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Adam M Fontebasso,
Jeremy Schwartzentruber,
Dong-Anh Khuong-Quang,
Xiao-Yang Liu,
Dominik Sturm,
Andrey Korshunov,
David T W Jones,
Hendrik Witt,
Marcel Kool,
Steffen Albrecht, [......],
Pawel P Liberski,
Peter Hauser,
Miklos Garami,
Almos Klekner,
Laszlo Bognar,
Gelareh Zadeh, Damien Faury,
Stefan M Pfister,
Nada Jabado,
Jacek Majewski
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ABSTRACT: Recurrent mutations affecting the histone H3.3 residues Lys27 or indirectly Lys36 are frequent drivers of pediatric high-grade gliomas (over 30 % of HGGs). To identify additional driver mutations in HGGs, we investigated a cohort of 60 pediatric HGGs using whole-exome sequencing (WES) and compared them to 543 exomes from non-cancer control samples. We identified mutations in SETD2, a H3K36 trimethyltransferase, in 15 % of pediatric HGGs, a result that was genome-wide significant (FDR = 0.029). Most SETD2 alterations were truncating mutations. Sequencing the gene in this cohort and another validation cohort (123 gliomas from all ages and grades) showed SETD2 mutations to be specific to high-grade tumors affecting 15 % of pediatric HGGs (11/73) and 8 % of adult HGGs (5/65) while no SETD2 mutations were identified in low-grade diffuse gliomas (0/45). Furthermore, SETD2 mutations were mutually exclusive with H3F3A mutations in HGGs (P = 0.0492) while they partly overlapped with IDH1 mutations (4/14), and SETD2-mutant tumors were found exclusively in the cerebral hemispheres (P = 0.0055). SETD2 is the only H3K36 trimethyltransferase in humans, and SETD2-mutant tumors showed a substantial decrease in H3K36me3 levels (P < 0.001), indicating that the mutations are loss-of-function. These data suggest that loss-of-function SETD2 mutations occur in older children and young adults and are specific to HGG of the cerebral cortex, similar to the H3.3 G34R/V and IDH mutations. Taken together, our results suggest that mutations disrupting the histone code at H3K36, including H3.3 G34R/V, IDH1 and/or SETD2 mutations, are central to the genesis of hemispheric HGGs in older children and young adults.
Acta Neuropathologica 02/2013; · 9.32 Impact Factor
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Dominik Sturm,
Hendrik Witt,
Volker Hovestadt,
Dong-Anh Khuong-Quang,
David T W Jones,
Carolin Konermann,
Elke Pfaff,
Martje Tönjes,
Martin Sill,
Sebastian Bender, [......],
Tom Mikkelsen,
Kenneth Aldape,
Guido Reifenberger,
V Peter Collins,
Jacek Majewski,
Andrey Korshunov,
Peter Lichter,
Christoph Plass,
Nada Jabado,
Stefan M Pfister
[show abstract]
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ABSTRACT: Glioblastoma (GBM) is a brain tumor that carries a dismal prognosis and displays considerable heterogeneity. We have recently identified recurrent H3F3A mutations affecting two critical amino acids (K27 and G34) of histone H3.3 in one-third of pediatric GBM. Here, we show that each H3F3A mutation defines an epigenetic subgroup of GBM with a distinct global methylation pattern, and that they are mutually exclusive with IDH1 mutations, which characterize a third mutation-defined subgroup. Three further epigenetic subgroups were enriched for hallmark genetic events of adult GBM and/or established transcriptomic signatures. We also demonstrate that the two H3F3A mutations give rise to GBMs in separate anatomic compartments, with differential regulation of transcription factors OLIG1, OLIG2, and FOXG1, possibly reflecting different cellular origins.
Cancer cell 10/2012; 22(4):425-37. · 25.29 Impact Factor
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Xiao-Yang Liu,
Noha Gerges,
Andrey Korshunov,
Nesrin Sabha,
Dong-Anh Khuong-Quang,
Adam M Fontebasso,
Adam Fleming,
Djihad Hadjadj,
Jeremy Schwartzentruber,
Jacek Majewski, [......],
Steffen Albrecht,
Sidney Croul,
David Tw Jones,
Marcel Kool,
Martje Tonjes,
Guido Reifenberger, Damien Faury,
Gelareh Zadeh,
Stefan Pfister,
Nada Jabado
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ABSTRACT: Gliomas are the most common primary brain tumors in children and adults. We recently identified frequent alterations in chromatin remodelling pathways including recurrent mutations in H3F3A and mutations in ATRX (α-thalassemia/mental-retardation-syndrome-X-linked) in pediatric and young adult glioblastoma (GBM, WHO grade IV astrocytoma). H3F3A mutations were specific to pediatric high-grade gliomas and identified in only 3.4 % of adult GBM. Using sequencing and/or immunohistochemical analyses, we investigated ATRX alterations (mutation/loss of expression) and their association with TP53 and IDH1 or IDH2 mutations in 140 adult WHO grade II, III and IV gliomas, 17 pediatric WHO grade II and III astrocytomas and 34 pilocytic astrocytomas. In adults, ATRX aberrations were detected in 33 % of grade II and 46 % of grade III gliomas, as well as in 80 % of secondary and 7 % of primary GBMs. They were absent in the 17 grade II and III astrocytomas in children, and the 34 pilocytic astrocytomas. ATRX alterations closely overlapped with mutations in IDH1/2 (p < 0.0001) and TP53 (p < 0.0001) in samples across all WHO grades. They were prevalent in astrocytomas and oligoastrocytomas, but were absent in oligodendrogliomas (p < 0.0001). No significant association of ATRX mutation/loss of expression and alternative lengthening of telomeres was identified in our cohort. In summary, our data show that ATRX alterations are frequent in adult diffuse gliomas and are specific to astrocytic tumors carrying IDH1/2 and TP53 mutations. Combined alteration of these genes may contribute to drive the neoplastic growth in a major subset of diffuse astrocytomas in adults.
Acta Neuropathologica 08/2012; 124(5):615-25. · 9.32 Impact Factor
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Dong-Anh Khuong-Quang,
Pawel Buczkowicz,
Patricia Rakopoulos,
Xiao-Yang Liu,
Adam M Fontebasso,
Eric Bouffet,
Ute Bartels,
Steffen Albrecht,
Jeremy Schwartzentruber,
Louis Letourneau, [......],
Marcel Kool,
Andreas von Deimling,
Dominik Sturm,
Andrey Korshunov, Damien Faury,
David T Jones,
Jacek Majewski,
Stefan M Pfister,
Nada Jabado,
Cynthia Hawkins
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ABSTRACT: Pediatric glioblastomas (GBM) including diffuse intrinsic pontine gliomas (DIPG) are devastating brain tumors with no effective therapy. Here, we investigated clinical and biological impacts of histone H3.3 mutations. Forty-two DIPGs were tested for H3.3 mutations. Wild-type versus mutated (K27M-H3.3) subgroups were compared for HIST1H3B, IDH, ATRX and TP53 mutations, copy number alterations and clinical outcome. K27M-H3.3 occurred in 71 %, TP53 mutations in 77 % and ATRX mutations in 9 % of DIPGs. ATRX mutations were more frequent in older children (p < 0.0001). No G34V/R-H3.3, IDH1/2 or H3.1 mutations were identified. K27M-H3.3 DIPGs showed specific copy number changes, including all gains/amplifications of PDGFRA and MYC/PVT1 loci. Notably, all long-term survivors were H3.3 wild type and this group of patients had better overall survival. K27M-H3.3 mutation defines clinically and biologically distinct subgroups and is prevalent in DIPG, which will impact future therapeutic trial design. K27M- and G34V-H3.3 have location-based incidence (brainstem/cortex) and potentially play distinct roles in pediatric GBM pathogenesis. K27M-H3.3 is universally associated with short survival in DIPG, while patients wild-type for H3.3 show improved survival. Based on prognostic and therapeutic implications, our findings argue for H3.3-mutation testing at diagnosis, which should be rapidly integrated into the clinical decision-making algorithm, particularly in atypical DIPG.
Acta Neuropathologica 06/2012; 124(3):439-47. · 9.32 Impact Factor
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Jeremy Schwartzentruber,
Andrey Korshunov,
Xiao-Yang Liu,
David T W Jones,
Elke Pfaff,
Karine Jacob,
Dominik Sturm,
Adam M Fontebasso,
Dong-Anh Khuong Quang,
Martje Tönjes, [......],
Olaf Witt,
Cindy Zhang,
Pedro Castelo-Branco,
Peter Lichter, Damien Faury,
Uri Tabori,
Christoph Plass,
Jacek Majewski,
Stefan M Pfister,
Nada Jabado
Nature 03/2012; · 36.28 Impact Factor
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Jeremy Schwartzentruber,
Andrey Korshunov,
Xiao-Yang Liu,
David T W Jones,
Elke Pfaff,
Karine Jacob,
Dominik Sturm,
Adam M Fontebasso,
Dong-Anh Khuong Quang,
Martje Tönjes, [......],
Olaf Witt,
Cindy Zhang,
Pedro Castelo-Branco,
Peter Lichter, Damien Faury,
Uri Tabori,
Christoph Plass,
Jacek Majewski,
Stefan M Pfister,
Nada Jabado
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ABSTRACT: Glioblastoma multiforme (GBM) is a lethal brain tumour in adults and children. However, DNA copy number and gene expression signatures indicate differences between adult and paediatric cases. To explore the genetic events underlying this distinction, we sequenced the exomes of 48 paediatric GBM samples. Somatic mutations in the H3.3-ATRX-DAXX chromatin remodelling pathway were identified in 44% of tumours (21/48). Recurrent mutations in H3F3A, which encodes the replication-independent histone 3 variant H3.3, were observed in 31% of tumours, and led to amino acid substitutions at two critical positions within the histone tail (K27M, G34R/G34V) involved in key regulatory post-translational modifications. Mutations in ATRX (α-thalassaemia/mental retardation syndrome X-linked) and DAXX (death-domain associated protein), encoding two subunits of a chromatin remodelling complex required for H3.3 incorporation at pericentric heterochromatin and telomeres, were identified in 31% of samples overall, and in 100% of tumours harbouring a G34R or G34V H3.3 mutation. Somatic TP53 mutations were identified in 54% of all cases, and in 86% of samples with H3F3A and/or ATRX mutations. Screening of a large cohort of gliomas of various grades and histologies (n = 784) showed H3F3A mutations to be specific to GBM and highly prevalent in children and young adults. Furthermore, the presence of H3F3A/ATRX-DAXX/TP53 mutations was strongly associated with alternative lengthening of telomeres and specific gene expression profiles. This is, to our knowledge, the first report to highlight recurrent mutations in a regulatory histone in humans, and our data suggest that defects of the chromatin architecture underlie paediatric and young adult GBM pathogenesis.
Nature 02/2012; 482(7384):226-31. · 36.28 Impact Factor
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ABSTRACT: Graft-versus-host disease is uncommon in autologous hematopoietic cell transplantation (HCT) and is typically brief and mild. We report unusual, protracted, and severe Omenn syndrome-like autoaggression following autologous HCT. We identified a profound FOXP3(+) regulatory T cell defect that coincided with hyperinflammatory T cell responses which were reversible with rapamycin in vitro.
Clinical and vaccine immunology: CVI 11/2011; 19(1):109-12. · 2.37 Impact Factor
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Karine Jacob,
Dongh-Anh Quang-Khuong,
David T W Jones,
Hendrik Witt,
Sally Lambert,
Steffen Albrecht,
Olaf Witt,
Catherine Vezina,
Margret Shirinian, Damien Faury, [......],
Laszlo Bognar,
Jean-Pierre Farmer,
Jose-Luis Montes,
Jeffrey Atkinson,
Cynthia Hawkins,
Andrey Korshunov,
V Peter Collins,
Stefan M Pfister,
Uri Tabori,
Nada Jabado
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ABSTRACT: Oncogenic BRAF/Ras or NF1 loss can potentially trigger oncogene-induced senescence (OIS) through activation of the mitogen-activated protein kinase (MAPK) pathway. Somatic genetic abnormalities affecting this pathway occur in the majority of pilocytic astrocytomas (PA), the most prevalent brain neoplasm in children. We investigated whether OIS is induced in PA.
We tested expression of established senescence markers in three independent cohorts of sporadic PA. We also assessed for OIS in vitro, using forced expression of wild-type and V600E-mutant BRAF in two astrocytic cell lines: human telomerase reverse transcriptase (hTERT)-immortalized astrocytes and fetal astrocytes.
Our results indicate that PAs are senescent as evidenced by marked senescence-associated acidic β-galactosidase activity, low KI-67 index, and induction of p16(INK4a) but not p53 in the majority of 52 PA samples (46 of 52; 88.5%). Overexpression of a number of senescence-associated genes [CDKN2A (p16), CDKN1A (p21), CEBPB, GADD45A, and IGFBP7] was shown at the mRNA level in two independent PA tumor series. In vitro, sustained activation of wild-type or mutant BRAF induced OIS in both astrocytic cell lines. Loss of p16(INK4a) in immortalized astrocytes abrogated OIS, indicative of the role of this pathway in mediating this phenomenon in astrocytes. OIS is a mechanism of tumor suppression that restricts the progression of benign tumors. We show that it is triggered in PAs through p16(INK4a) pathway induction following aberrant MAPK activation.
OIS may account for the slow growth pattern in PA, the lack of progression to higher-grade astrocytomas, and the high overall survival of affected patients.
Clinical Cancer Research 05/2011; 17(14):4650-60. · 7.74 Impact Factor
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Hui-Qi Qu,
Karine Jacob,
Sarah Fatet,
Bing Ge,
David Barnett,
Olivier Delattre, Damien Faury,
Alexandre Montpetit,
Lauren Solomon,
Peter Hauser,
Miklos Garami,
Laszlo Bognar,
Zoltan Hansely,
Robert Mio,
Jean-Pierre Farmer,
Steffen Albrecht,
Constantin Polychronakos,
Cynthia Hawkins,
Nada Jabado
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ABSTRACT: Available data on genetic events in pediatric grade IV astrocytomas (glioblastoma [pGBM]) are scarce. This has traditionally been a major impediment in understanding the pathogenesis of this tumor and in developing ways for more effective management. Our aim is to chart DNA copy number aberrations (CNAs) and get insight into genetic pathways involved in pGBM. Using the Illumina Infinium Human-1 bead-chip-array (100K single-nucleotide polymorphisms [SNPs]), we genotyped 18 pediatric and 6 adult GBMs. Results were compared to BAC-array profiles harvested on 16 of the same pGBM, to an independent data set of 9 pediatric high-grade astrocytomas (HGAs) analyzed on Affymetrix 250K-SNP arrays, and to existing data sets on HGAs. CNAs were additionally validated by real-time qPCR in a set of genes in pGBM. Our results identify with nonrandom clustering of CNAs in several novel, previously not reported, genomic regions, suggesting that alterations in tumor suppressors and genes involved in the regulation of RNA processing and the cell cycle are major events in the pathogenesis of pGBM. Most regions were distinct from CNAs in aGBMs and show an unexpectedly low frequency of genetic amplification and homozygous deletions and a high frequency of loss of heterozygosity for a high-grade I rapidly dividing tumor. This first, complete, high-resolution profiling of the tumor cell genome fills an important gap in studies on pGBM. It ultimately guides the mapping of oncogenic networks unique to pGBM, identification of the related therapeutic predictors and targets, and development of more effective therapies. It further shows that, despite commonalities in a few CNAs, pGBM and aGBMs are two different diseases.
Neuro-Oncology 02/2010; 12(2):153-63. · 5.72 Impact Factor
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ABSTRACT: Glioblastoma multiforme (GBM) is an aggressive type of brain tumor where <3% of newly diagnosed cases in the patients will survive >5 years. In adults, GBM is the most common type of brain tumor. It is rarer in children, where it constitutes approximately 15% of all brain tumors diagnosed. These tumors are often invasive, making surgical resection difficult. Further, they can be refractory to current therapies such as temozolomide. The current dogma is that temozolomide resistance rests on the expression of O6-methylguanine-DNA methyltransferase (MGMT) because it cleaves methylated DNA adducts formed by the drug. Our laboratory recently reported that another drug resistance gene known as the Y-box binding protein-1 (YB-1) is highly expressed in primary GBM but not in normal brain tissues based on the evaluation of primary tumors. We therefore questioned whether GBM depend on YB-1 for growth and/or response to temozolomide. Herein, we report that YB-1 inhibition reduced tumor cell invasion and growth in monolayer as well as in soft agar. Moreover, blocking this protein ultimately delayed tumor onset in mice. Importantly, inhibiting YB-1 enhanced temozolomide sensitivity in a manner that was independent of MGMT in models of adult and pediatric GBM. In conclusion, inhibiting YB-1 may be a novel way to improve the treatment of GBM.
Molecular Cancer Therapeutics 12/2009; 8(12):3276-84. · 5.23 Impact Factor
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Takrima Haque, Damien Faury,
Steffen Albrecht,
Enrique Lopez-Aguilar,
Péter Hauser,
Miklós Garami,
Zoltán Hanzély,
László Bognár,
Rolando F Del Maestro,
Jeffrey Atkinson,
Andre Nantel,
Nada Jabado
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ABSTRACT: Gene expression profiling has proved crucial for understanding the biology of cancer. In rare diseases, including pediatric glioblastoma (pGBM), the lack of readily available fresh frozen (FF) material limits the feasibility of this analysis, as well as its validation, on independent data sets, a step needed to ensure relevance, mandating the use of alternate RNA sources. To overcome the limitation of material number and to validate results we obtained on FF pGBM, we did microarray analysis on RNA extracted from formalin-fixed, paraffin-embedded archival samples from pGBM and control brains, wherein we had no control on the fixation process.
RNA from 16 pGBM and 3 control brains was extracted and linearly amplified. Reverse transcription-PCR on housekeeping and formerly identified tumor-associated genes and microarray analysis were done on this RNA source. Results were validated by immunohistochemistry.
Despite extensive RNA degradation, microarray analysis was possible on 16 of 19 samples and reproduced the pattern of results obtained on FF pGBM. Gene lists and ontology subgrouping were highly concordant in both sample types. Similar to the findings on FF samples, we were able to identify two subsets of pGBM based on their association/lack of association with evidence consistent with an active Ras pathway.
Archival formalin-fixed, paraffin-embedded tissues are an invaluable resource as they are the most widely available materials often accessible in conjunction with clinical and follow-up data. Gene expression profiling on this material is feasible and may represent a significant advance for understanding the biology of rare human diseases.
Clinical Cancer Research 12/2007; 13(21):6284-92. · 7.74 Impact Factor
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Damien Faury,
André Nantel,
Sandra E Dunn,
Marie-Christine Guiot,
Takrima Haque,
Péter Hauser,
Miklós Garami,
László Bognár,
Zoltán Hanzély,
Pawel P Liberski,
Enrique Lopez-Aguilar,
Elvis T Valera,
Luis G Tone,
Anne-Sophie Carret,
Rolando F Del Maestro,
Martin Gleave,
Jose-Luis Montes,
Torsten Pietsch,
Stephen Albrecht,
Nada Jabado
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ABSTRACT: Pediatric glioblastoma (pGBM) is a rare, but devastating brain tumor. In contrast to GBM in adults (aGBM), little is known about the mechanisms underlying its development. Our aim is to gain insight into the molecular pathways of pGBM.
Thirty-two pGBM and seven aGBM samples were investigated using biochemical and transcriptional profiling. Ras and Akt pathway activation was assessed through the phosphorylation of downstream effectors, and gene expression profiles were generated using the University Health Network Human 19K cDNA arrays. Results were validated using real-time polymerase chain reaction and immunohistochemistry and compared with existing data sets on aGBM.
There are at least two subsets of pGBM. One subset, associated with Ras and Akt pathway activation, has very poor prognosis and exhibits increased expression of genes related to proliferation and to a neural stem-cell phenotype, similar to findings in aggressive aGBM. This subset was still molecularly distinguishable from aGBM after unsupervised and supervised analysis of expression profiles. A second subset, with better prognosis, is not associated with activation of Akt and Ras pathways, may originate from astroglial progenitors, and does not express gene signatures and markers shown to be associated with long-term survival in aGBM. Both subsets of pGBM show overexpression of Y-box-protein-1 that may help drive oncogenesis in this tumor.
Our work, the first study of gene expression profiles in pGBM, provides valuable insight into active pathways and targets in a cancer with minimal survival, and suggests that these tumors cannot be understood exclusively through studies of aGBM.
Journal of Clinical Oncology 04/2007; 25(10):1196-208. · 18.37 Impact Factor
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ABSTRACT: Malignant central nervous system (CNS) tumors, such as glioblastoma multiforme, invade the brain and disrupt normal tissue architecture, making complete surgical removal virtually impossible. Here, we have developed and optimized a purification strategy to isolate and identify natural inhibitors of glioma cell invasion in a three-dimensional collagen type I matrix. Inter alpha-trypsin inhibitor heavy chain 2 (ITI H2) was identified from the most inhibitory fractions and its presence was confirmed both as a single protein and in a bikunin-bound form. Stable overexpression in U251 glioma cells validated ITI H2's strong inhibition of human glioma cell invasion together with significant inhibition of cell proliferation and promotion of cell-cell adhesion. Analysis of primary human brain tumors showed significantly higher levels of ITI H2 in normal brain and low-grade tumors compared with high-grade gliomas, indicating an inverse correlation with malignancy. The phosphatidylinositol 3-kinase/Akt signaling cascade seemed to be one of the pathways involved in the effect of ITI H2 on U251 cells. These findings suggest that reduction of ITI H2 expression correlates with brain tumor progression and that targeting factors responsible for its loss or restoring the ITI supply exogenously may serve as potential therapeutic strategies for a variety of CNS tumors.
Cancer Research 03/2006; 66(3):1464-72. · 7.86 Impact Factor
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ABSTRACT: Xylanase C (XlnC) is a cofactorless protein secreted through the twin arginine translocation (Tat)-dependent secretion pathway by Streptomyces lividans. Its signal peptide contains the SRRGFLG sequence, which is similar to the twin-arginine consensus motif. The 49 amino acid-long signal peptide was analyzed by random, site-directed and site-saturation mutagenesis and the effect of these mutations on XlnC secretion determined. None of the mutations abolished XlnC production and the decreased yields were attributed to the low processing rate of precursors ranging from 2 to 5 h instead of 11 min for the wild-type precursor. Replacement of phenylalanine in the consensus motif by other amino acid residues decreased XlnC secretion by 75%, except for a tryptophan substitution which demonstrated a 50% decrease. Charge distribution in the n-domain of the signal peptide was more important than the net charge. Replacement of the signal peptidase recognition site A-H-A by either A-H-E, V-D-S or R-L-E did not affect precursor processing, indicating that the presence of the conserved residues found in the signal peptidase recognition site is not a prerequisite for the processing of Tat-substrates as it is for the processing of Sec-substrates in S. lividans.
FEMS Microbiology Letters 03/2006; 255(2):268-74. · 2.04 Impact Factor
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ABSTRACT: The bacterial twin-arginine translocation (Tat) pathway transports folded proteins across the cytoplasmic membrane. The precursors targeted to the Tat pathway have signal peptides bearing the consensus motif (S/T-R-R-X-F-L-K). The xylanase C (XlnC) of Streptomyces lividans is a 20-kDa secreted enzyme. The XlnC signal peptide is 49 amino acids long and contains the S-R-R-G-F-L-G sequence, which is similar to the twin-arginine consensus motif. In S. lividans, XlnC secretion was impaired in a tatC insertion mutant, which is unable to secrete proteins that are dependent on the Tat system. When the signal peptide of XlnC was replaced by the Sec-dependent signal peptide of xylanase A, XlnC was secreted as an inactive form and demonstrated rapid proteolytic degradation in the culture supernatant, thus indicating that XlnC was specifically secreted through the Tat system. Deletions of the n-region of the XlnC signal sequence showed that a minimum of six amino acids residues preceding the twin-arginine motif was required to secrete XlnC. Replacement of one or both arginines by lysine residues in the twin arginine motif decreased four- and sevenfold, respectively, the enzyme production but did not abolish it. However, pulse chase experiments showed that the half-life of the precursor was from 2 to 3 h instead of 11 min for the wild- type precursor. Since XlnC is not associated with cofactors to exhibit activity, it is therefore a newly identified prokaryotic non-redox Tat substrate.
Biochimica et Biophysica Acta 07/2004; 1699(1-2):155-62. · 4.66 Impact Factor
