The Experts below are selected from a list of 1146 Experts worldwide ranked by ideXlab platform
Jean-marc Egly - One of the best experts on this subject based on the ideXlab platform.
-
Distinct Regions of MAT1 Regulate cdk7 Kinase and TFIIH Transcription Activities
Journal of Biological Chemistry, 2000Co-Authors: Didier Busso, Arnaud Poterszman, Anne Keriel, Björn Sandrock, Opher Gileadi, Jean-marc EglyAbstract:Abstract THe transcription/DNA repair factor TFIIH may be resolved into at least two subcomplexes: tHe core TFIIH and tHe cdk-activating kinase (CAK) complex. THe CAK complex, wHicH is also found free in tHe cell, is composed of cdk7, Cyclin H, and MAT1. In tHe present work, we found tHat tHe C terminus of MAT1 binds to tHe cdk7·Cyclin H complex and activates tHe cdk7 kinase activity. THe median portion of MAT1, wHicH contains a coiled-coil motif, allows tHe binding of CAK to tHe TFIIH core tHrougH interactions witH botH XPD and XPB Helicases. FurtHermore, using recombinant TFIIH complexes, it is demonstrated tHat tHe N-terminal RING finger domain of MAT1 is crucial for transcription activation and participates to tHe pHospHorylation of tHe C-terminal domain of tHe largest subunit of tHe RNA polymerase II.
-
Stimulation of RARα Activation Function AF-1 tHrougH Binding to tHe General Transcription Factor TFIIH and PHospHorylation by CDK7
Cell, 1997Co-Authors: Cécile Rochette-egly, Jean-marc Egly, Michel Rossignol, Sylvie Adam, Pierre ChambonAbstract:Abstract THe activity of tHe N-terminal activation function AF-1 of RARα1 is abrogated upon mutation of a pHospHorylatable serine residue (Ser-77). Recombinant RARα was pHospHorylated by a variety of proline-directed protein kinases in vitro. However, only tHe coexpression of cdk7 stimulated Ser-77 pHospHorylation in vivo and enHanced transactivation by RARα, but not by a S77A RAR mutant. BotH free CAK (cdk7, Cyclin H, MAT1) and tHe CAK-containing general transcription factor TFIIH pHospHorylated Ser-77 in vitro. FurtHermore RARα bound free CAK and purified TFIIH in vitro, and RARα–TFIIH complexes could be isolated from HeLa nuclear extracts. THese findings represent tHe first example of activation of a transactivator tHrougH binding to and pHospHorylation by a general transcription factor.
-
Substrate specificity of tHe cdk-activating kinase (CAK) is altered upon association witH TFIIH.
The EMBO Journal, 1997Co-Authors: Michel Rossignol, Isabelle Kolb-cheynel, Jean-marc EglyAbstract:THe transcription/DNA repair factor TFIIH consists of nine subunits, several exHibiting known functions: Helicase/ATPase, kinase activity and DNA binding. THree subunits of TFIIH, cdk7, Cyclin H and MAT1, form a ternary complex, cdk-activating kinase (CAK), found eitHer on its own or as part of TFIIH. In tHe present work, we demonstrate tHat purified Human CAK complex (free CAK) and recombinant CAK (rCAK) produced in insect cells exHibit a strong preference for tHe Cyclin-dependent kinase 2 (cdk2) over a ctd oligopeptide substrate (wHicH mimics tHe carboxy-terminal domain of tHe RNA polymerase II). In contrast, TFIIH preferentially pHospHorylates tHe ctd as well as TFIIE alpHa, but not cdk2. TFIIH was resolved into four subcomplexes: tHe kinase complex composed of cdk7, Cyclin H and MAT1; tHe core TFIIH wHicH contains XPB, p62, p52, p44 and p34; and two otHer subcomplexes in wHicH XPD is found associated witH eitHer tHe kinase complex or witH tHe core TFIIH. Using tHese fractions, we demonstrate tHat TFIIH lacking tHe CAK subcomplex completely recovers its transcriptional activity in tHe presence of free CAK. FurtHermore, studies examining tHe interactions between TFIIH subunits provide evidence tHat CAK is integrated witHin TFIIH via XPB and XPD.
-
tHe structure of Cyclin H common mode of kinase activation and specific features
The EMBO Journal, 1997Co-Authors: G R Andersen, Arnaud Poterszman, J C Thierry, Jean-marc Egly, Didier Busso, Jae Ryoung Hwang, Jeanmarie Wurtz, Raymond Ripp, Dino MorasAbstract:THe crystal structure of Human Cyclin H refined at 2.6 A resolution is compared witH tHat of Cyclin A. THe core of tHe molecule consists of two repeats containing five Helices eacH and forming tHe canonical Cyclin fold also observed in TFIIB. One Hundred and tHirty‐two out of tHe 217 Cα atoms from tHe Cyclin fold can be superposed witH a root‐mean‐square difference of 1.8 A. THe structural Homology is even HigHer for tHe residues at tHe interface witH tHe kinase, wHicH is of functional significance, as sHown by our observation tHat Cyclin H binds to Cyclin‐dependent kinase 2 (cdk2) and tHat Cyclin A is able to activate cdk7 in tHe presence of MAT1. Based on tHis superposition, a new signature sequence for Cyclins was found. THe specificity of tHe Cyclin H molecule is provided mainly by two long Helices wHicH extend tHe Cyclin fold at its N‐ and C‐termini and pack togetHer against tHe first repeat on tHe side opposite to tHe kinase. Deletion mutants sHow tHat tHe terminal Helices are required for a functionally active Cyclin H.
-
expression in escHericHia coli purification and cHaracterization of Cyclin H a subunit of tHe Human general transcription dna repair factor tfiiH
Protein Expression and Purification, 1997Co-Authors: Arnaud Poterszman, Jean-marc Egly, Michel Rossignol, G R Andersen, Didier Busso, J C ThierryAbstract:Abstract THe Human Cyclin H, a protein normally associated witH tHe Cyclin-dependent kinase cdk7, was overexpressed in EscHericHia coli using a T7 RNA polymerase expression system and furtHer purified to apparent Homogeneity. THe purified recombinant Cyclin H is similar to tHe endogenous one according to tHe following criteria: molecular weigHt, microsequencing and mass spectra studies, ability to interact witH cdk7, and regulatory kinase activity. THe scale-up of Cyclin H purification is described.
Erich A Nigg - One of the best experts on this subject based on the ideXlab platform.
-
Cyclin-dependent kinase 7: at tHe cross-roads of transcription, DNA repair and cell cycle control?
Current Opinion in Cell Biology, 1996Co-Authors: Erich A NiggAbstract:Cyclin-dependent kinase (CDK) 7 was originally implicated in cell cycle control by virtue of its ability to pHospHorylate and activate otHer CDKs. Subsequently, botH CDK7 and its partner, Cyclin H, were found to be associated witH tHe general transcription factor TFIIH, suggesting additional roles for CDK7 in transcription and DNA repair. During tHe past year, a tHird subunit associated witH CDK7 and Cyclin H Has been cHaracterized, and tHe functional link between CDK7 and RNA polymerase II Has been strengtHened.
-
mat1 cdk7 and Cyclin H form a kinase complex wHicH is uv ligHt sensitive upon association witH tfiiH
The EMBO Journal, 1996Co-Authors: J P Adamczewski, Michel Rossignol, J P Tassan, Erich A Nigg, Vincent Moncollin, Jean-marc EglyAbstract:MAT1, Cyclin H and cdk7 are part of TFIIH, a class II transcription factor wHicH possesses numerous subunits of wHicH several Have been sHown to be involved in processes otHer tHan transcription. Two of tHem, XPD (ERCC2) and XPB (ERCC3), are Helicases involved in nucleotide excision repair (NER), wHereas cdk7, Cyclin H and MAT1 are tHougHt to participate in cell cycle regulation. MAT1, Cyclin H and cdk7 exist as a ternary complex eitHer free or associated witH TFIIH from wHicH tHe latter can be dissociated at HigH salt concentration. MAT1 is strongly associated witH cdk7 and Cyclin H. AltHougH not strictly required for tHe formation and activity of tHe complex, it stimulates its kinase activity. THe kinase activity of TFIIH, wHicH is constant during tHe cell cycle, is reduced after UV ligHt irradiation.
-
MAT1, cdk7 and Cyclin H form a kinase complex wHicH is UV ligHt‐sensitive upon association witH TFIIH.
The EMBO Journal, 1996Co-Authors: J P Adamczewski, Michel Rossignol, J P Tassan, Erich A Nigg, Vincent Moncollin, Jean-marc EglyAbstract:MAT1, Cyclin H and cdk7 are part of TFIIH, a class II transcription factor wHicH possesses numerous subunits of wHicH several Have been sHown to be involved in processes otHer tHan transcription. Two of tHem, XPD (ERCC2) and XPB (ERCC3), are Helicases involved in nucleotide excision repair (NER), wHereas cdk7, Cyclin H and MAT1 are tHougHt to participate in cell cycle regulation. MAT1, Cyclin H and cdk7 exist as a ternary complex eitHer free or associated witH TFIIH from wHicH tHe latter can be dissociated at HigH salt concentration. MAT1 is strongly associated witH cdk7 and Cyclin H. AltHougH not strictly required for tHe formation and activity of tHe complex, it stimulates its kinase activity. THe kinase activity of TFIIH, wHicH is constant during tHe cell cycle, is reduced after UV ligHt irradiation.
-
in vitro assembly of a functional Human cdk7 Cyclin H complex requires mat1 a novel 36 kda ring finger protein
The EMBO Journal, 1995Co-Authors: Jeanpierre Tassan, Malika Jaquenoud, S Frutiger, G J Hughes, Erich A NiggAbstract:It is proposed tHat tHe CDK7-Cyclin H complex functions in cell cycle progression, basal transcription and DNA repair. Here we report tHat in vitro reconstitution of an active CDK7-Cyclin H complex requires stoicHiometric amounts of a novel 36 kDa assembly factor termed MAT1 (menage a trois 1). Sequencing of MAT1 reveals a putative zinc binding motif (a C3HC4 RING finger) in tHe N-terminus; However, tHis domain is not required for ternary complex formation witH CDK7-Cyclin H. MAT1 is associated witH nuclear CDK7-Cyclin H at all stages of tHe cell cycle in vivo. Ternary complexes of CDK7, Cyclin H and MAT1 display kinase activity towards substrates mimicking botH tHe T-loop in CDKs and tHe C-terminal domain of RNA polymerase II, regardless of wHetHer tHey are immunoprecipitated from HeLa cells or reconstituted in a reticulocyte lysate. MAT1 constitutes tHe first example of an assembly factor tHat appears to be essential for tHe formation of an active CDK-Cyclin complex.
-
mat1 menage a trois a new ring finger protein subunit stabilizing Cyclin H cdk7 complexes in starfisH and xenopus cak
The EMBO Journal, 1995Co-Authors: Alain Devault, Annemarie Martinez, Jeanclaude Labbe, Erich A Nigg, Jeanpierre Tassan, Didier Fesquet, Nathalie Morin, J C Cavadore, Marcel DoreeAbstract:Abstract THe kinase responsible for THr161-THr160 pHospHorylation and activation of cdc2/cdk2 (CAK:cdk-activating kinase) Has been sHown previously to comprise at least two subunits, cdk7 and Cyclin H. An additional protein co-purified witH CAK in starfisH oocytes, but its sequencing did not reveal any similarity witH any known protein. In tHe present work, a cDNA encoding tHis protein is cloned and sequenced in botH starfisH and Xenopus oocytes. It is sHown to encode a new member of tHe RING finger family of proteins witH a cHaracteristic C3HC4 motif located in tHe N-terminal domain. We demonstrate tHat tHe RING finger protein (MAT1: 'menage a trois') is a new subunit of CAK in botH vertebrate and invertebrates. However, CAK may also exist in oocytes as Heterodimeric complexes between Cyclin H and cdk7 only. Stable Heterotrimeric CAK complexes were generated in reticulocyte lysates programmed witH mRNAs encoding Xenopus cdk7, Cyclin H and MAT1. In contrast, no Heterodimeric Cyclin H-cdk7 complex could be immunoprecipitated from reticulocyte lysates programmed witH cdk7 and Cyclin H mRNAs only. Stabilization of CAK complexes by MAT1 does not involve pHospHorylation of THr176, as tHe THr176-->Ala mutant of Xenopus cdk7 could engage as efficiently as wild-type cdk7 in ternary complexes. Even tHougH starfisH MAT1 is almost identical to Xenopus MAT1 in tHe RING finger domain, tHe starfisH subunit could not replace tHe Xenopus subunit and stabilize Cyclin H-cdk7 in reticulocyte lysate, suggesting tHat tHe MAT1 subunit does not (or not only) interact witH Cyclin H-cdk7 tHrougH tHe RING finger domain.
Jerard Hurwitz - One of the best experts on this subject based on the ideXlab platform.
-
isolation and cHaracterization of two Human transcription factor iiH tfiiH related complexes ercc2 cak and tfiiH
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Joyce T Reardon, Hui Ge, Aziz Sancar, Emma Gibbs, Jerard HurwitzAbstract:Abstract Transcription factor IIH (TFIIH) is a multisubunit protein complex essential for botH tHe initiation of RNA polymerase class II (pol II)-catalyzed transcription and nucleotide excision repair of DNA. Recent studies Have sHown tHat TFIIH copurifies witH tHe Cyclin-dependent kinase (cdk)-activating kinase complex (CAK) tHat includes cdk7, Cyclin H, and p36/MAT1. Here we report tHe isolation of two TFIIH-related complexes: TFIIH* and ERCC2/CAK. TFIIH* consists of a subset of tHe TFIIH complex proteins including ERCC3 (XPB), p62, p44, p41, and p34 but is devoid of detectable levels of ERCC2 (XPD) and CAK. ERCC2/CAK was isolated as a complex tHat exHibits CAK activity tHat cosediments witH tHe tHree CAK components (cdk7, Cyclin H, and p36/MAT1) as well as tHe ERCC2 (XPD) protein. TFIIH* can support pol II-catalyzed transcription in vitro witH lower efficiency compared witH TFIIH. THis TFIIH*-dependent transcription reaction was stimulated by ERCC2/CAK. THe ERCC2/CAK and TFIIH* complexes are eacH active in DNA repair as sHown by tHeir ability to complement extracts prepared from ERCC2 (XPD)- and ERCC3 (XPB)-deficient cells, respectively, in supporting tHe excision of DNA containing a cHolesterol lesion. THese data suggest tHat TFIIH* and ERCC2/CAK interact to form tHe TFIIH Holoenzyme capable of efficiently assembling tHe pol II transcription initiation complex and directly participating in excision repair reactions.
-
Isolation and cHaracterization of two Human transcription factor IIH (TFIIH)-related complexes: ERCC2/CAK and TFIIH.
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Joyce T Reardon, Hui Ge, Aziz Sancar, Emma Gibbs, Jerard HurwitzAbstract:Abstract Transcription factor IIH (TFIIH) is a multisubunit protein complex essential for botH tHe initiation of RNA polymerase class II (pol II)-catalyzed transcription and nucleotide excision repair of DNA. Recent studies Have sHown tHat TFIIH copurifies witH tHe Cyclin-dependent kinase (cdk)-activating kinase complex (CAK) tHat includes cdk7, Cyclin H, and p36/MAT1. Here we report tHe isolation of two TFIIH-related complexes: TFIIH* and ERCC2/CAK. TFIIH* consists of a subset of tHe TFIIH complex proteins including ERCC3 (XPB), p62, p44, p41, and p34 but is devoid of detectable levels of ERCC2 (XPD) and CAK. ERCC2/CAK was isolated as a complex tHat exHibits CAK activity tHat cosediments witH tHe tHree CAK components (cdk7, Cyclin H, and p36/MAT1) as well as tHe ERCC2 (XPD) protein. TFIIH* can support pol II-catalyzed transcription in vitro witH lower efficiency compared witH TFIIH. THis TFIIH*-dependent transcription reaction was stimulated by ERCC2/CAK. THe ERCC2/CAK and TFIIH* complexes are eacH active in DNA repair as sHown by tHeir ability to complement extracts prepared from ERCC2 (XPD)- and ERCC3 (XPB)-deficient cells, respectively, in supporting tHe excision of DNA containing a cHolesterol lesion. THese data suggest tHat TFIIH* and ERCC2/CAK interact to form tHe TFIIH Holoenzyme capable of efficiently assembling tHe pol II transcription initiation complex and directly participating in excision repair reactions.
Marcel Doree - One of the best experts on this subject based on the ideXlab platform.
-
Is Cdk7/Cyclin H/MAT1 tHe genuine cdk activating kinase in Cycling xenopus egg extracts?
Oncogene, 1997Co-Authors: Didier Fesquet, Marcel Doree, Nathalie Morin, Alain DevaultAbstract:: Formation of active cdk (Cyclin dependent kinase)/ Cyclin kinases involves pHospHorylation of a conserved tHreonine residue in tHe T loop of tHe cdk catalytic-subunit by CAK (Cdk Activating Kinase). CAK was first purified biocHemically from HigHer eukaryotes and identified as a trimeric complex containing a cdk7 catalytic subunit, Cyclin H and MAT1 (Menage a trois), a member of tHe RING finger family. THe same trimeric complex is also part of basal transcription factor TFIIH. In budding yeast, tHe closest Homologs of cdk7 and Cyclin H, KIN28 and CCL1, respectively, also associate witH TFIIH. However, tHe KIN28/CCL1 complex does not display CAK activity and a distinct protein kinase able to pHospHorylate monomeric CDC28 and GST-cdk2 was recently identified, cHallenging tHe identification of cdk7 as tHe pHysiological CAK in HigHer eukaryotes. Here we demonstrate tHat immunodepletion of cdk7 suppresses CAK activity from Cycling Xenopus egg extracts, and arrest tHem before M-pHase. We also sHow tHat specific translation of mRNAs encoding Xenopus cdk7 and its associated subunits restores CAK activity in cdk7-immunodepleted Xenopus egg extracts. Hence, tHe cdk7 complex is necessary and sufficient for activation of cdk-Cyclin complexes in Cycling Xenopus egg extracts.
-
is cdk7 Cyclin H mat1 tHe genuine cdk activating kinase in Cycling xenopus egg extracts
Oncogene, 1997Co-Authors: Didier Fesquet, Marcel Doree, Nathalie Morin, Alain DevaultAbstract:: Formation of active cdk (Cyclin dependent kinase)/ Cyclin kinases involves pHospHorylation of a conserved tHreonine residue in tHe T loop of tHe cdk catalytic-subunit by CAK (Cdk Activating Kinase). CAK was first purified biocHemically from HigHer eukaryotes and identified as a trimeric complex containing a cdk7 catalytic subunit, Cyclin H and MAT1 (Menage a trois), a member of tHe RING finger family. THe same trimeric complex is also part of basal transcription factor TFIIH. In budding yeast, tHe closest Homologs of cdk7 and Cyclin H, KIN28 and CCL1, respectively, also associate witH TFIIH. However, tHe KIN28/CCL1 complex does not display CAK activity and a distinct protein kinase able to pHospHorylate monomeric CDC28 and GST-cdk2 was recently identified, cHallenging tHe identification of cdk7 as tHe pHysiological CAK in HigHer eukaryotes. Here we demonstrate tHat immunodepletion of cdk7 suppresses CAK activity from Cycling Xenopus egg extracts, and arrest tHem before M-pHase. We also sHow tHat specific translation of mRNAs encoding Xenopus cdk7 and its associated subunits restores CAK activity in cdk7-immunodepleted Xenopus egg extracts. Hence, tHe cdk7 complex is necessary and sufficient for activation of cdk-Cyclin complexes in Cycling Xenopus egg extracts.
-
Interactions of Cyclins witH Cyclin-Dependent Kinases: A Common Interactive MecHanism†
Biochemistry, 1997Co-Authors: Frédéric Heitz, Jeanclaude Cavadore, Marcel Doree, Didier Fesquet, May C. Morris, Gilles DivitaAbstract:THe formation of cdk−Cyclin complexes Has been investigated at tHe molecular level and quantified using spectroscopic approacHes. In tHe absence of pHospHorylation, cdk2, cdc2, and cdk7 form HigHly stable complexes witH tHeir “natural” Cyclin partners witH dissociation constants in tHe nanomolar range. In contrast, nonpHospHorylated cdc2−Cyclin H, cdk2−Cyclin H, and cdk7−Cyclin A complexes present a 25-fold lower stability. On tHe basis of botH tHe structure of tHe cdk2−Cyclin A complex and on our kinetic results, we suggest tHat interaction of any Cyclin witH any cdk involves tHe same HydropHobic contacts and induces a marked conformational cHange in tHe catalytic cleft of tHe cdks. AltHougH cdks bind ATP strongly, tHey remain in a catalytically inactive conformation. In contrast, binding of tHe Cyclin induces structural rearrangements wHicH result in tHe selective reorientation of ATP, a concomitant 3-fold increase in its affinity, and a 5-fold decrease of its release from tHe active site of cdks.
-
dual pHospHorylation of tHe t loop in cdk7 its role in controlling Cyclin H binding and cak activity
The EMBO Journal, 1997Co-Authors: Annemarie Martinez, Mohammad Afshar, F Martin, Jeanclaude Cavadore, Jeanclaude Labbe, Marcel DoreeAbstract:A Cyclin-dependent kinase (cdk)-activating kinase (CAK) Has been sHown previously to catalyze T-loop pHospHorylation of cdks in most eukaryotic cells. THis enzyme exists in eitHer of two forms: tHe major one contains cdk7, Cyclin H and an assembly factor called MAT-1, wHilst tHe minor one lacks MAT-1. Cdk7 is unusual among cdks because it contains not one but two residues (S170 and T176 in Xenopus cdk7) in its T-loop tHat are pHospHorylated in vivo. We Have investigated tHe role of S170 and T176 pHospHorylation in tHe assembly and activity of Cyclin H-cdk7 dimers. In tHe absence of MAT-1, pHospHorylation of tHe T-loop appears to be required for cdk7 to bind Cyclin H. PHospHorylation of botH residues does not require Cyclin H binding in vitro. PHospHorylation of S170 is sufficient for cdk7 to bind Cyclin H witH low affinity, but HigH affinity binding requires T176 pHospHorylation. By mutational analysis, we demonstrate tHat in addition to its role in promotion of Cyclin H binding, S170 pHospHorylation plays a direct role in tHe control of CAK activity. Finally, we sHow tHat dual pHospHorylation of S170 and T176, or substitution of botH pHospHorylatable residues by aspartic residues, is sufficient to generate CAK activity to one-tHird of its maximal value in vitro, even in tHe absence of Cyclin H and MAT-1, and may tHus provide furtHer clues as to How Cyclins activate cdk subunits.
-
Dual pHospHorylation of tHe T‐loop in cdk7: its role in controlling Cyclin H binding and CAK activity
The EMBO Journal, 1997Co-Authors: Anne‐marie Martinez, Mohammad Afshar, F Martin, Jeanclaude Cavadore, Jeanclaude Labbe, Marcel DoreeAbstract:A Cyclin-dependent kinase (cdk)-activating kinase (CAK) Has been sHown previously to catalyze T-loop pHospHorylation of cdks in most eukaryotic cells. THis enzyme exists in eitHer of two forms: tHe major one contains cdk7, Cyclin H and an assembly factor called MAT-1, wHilst tHe minor one lacks MAT-1. Cdk7 is unusual among cdks because it contains not one but two residues (S170 and T176 in Xenopus cdk7) in its T-loop tHat are pHospHorylated in vivo. We Have investigated tHe role of S170 and T176 pHospHorylation in tHe assembly and activity of Cyclin H-cdk7 dimers. In tHe absence of MAT-1, pHospHorylation of tHe T-loop appears to be required for cdk7 to bind Cyclin H. PHospHorylation of botH residues does not require Cyclin H binding in vitro. PHospHorylation of S170 is sufficient for cdk7 to bind Cyclin H witH low affinity, but HigH affinity binding requires T176 pHospHorylation. By mutational analysis, we demonstrate tHat in addition to its role in promotion of Cyclin H binding, S170 pHospHorylation plays a direct role in tHe control of CAK activity. Finally, we sHow tHat dual pHospHorylation of S170 and T176, or substitution of botH pHospHorylatable residues by aspartic residues, is sufficient to generate CAK activity to one-tHird of its maximal value in vitro, even in tHe absence of Cyclin H and MAT-1, and may tHus provide furtHer clues as to How Cyclins activate cdk subunits.
Joyce T Reardon - One of the best experts on this subject based on the ideXlab platform.
-
isolation and cHaracterization of two Human transcription factor iiH tfiiH related complexes ercc2 cak and tfiiH
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Joyce T Reardon, Hui Ge, Aziz Sancar, Emma Gibbs, Jerard HurwitzAbstract:Abstract Transcription factor IIH (TFIIH) is a multisubunit protein complex essential for botH tHe initiation of RNA polymerase class II (pol II)-catalyzed transcription and nucleotide excision repair of DNA. Recent studies Have sHown tHat TFIIH copurifies witH tHe Cyclin-dependent kinase (cdk)-activating kinase complex (CAK) tHat includes cdk7, Cyclin H, and p36/MAT1. Here we report tHe isolation of two TFIIH-related complexes: TFIIH* and ERCC2/CAK. TFIIH* consists of a subset of tHe TFIIH complex proteins including ERCC3 (XPB), p62, p44, p41, and p34 but is devoid of detectable levels of ERCC2 (XPD) and CAK. ERCC2/CAK was isolated as a complex tHat exHibits CAK activity tHat cosediments witH tHe tHree CAK components (cdk7, Cyclin H, and p36/MAT1) as well as tHe ERCC2 (XPD) protein. TFIIH* can support pol II-catalyzed transcription in vitro witH lower efficiency compared witH TFIIH. THis TFIIH*-dependent transcription reaction was stimulated by ERCC2/CAK. THe ERCC2/CAK and TFIIH* complexes are eacH active in DNA repair as sHown by tHeir ability to complement extracts prepared from ERCC2 (XPD)- and ERCC3 (XPB)-deficient cells, respectively, in supporting tHe excision of DNA containing a cHolesterol lesion. THese data suggest tHat TFIIH* and ERCC2/CAK interact to form tHe TFIIH Holoenzyme capable of efficiently assembling tHe pol II transcription initiation complex and directly participating in excision repair reactions.
-
Isolation and cHaracterization of two Human transcription factor IIH (TFIIH)-related complexes: ERCC2/CAK and TFIIH.
Proceedings of the National Academy of Sciences of the United States of America, 1996Co-Authors: Joyce T Reardon, Hui Ge, Aziz Sancar, Emma Gibbs, Jerard HurwitzAbstract:Abstract Transcription factor IIH (TFIIH) is a multisubunit protein complex essential for botH tHe initiation of RNA polymerase class II (pol II)-catalyzed transcription and nucleotide excision repair of DNA. Recent studies Have sHown tHat TFIIH copurifies witH tHe Cyclin-dependent kinase (cdk)-activating kinase complex (CAK) tHat includes cdk7, Cyclin H, and p36/MAT1. Here we report tHe isolation of two TFIIH-related complexes: TFIIH* and ERCC2/CAK. TFIIH* consists of a subset of tHe TFIIH complex proteins including ERCC3 (XPB), p62, p44, p41, and p34 but is devoid of detectable levels of ERCC2 (XPD) and CAK. ERCC2/CAK was isolated as a complex tHat exHibits CAK activity tHat cosediments witH tHe tHree CAK components (cdk7, Cyclin H, and p36/MAT1) as well as tHe ERCC2 (XPD) protein. TFIIH* can support pol II-catalyzed transcription in vitro witH lower efficiency compared witH TFIIH. THis TFIIH*-dependent transcription reaction was stimulated by ERCC2/CAK. THe ERCC2/CAK and TFIIH* complexes are eacH active in DNA repair as sHown by tHeir ability to complement extracts prepared from ERCC2 (XPD)- and ERCC3 (XPB)-deficient cells, respectively, in supporting tHe excision of DNA containing a cHolesterol lesion. THese data suggest tHat TFIIH* and ERCC2/CAK interact to form tHe TFIIH Holoenzyme capable of efficiently assembling tHe pol II transcription initiation complex and directly participating in excision repair reactions.
