics of single-nucleotide incorporAtion. Primer extension AssAys performed in the presence of trAp dsDNA disclosed thAt RHA enhAnces the processivity of HIV-1 reverse trAnscriptAse (RT). The biochemicAl AssAys used here effectively reflected And explAined the low RT Activity in HIV-1 virions produced from RHA-depleted cells. Moreover, RT Activity in our AssAys indicAted thAt RHA in HIV-1 virions is required for the efficient cAtAlysis of (-)cDNA synthesis during virAl infection before cApsid uncoAting. Our study identifies RHA As A processivity fActor of HIV-1 RT.
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RNA HelicAse A modulAtes trAnslAtion of HIV-1 And infectivity of progeny virions
Nucleic acids research, 2009Co-Authors: Cheryl Bolinger, Amit Sharma, Deepali Singh, Kathleen Boris-lawrieAbstract:Retroviruses rely on host RNA-binding proteins to modulAte vArious steps in their replicAtion. Previously severAl AnimAl retroviruses were determined to mediAte Dhx9/RNA HelicAse A (RHA) interAction with A 5′ terminAl post-trAnscriptionAl control element (PCE) for efficient trAnslAtion. Herein PCE reporter AssAys determined HTLV-1 And HIV-1 RU5 confer orientAtion-dependent PCE Activity. The effect of Dhx9/RHA down-regulAtion And rescue with siRNA-resistAnt RHA on expression of HIV-1NL4–3 provirus determined thAt RHA is necessAry for efficient HIV-1 RNA trAnslAtion And requires ATPAse-dependent HelicAse function. QuAntitAtive AnAlysis determined HIV-1 RNA steAdy-stAte And cytoplAsmic AccumulAtion were not reduced; rAther the trAnslAtionAl Activity of virAl RNA wAs reduced. Western blotting determined thAt RHA-deficient virions Assemble with Lys-tRNA synthetAse, exhibit processed reverse trAnscriptAse And contAin similAr level of virAl RNA, but they Are poorly infectious on primAry lymphocytes And HeLA cells. The results demonstrAte RHA is An importAnt host fActor within the virus-producer cell And within the virAl pArticle. The identificAtion of RHA-dependent PCE Activity in cellulAr junD RNA And in six of seven generA of RetroviridAe suggests conservAtion of this trAnslAtionAl control mechAnism Among vertebrAtes, And convergent evolution of RetroviridAe to utilize this host mechAnism.
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MechAnisms employed by retroviruses to exploit host fActors for trAnslAtionAl control of A complicAted proteome
Retrovirology, 2009Co-Authors: Cheryl Bolinger, Kathleen Boris-lawrieAbstract:Retroviruses hAve evolved multiple strAtegies to direct the synthesis of A complex proteome from A single primAry trAnscript. Their mechAnisms Are modulAted by A breAdth of virus-host interActions, which Are of significAnt fundAmentAl interest becAuse they ultimAtely Affect the efficiency of virus replicAtion And diseAse pAthogenesis. Motifs locAted within the untrAnslAted region (UTR) of the retrovirAl RNA hAve estAblished roles in trAnscriptionAl trAns-ActivAtion, RNA pAckAging, And genome reverse trAnscription; And A growing literAture hAs reveAled A necessAry role of the UTR in modulAting the efficiency of virAl protein synthesis. ExAmples include A 5' UTR post-trAnscriptionAl control element (PCE), present in At leAst eight retroviruses, thAt interActs with cellulAr RNA HelicAse A to fAcilitAte cAp-dependent polyribosome AssociAtion; And 3' UTR constitutive trAnsport element (CTE) of MAson-Pfizer monkey virus thAt interActs with TAp/NXF1 And SR protein 9G8 to fAcilitAte RNA export And trAnslAtionAl utilizAtion. By contrAst, nucleAr protein hnRNP E1 negAtively modulAtes HIV-1 GAg, Env, And Rev protein synthesis. AlteRNAtive initiAtion strAtegies by ribosomAl frAmeshifting And leAky scAnning enAble polycistronic trAnslAtion of the cAp-dependent virAl trAnscript. Other studies posit cAp-independent trAnslAtion initiAtion by inteRNAl ribosome entry At structurAl feAtures of the 5' UTR of selected retroviruses. The retrovirAl ArmAmentArium Also commAnds mechAnisms to counter cellulAr post-trAnscriptionAl innAte defenses, including protein kinAse R, 2',5'-oligoAdenylAte synthetAse And the smAll RNA pAthwAy. This review will discuss recent And historicAlly-recognized insights into retrovirus trAnslAtionAl control. The expAnding knowledge of retrovirAl post-trAnscriptionAl control is vitAl to understAnding the biology of the retrovirAl proteome. In A broAd perspective, eAch new insight offers A prospective tArget for AntivirAl therApy And strAtegic improvement of gene trAnsfer vectors.
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RNA HelicAse A is necessAry for trAnslAtion of selected messenger RNAs
Nature Structural & Molecular Biology, 2006Co-Authors: Tiffiney Roberts Hartman, Shuiming Qian, Cheryl Bolinger, Soledad Fernandez, Daniel R. Schoenberg, Kathleen BorislawrieAbstract:RNA HelicAse A (RHA) is A highly conserved DEAD-box protein thAt ActivAtes trAnscription, modulAtes RNA splicing And binds the nucleAr pore complex. The life cycle of typicAl mRNA involves RNA processing And trAnslAtion After ribosome scAnning of A relAtively unstructured 5′ untrAnslAted region (UTR). The precursor RNAs of retroviruses And selected cellulAr genes hArbor A complex 5′ UTR And use A yet-to-be-identified host post-trAnscriptionAl effector to stimulAte efficient trAnslAtion. Here we show thAt RHA recognizes A structured 5′-terminAl post-trAnscriptionAl control element (PCE) of A retrovirus And the JUND growth-control gene. RHA interActs with PCE RNA in the nucleus And cytoplAsm, fAcilitAtes polyribosome AssociAtion And is necessAry for its efficient trAnslAtion. Our results reveAl A previously unidentified role for RHA in trAnslAtion And implicAte RHA As An integrAtive effector in the continuum of gene expression from trAnscription to trAnslAtion.
Takaji Wakita - One of the best experts on this subject based on the ideXlab platform.
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RNA exosome complex regulAtes stAbility of the hepAtitis b virus x mRNA trAnscript in A non stop mediAted nsd RNA quAlity control mechAnism
Journal of Biological Chemistry, 2016Co-Authors: Hussein Hassan Aly, Junya Suzuki, Koichi Watashi, Kazuaki Chayama, Shinichi Hoshino, Makoto Hijikata, Takanobu Kato, Takaji WakitaAbstract:HepAtitis B virus (HBV) is A steAlth virus, minimAlly inducing the interferon system required for efficient induction of both innAte And AdAptive immune responses. However, 90% of Acutely infected Adults cAn cleAr the virus, suggesting the presence of other, interferon-independent pAthwAys leAding to virAl cleArAnce. Given the known Ability of HelicAses to bind virAl nucleic Acids, we performed A functionAl screening AssAy to identify HelicAses thAt regulAte HBV replicAtion. We identified the superkiller virAlicidic Activity 2-like (SKIV2L) RNA HelicAse (A homolog of the SAcchAromyces cerevisiAe Ski2 protein) on the bAsis of its direct And preferentiAl interAction with HBV X-mRNA. This interAction wAs essentiAl for HBV X-mRNA degrAdAtion At the RNA exosome. The degrAdAtion of HBV X-mRNA At the RNA exosome wAs Also mediAted by HBS1L (HBS1-like trAnslAtionAl GTPAse) protein, A known component of the host RNA quAlity control system. We found thAt the redundAnt HBV-precore trAnslAtion initiAtion site present At the 3'-end of HBV X-mRNA (3' precore) is trAnslAtionAlly Active. The initiAtion of trAnslAtion from this site without A proper stop codon wAs identified by the non-stop-mediAted RNA decAy mechAnism leAding to its degrAdAtion. Although 3' precore is present in the five mAin HBV-RNA trAnscripts, only X-mRNA lAcks the presence of An upstreAm stArt codons for lArge, middle, And smAll (L, M, And S) HBV surfAce proteins. These upstreAm codons Are in-frAme with 3' precore trAnslAtion initiAtion site, blocking its trAnslAtion from the other HBV-mRNA trAnscripts. To our knowledge, this is the first demonstrAtion of the Anti-virAl function of the non-stop-mediAted RNA decAy mechAnism.
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RNA exosome complex regulAtes stAbility of the hepAtitis b virus x mRNA trAnscript in A non stop mediAted nsd RNA quAlity control mechAnism
Journal of Biological Chemistry, 2016Co-Authors: Hussein Hassan Aly, Junya Suzuki, Koichi Watashi, Kazuaki Chayama, Shinichi Hoshino, Makoto Hijikata, Takanobu Kato, Takaji WakitaAbstract:AbstrAct HepAtitis B virus (HBV) is A steAlth virus, minimAlly inducing the Interferon system required for efficient induction of both innAte And AdAptive immune responses. However, 90% of Acutely infected Adults cAn cleAr the virus, suggesting the presence of other, interferon-independent pAthwAys leAding to virAl cleArAnce. Given the known Ability of HelicAses to bind virAl nucleic Acids, we performed A functionAl screening AssAy to identify HelicAses thAt regulAte HBV replicAtion. We identified the superkiller virAlicidic Activity 2-like (SKIV2L) RNA HelicAse (A homolog of the S. cerevisiAe Ski2 protein) on the bAsis of its direct And preferentiAl interAction with HBV X-mRNA. This interAction wAs essentiAl for HBV X-mRNA degrAdAtion At the RNA exosome. The degrAdAtion of HBV X-mRNA At the RNA exosome wAs Also mediAted by HBS1L protein, A known component of the host RNA quAlity control system. We found thAt the redundAnt HBV-precore trAnslAtion initiAtion site present At the 3′ end of HBV X-mRNA (3′ precore) is trAnslAtionAlly Active. The initiAtion of trAnslAtion from this site without A proper stop codon wAs identified by the Non-Stop mediAted RNA decAy mechAnism (NSD) leAding to its degrAdAtion. Although 3′ precore is present in the 5 mAin HBV-RNA trAnscripts; only X-mRNA lAcks the presence of An upstreAm stArt codons for LArge, Middle, And SmAll (L, M, And S) HBV surfAce proteins. These upstreAm codons Are in-frAme with 3′ precore trAnslAtion initiAtion site, blocking its trAnslAtion from the other HBV-mRNA trAnscripts. To our knowledge, this is the first demonstrAtion of the Anti-virAl function of NSD.
Gideon Dreyfuss - One of the best experts on this subject based on the ideXlab platform.
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the survivAl of motor neurons smn protein interActs with the snornp proteins fibrillArin And gAr1
Current Biology, 2001Co-Authors: Livio Pellizzoni, Jennifer Baccon, Bernard Charroux, Gideon DreyfussAbstract:AbstrAct BAckground: The survivAl of motor neurons (SMN) protein is the protein product of the spinAl musculAr Atrophy (SMA) diseAse gene. SMN And its AssociAted proteins Gemin2, Gemin3, And Gemin4 form A lArge complex thAt plAys A role in snRNP Assembly, pre-mRNA splicing, And trAnscription. The functions of SMN in these processes Are mediAted by A direct interAction of SMN with components of these mAchineries, such As Sm proteins And RNA HelicAse A. Results: We show thAt SMN binds directly to fibrillArin And GAR1. FibrillArin And GAR1 Are specific mArkers of the two clAsses of smAll nucleolAr ribonucleoprotein pArticles (snoRNPs) thAt Are involved in posttrAnscriptionAl processing And modificAtion of ribosomAl RNA. SMN interAction requires the Arginine- And glycine-rich domAins of both fibrillArin And GAR1 And is defective in SMN mutAnts found in some SMA pAtients. CoimmunoprecipitAtions demonstrAte thAt the SMN complex AssociAtes with fibrillArin And with GAR1 in vivo. The inhibition of RNA polymerAse I trAnscription cAuses A trAnsient redistribution of SMN to the nucleolAr periphery And loss of fibrillArin And GAR1 colocAlizAtion with SMN in gems. Furthermore, the expression of A dominAnt-negAtive mutAnt of SMN (SMNΔN27) cAuses snoRNPs to AccumulAte outside of the nucleolus in structures thAt Also contAin components of gems And coiled (CAjAl) bodies. Conclusions: These findings identify fibrillArin And GAR1 As novel interActors of SMN And suggest A function for the SMN complex in the Assembly And metAbolism of snoRNPs. We propose thAt the SMN complex performs functions necessAry for the biogenesis And function of diverse ribonucleoprotein complexes.
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A functionAl interAction between the survivAl motor neuron complex And RNA polymerAse ii
Journal of Cell Biology, 2001Co-Authors: Livio Pellizzoni, Bernard Charroux, Juri Rappsilber, Matthias Mann, Gideon DreyfussAbstract:The survivAl motor neuron (SMN) protein, the protein product of the spinAl musculAr Atrophy (SMA) diseAse gene, plAys A role in the Assembly And regenerAtion of smAll nucleAr ribonucleoproteins (snRNPs) And spliceosomes. By nAnoelectrosprAy mAss spectrometry, we identified RNA HelicAse A (RHA) As An SMN complex–AssociAted protein. RHA is A DEAH box RNA HelicAse which binds RNA polymerAse II (pol II) And reportedly functions in trAnscription. SMN interActs with RHA in vitro, And this interAction is impAired in mutAnt SMNs found in SMA pAtients. CoimmunoprecipitAtion demonstrAted thAt the SMN complex is AssociAted with pol II, snRNPs, And RHA in vivo. In vitro experiments suggest thAt RHA mediAtes the AssociAtion of SMN with the COOH-terminAl domAin of pol II. Moreover, trAnsfection of cells with A dominAnt negAtive mutAnt of SMN, SMNΔN27, cAuses AccumulAtion of pol II, snRNPs, And RHA in nucleAr structures thAt contAin the known mArkers of gems And coiled bodies, And inhibits RNA pol I And pol II trAnscription in vivo. These findings indicAte A functionAl As well As physicAl AssociAtion of the SMN complex with pol II And suggest A role for the SMN complex in the Assembly of the pol II trAnscription/processing mAchinery.
