Brian D Dynlacht
New York University School Of Medicine
Project start date: 2003-02-01
Project end date: 2016-01-31
Sponsored Links Excellgen http://Excellgen.com
Role Of The PRB Family In Quiescence And Differentiation
Brian D Dynlacht, Professor
New York University School Of Medicine New York, Ny 10016
Grant 2R01GM067132-05A1 from National Institute Of General Medical Sciences IRG: ZRG1
Abstract: A major focus in the area of human cancer pertains to the programs involved in cell cycle exit, differentiation, and maintenance of the differentiated state. Research over nearly two decades has suggested that the retinoblastoma tumor suppressor (pRB) and two related proteins (p107 and p130; collectively referred to as pocket proteins) play a fundamental role in regulating the cell cycle, and pRB has been shown to be a prototypical tumor suppressor mutated in a large portion of human tumors. In addition, pRB has been shown to play a pivotal role in differentiation of several tissues, including muscle and bone. Our work during the previous funding period has highlighted unique roles for each of the pocket proteins in responding to growth arrest cues and in both promoting and maintaining differentiation of muscle. In addition, we have succeeded for the first time in purifying pRB complexes from proliferating and differentiated muscle cells. In this grant, we propose the following aims to further dissect the mechanisms underlying pocket protein involvement in cell cycle exit and differentiation. (1) We will characterize pRB complexes in proliferating and differentiated cells and examine the impact of depleting associated proteins on gene expression and differentiation; (2) we will identify targets of the pRB complexes in cultured cells and in muscle tissue and examine the role of pRB complexes in modification of target gene chromatin; and (3) we will examine whether mechanisms analogous to those discovered in Aims 1 and 2 pertain to reversible growth arrest as well and determine if there are mechanisms that distinguish reversible and irreversible cell cycle exit. These studies will enhance our understanding of regulatory controls that are essential for both reversible and permanent withdrawal from the cell cycle and differentiation.
Project start date: 2003-02-01
Project end date: 2011-01-31
2R01GM067132-05A1 (2007): $253529
Grants awarded to Brian D Dynlacht
TRANSCRIPTIONAL REGULATION BY THE PRB PROTEIN FAMILY
Brian D Dynlacht, Professor
Molecular And Cellular Biologyharvard University
1350 Massachusetts Ave
cambridge, Ma 02138
Grant 5R01CA077245-04 from National Cancer Institute IRG: MBY
Abstract: This proposal investigates the mechanisms by which the retinoblastoma tumor suppressor protein (pRB) and two related proteins, p107 and p130, modulate transcription. The pRB family plays an important role in restraining cell proliferation, and mutations of pRB have been found in a variety of cancers. It is thought that these proteins suppress growth in part through an ability to regulate transcriptional activity of proteins to which they bind. We propose a biochemical approach toward studying the function of the pRB family of proteins. Our objectives include the following (1) determining whether the pRB family targets proteins in the basal transcription machinery; (2) understanding how pRB can inhibit and activate transcription; and (3) understanding how the pRB-related proteins, p107 and p130, and p107/p130 complexes with cyclin-dependent kinases, modulate gene expression and growth. Each of these specific aims is part of an overall attempt to understand how certain critical cell cycle regulators modulate transcriptional responses during cell proliferation. These studies will be carried out using a mammalian in vitro transcription system reconstituted with purified basal and sequence- specific transcription factors and recombinant cell cycle proteins. The well-defined nature of these in vitro assays circumvents many of the complications of studying cell cycle-regulated events in vivo. In vivo growth suppression, protein-protein interaction assays, and DNA-binding analyses with purified or partially purified proteins will be used in parallel to confirm and strengthen our in vitro transcription results. Although much research has focused on the regulatory cues that promote cell growth, our understanding of the mechanisms that drive proliferation is still rudimentary. It is clear that a thorough knowledge of the interplay between gene expression and cell cycle regulatory proteins will be of fundamental importance in understanding the mechanisms for cell cycle progression in both normal and cancer cells
Keywords: cell growth regulation, genetic transcription, retinoblastoma protein, transcription factor DNA binding protein, cell cycle, cell cycle protein, cell proliferation, cyclin dependent kinase, enzyme activity, gene induction /repression, genetic promoter element, neoplastic growth, phosphorylation, protein binding, protein protein interaction, recombinant protein cell line, gel mobility shift assay, transfection, western blotting
Project start date: 1998-06-19
Project end date: 2004-02-29
5R01CA077245-04 (2001): $192888
5R01CA077245-03 (2000): $187352
5R01CA077245-02 (1999): $162817
1R01CA077245-01 (1998): $158158
7R01CA077245-06 (2003): $195225
Role Of The PRB Family In Quiescence And Differentiation
Brian D Dynlacht, Professor
New York University School Of Medicine New York, Ny 10016
Grant 5R01GM067132-04 from National Institute Of General Medical Sciences IRG: CDF
Abstract: The retinoblastoma tumor suppressor protein (pRB) and the related proteins p107 and p130 (collectively termed "pocket" proteins) play an established role in suppressing cell growth through inhibition of the E2F transcription factor. A role for the pRB family in cell cycle exit and muscle differentiation has also been documented. While cellular quiescence and p16INK4a-induced growth arrest appear to require combinations of "pocket" proteins, specific pRB family members have been implicated in terminal differentiation of muscle cells. However, very few direct, physiological targets have been linked to cellular quiescence, and fewer direct targets associated with differentiation have been identified. Furthermore, pRB binding to promoters has not been widely observed in cultured fibroblasts during the cell cycle, raising interesting and important questions regarding the role of pRB in tumor suppression and suggesting that pRB s tumor suppressive function may involve a much more extensive role in promoting differentiation than previously imagined. One goal of this proposal is to identify and characterize (1) direct, physiological targets of the pRB family involved in achieving cellular quiescence and p161NK4a -mediated growth arrest and (2) those gene targets that cooperate to confer irreversible cell cycle exit and terminal differentiation of muscle. It will also attempt to distinguish between those controls involved in cell cycle withdrawal and phenotypic differentiation. This will be accomplished through large-scale analyses of "pocket" protein binding to the genome of living cells (factor location analysis) during the process of cell cycle exit and differentiation, through simultaneous analysis of gene expression profiles, and through biochemical dissection of target promoters. By examining three cell cycle exit pathways that appear to require certain pRB family members but not others, this work will have a fundamental impact on our understanding of the existence of gene regulatory networks effecting cell cycle exit in response to distinct biological cues. pRB plays a well-documented role in growth control, and inactivation of this tumor suppressor has been associated with a large proportion of human cancers. This Proposal is therefore highly relevant to our understanding of tumor suppressive mechanisms and cancer.
Keywords: cell cycle, cell differentiation, gene expression, genetic regulation, muscle cell, protein structure function, retinoblastoma protein, acetylation, amidohydrolase, binding site, chromatin, developmental genetics, fibroblast, genetic promoter element, genome, protein binding, striated muscle, transcription factor, tumor suppressor protein, cell line, immunoprecipitation, microarray technology
Project start date: 2003-02-01
Project end date: 2007-09-19
5R01GM067132-04 (2006): $354812
5R01GM067132-03 (2005): $363350
5R01GM067132-02 (2004): $363350
1R01GM067132-01 (2003): $362723
Brian D Dynlacht, Professor
New York University School Of Medicine, New York, Ny 10016
Abstract: Successful unraveling of the nature and functional dynamics of the human genome together with significant advances in computational biology, scientific research of genome-related phenomena have accelerated our understanding of pathogenesis of cancer.and other human diseases. The focus is at fundamental molecular pathomechanisms such as chromosomal abnormalities, genetic susceptibility, and genome structure as well as the epigenetic, transcriptional and post-transcriptional regulation of gene function. In order to realize the goals of genome-oriented research and to make relevant methodologies readily available to individual NYU Cancer Institute researchers as they embark on their research career, it is imperative to develop institutional core facilities that will support such an effort. The NYU Cancer Institute (NYU Cl) under the leadership of Dr. Steven Burakoff has made a significant effort to respond to the challenges of modern functional genomic science by establishing a state-of-the-art Genomics Facility and most recently the RNAi Core Facility. These two core laboratories work in close co-operation to provide NYU Cl investigators with services and expertise in microarray- or quantitative real-time PCR-based profiling of gene and microRNA expression, of protein- DNA interactions in chromatin structure studies, in DMA analysis such as SNP analysis for genotyping, copy number studies, and DNA sequencing for SNP, mutation or CpG-island methylation studies, and finally with high-throughput RNA interference screens
Keywords: Activities, Educational; Analysis, Data; Animal Model; Animal Models and Related Studies; Arts; Assay; Bio-Informatics; Bioassay; Bioinformatics; Biologic Assays; Biological Assay; Biology; C elegans; C. elegans genome; C.elegans; CCSG; Caenorhabditis elegans; Cancer Biology; Cancer Center Support Grant; Cancers; Cells; Chemistry; Chromatin Structure; Chromosomes; Communities; Comparative Genome Hybridization; Computing Methodologies; Consultations; Core Facility; Core Grant; Country; DNA; DNA Chips; DNA Microarray; DNA Microarray Chip; DNA Microchips; Data Analyses; Deoxyribonucleic Acid; Development; Double-Stranded RNA; Drosophila; Drosophila genome; Drosophila genus; Drugs; Educational Activities; Environment; Equipment; Experimental Designs; Expression Profiling; Expression Signature; Fostering; Fruit Fly, Drosophila; Gene Expression; Genes; Genome; Genome, Human; Genomics; Goals; Guidelines; Housing; Human; Human Genome; Human, General; Institutes; Investigators; Laboratories; Lead; Leadership; Libraries; Logistics; Malignant Neoplasms; Malignant Tumor; Mammals, Mice; Man (Taxonomy); Man, Modern; Medication; Method LOINC Axis 6; Methodology; Methods and Techniques; Methods, Other; Mice; Mission; Molecular Fingerprinting; Molecular Profiling; Murine; Mus; Nucleic Acids; P30 Grant; Pb element; Pharmaceutic Preparations; Pharmaceutical Preparations; Policies; Post-Transcriptional Gene Silencing; Post-Transcriptional Gene Silencings; Posttranscriptional Gene Silencing; Posttranscriptional Gene Silencings; Process; Publications; Quelling; RNA Interference; RNA Silencing; RNA Silencings; RNA, Double-Stranded; RNA, Small Interfering; RNAi; Research; Research Personnel; Researchers; Resolution; SNP genotyping; Science; Science of Chemistry; Scientific Publication; Screening procedure; Sequence-Specific Posttranscriptional Gene Silencing; Services; Small Interfering RNA; Techniques; Validation; Work; base; chromatin protein; comparative genomic hybridization; computational methodology; computational methods; computer methods; data acquisition; drug/agent; dsRNA; experiment; experimental research; experimental study; falls; feeding; fruit fly; functional genomics; genome, C elegans; genome, C. elegans; genome, C.elegans; genome, Caenorhabditis elegans; genome, Drosophila; genome, fruit fly; genome-wide; heavy metal Pb; heavy metal lead; malignancy; member; model organism; molecuar profile; molecular signature; neoplasm/cancer; research study; screening; screenings; shRNA; sharing data; short hairpin RNA; siRNA; small hairpin RNA
Budget start date: 1-JUN-2009
Budget end date: 30-SEP-2010
3P30CA016087-29S1_9032 (2009): $5531
Sponsored Links Excellgen http://Excellgen.com
ROLE OF THE PRB FAMILY IN QUIESCENCE AND DIFFERENTIATION
Brian D Dynlacht, Professor
New York University School Of Medicine, 550 1st Ave, New York, Ny 10016
Grant 3R01GM067132-07S1 from National Institute Of General Medical Sciences
Abstract: Research during the past two decades has suggested that the retinoblastoma tumor suppressor (pRB) and two related proteins (p107 and p130; collectively referred to as pocket proteins) play a fundamental role in regulating the cell cycle, and pRB has been shown to be a prototypical tumor suppressor mutated in a large portion of human tumors. In addition, pRB has been shown to play a pivotal role in differentiation of several tissues, including muscle and bone. Our work during the previous funding period, under the auspices of our parent grant application, has highlighted unique roles for each of the pocket proteins in responding to growth arrest cues and in both promoting and maintaining differentiation of muscle. In addition, we have succeeded for the first time in purifying pRB complexes from proliferating and differentiated muscle cells. In this grant, we propose the following aims to further dissect the mechanisms underlying pocket protein involvement in cell cycle exit and differentiation. (1) We will characterize pRB complexes in proliferating and differentiated cells and examine the impact of depleting associated proteins on gene expression and differentiation; (2) we will identify targets of the pRB complexes in cultured cells and in muscle tissue and examine the role of pRB complexes in modification of target gene chromatin; and (3) we will examine whether mechanisms analogous to those discovered in Aims 1 and 2 pertain to reversible growth arrest as well and determine if there are mechanisms that distinguish reversible and irreversible cell cycle exit. In the current Revision to the original parent application, we propose broadening our investigation of chromatin modifications associated with myogenic differentiation, focusing in particular on those changes that are dependent on the pRB family of proteins. We will incorporate the new, state-of-the-art ChIP-sequencing approach to examine these modifications in an unbiased, genome-wide manner. These studies will enhance our understanding of regulatory controls that are essential for both reversible and permanent withdrawal from the cell cycle and differentiation. In addition, they will elucidate critical interactions between repressors, co-repressors, and chromatin modifications as they occur in a developmentally relevant setting. Cancer results in some cases when cells fail to properly differentiate. Differentiation is coupled to exit from the cell cycle, and the retinoblastoma (pRB) tumor suppressor plays a pivotal role in controlling growth arrest and differentiation. This proposal seeks to understand the underlying mechanisms whereby pRB regulates gene expression and controls the decision to permanently stop dividing and to terminally differentiate
Keywords: Algorithms; Arts; Body Tissues; Bone; Bone and Bones; Bones and Bone Tissue; CHIP assay; Cancers; Cell Cycle; Cell Division Cycle; Cells; ChIP (chromatin immunoprecipitation); Chromatin; Complex; Coupled; Cues; Cultured Cells; Family; Family member; Funding; Gene Expression; Gene Targeting; Generalized Growth; Genome; Grant; Growth; Human; Human, General; Investigation; Link; Location; Malignant Neoplasms; Malignant Tumor; Man (Taxonomy); Man, Modern; Modification; Muscle; Muscle Cells; Muscle Cells, Embryonic; Muscle Cells, Mature; Muscle Cells, Precursor; Muscle Fibers; Muscle Tissue; Mutagenesis, Site-Directed; Mutate; Myoblasts; Myocytes; Myotubes; Neuroblastoma of the Retina; Neuroblastoma, Retinal; P105-RB; P130; PP110; Parents; Play; Post-Transcriptional Gene Silencing; Post-Transcriptional Gene Silencings; Posttranscriptional Gene Silencing; Posttranscriptional Gene Silencings; Proliferating; Protein Family; Proteins; Quelling; RB1; RBL1 protein; RBL2; RBL2 gene; RNA Interference; RNA Silencing; RNA Silencings; RNAi; Rb Gene Product; Rb Protein; Rb1 Gene Product; Rb2; Research; Retinoblastoma; Retinoblastoma Associated Protein; Retinoblastoma Protein; Rhabdomyocyte; Role; Sequence-Specific Posttranscriptional Gene Silencing; Site-Directed Mutagenesis; Site-Specific Mutagenesis; Skeletal Fiber; Skeletal Muscle Cell; Skeletal Muscle Fiber; Skeletal Myocytes; Targeted DNA Modification; Targeted Modification; Targetings, Gene; Time; Tissue Growth; Tissues; Tumor Suppressor Proteins; Withdrawal; Work; bone; chromatin immunoprecipitation; chromatin modification; experiment; experimental research; experimental study; gene product; genome-wide; malignancy; neoplasm/cancer; ontogeny; p107 protein; p107 tumor suppressor protein; pRB; parent grant; public health relevance; research study; retinoblastoma like protein p107; retinoblastoma tumor suppressor; retinoblastoma-like 1 protein; social role; tumor; tumor suppressor
Relevance: Relevance Cancer results in some cases when cells fail to properly differentiate. Differentiation is coupled to exit from the cell cycle, and the retinoblastoma (pRB) tumor suppressor plays a pivotal role in controlling growth arrest and differentiation. This proposal seeks to understand the underlying mechanisms whereby pRB regulates gene expression and controls the decision to permanently stop dividing and to terminally differentiate
Project start date: 2009-09-30
Project end date: 2011-01-31
Budget start date: 30-SEP-2009
Budget end date: 31-JAN-2011
PFA/PA: PA-07-070
3R01GM067132-07S1 (2009): $219700
THE ROLE OF PRB AND CO-REPRESSORS IN TRANSCRIPTIONAL REGULATION
Brian D Dynlacht, Professor
New York University School Of Medicine, New York, Ny 10016
Grant 5R01CA077245-13 from National Cancer Institute
Abstract: Cellular differentiation is tightly coordinated with, and dependent upon, permanent cell cycle exit. The tumor suppressor pRB prevents tumorigenesis by virtue of its ability to suppress proliferation. It is clear that the pRB tumor family, which is comprised of pRB and the related proteins, p107 and p130 (also known as pocket proteins), is recruited to promoters through the E2F transcription factor. Once recruited, pocket proteins coordinate changes in gene expression with cell cycle progression and cell cycle exit, during transient growth arrest (quiescence) and permanent growth arrest that occurs upon cellular differentiation. Pocket proteins play a role in recruiting a cadre of co-repressors, such as Sin3-HDAC and histone methyltransferases, that modify chromatin and silence gene expression. Yet the mechanisms underlying the pRB-dependent changes in gene expression are not well understood. It is important to unravel these mechanisms, since they are likely to shed light on transcriptional controls associated with normal cell growth and tumor suppressive mechanisms. It is also unclear what transcriptional controls distinguish cells progressing through the M/G1 phase transition from those observed in cells re-emerging from growth arrest (the G0/G1 phase transition) and how differences in factor recruitment and chromatin modifications dictate transient growth arrest versus permanent cell cycle exit. We have found that pRB and the Sin3 co-repressor play a role in both settings. In this proposal, we seek to build upon results of the previous funding period and explore answers to these fundamental issues in the following Aims (1) We will examine the role of E2F, pocket proteins, and Sin3 isoforms in factor recruitment, chromatin modification, and nucleosome remodeling during permanent cell cycle exit in muscle cells. (2) We will determine whether Sin3A and Sin3B isoforms can specifically regulate target gene transcription and purify endogenous complexes. (3) We will investigate the phenotype of Sin3 conditional knock-outs and attempt to unravel a role for Sin3 isoforms in maintaining cell cycle arrest in an animal model. Cells that fail to exit the cell cycle and differentiate can over-proliferate, promoting tumorigenesis. Loss of the pRB tumor suppressor can lead to de-repression of cell cycle genes and abnormal proliferation, expanding the population of cells that can sustain a second genetic mutation. Permanent cell cycle exit and differentiation are tightly coupled. It is known that the retinoblastoma (pRB) tumor suppressor family plays a pivotal role in controlling growth arrest and differentiation, in part through its association with co-repressors such as Sin3, and its ability to epigenetically modify chromatin. This proposal seeks to understand the underlying chromatin modifications and remodeling mechanisms whereby the pRB family and the Sin3 co-repressor regulate gene expression during cell cycle progression and growth arrest
Keywords: Ablation; Animal Model; Animal Models and Related Studies; Apoptosis; Apoptosis Pathway; Binding; Binding (Molecular Function); Biochemical; Cell Cycle; Cell Cycle Arrest; Cell Cycle Genes; Cell Cycle Progression; Cell Death, Programmed; Cell Division Cycle; Cells; Cellular Expansion; Cellular Growth; Chromatin; Chromatin Remodeling Complex; Chromatin Remodeling Factor; Complex; Coupled; DNA; DNA Alteration; DNA mutation; Deoxyribonucleic Acid; E2F transcription factors; Enzymes; Exhibits; Family; First Gap Phase; Funding; G1 Phase; G1 period; Gap Phase 1; Gene Action Regulation; Gene Alteration; Gene Down-Regulation; Gene Expression; Gene Expression Regulation; Gene Mutation; Gene Regulation; Gene Regulation Process; Gene Targeting; Gene Transcription; Generalized Growth; Genes; Genes, Cell Division Cycle; Genes, cdc; Genetic Transcription; Genetic mutation; Growth; HDAC; HDAC Proteins; Histone Deacetylase; In Vitro; Individual; Isoforms; Knockout Mice; Lead; Light; Maintenance; Maintenances; Mammalian Cell; Mice, Knock-out; Mice, Knockout; Molecular Interaction; Muscle; Muscle Cells; Muscle Cells, Mature; Muscle Fibers; Muscle Tissue; Myocytes; Myotubes; Neuroblastoma of the Retina; Neuroblastoma, Retinal; Normal Cell; Nucleosomes; Null Mouse; Oncogenesis; P105-RB; P130; PP110; Pb element; Phase Transition; Phenotype; Photoradiation; Physiologic; Physiological; Play; Population; Position; Positioning Attribute; Post-Transcriptional Gene Silencing; Post-Transcriptional Gene Silencings; Posttranscriptional Gene Silencing; Posttranscriptional Gene Silencings; Proliferating; Promoter; Promoters (Genetics); Promotor; Promotor (Genetics); Protein Isoforms; Proteins; Quelling; RB1; RBL1 protein; RBL2; RBL2 gene; RNA Expression; RNA Interference; RNA Silencing; RNA Silencings; RNAi; Rb Gene Product; Rb Protein; Rb1 Gene Product; Rb2; Recruitment Activity; Regulation; Repression; Repressor Proteins; Retinoblastoma; Retinoblastoma Associated Protein; Retinoblastoma Protein; Rhabdomyocyte; Role; Sequence Alteration; Sequence-Specific Posttranscriptional Gene Silencing; Skeletal Fiber; Skeletal Muscle Cell; Skeletal Muscle Fiber; Skeletal Myocytes; Specificity; Targetings, Gene; Tissue Growth; Transcription; Transcription Regulation; Transcription Repression; Transcription, Genetic; Transcriptional Control; Transcriptional Regulation; Transcriptional Repression; Tumor Suppressor Proteins; base; cdc Genes; cell growth; chromatin modification; gene product; gene repression; genome-wide; heavy metal Pb; heavy metal lead; histone H3 methyltransferase; histone methylase; histone methyltransferase; histone modification; in vivo; model organism; ontogeny; p107 protein; p107 tumor suppressor protein; pRB; prevent; preventing; public health relevance; recruit; retinoblastoma like protein p107; retinoblastoma-like 1 protein; social role; tumor; tumor suppressor; tumorigenesis
Relevance: Relevance Cells that fail to exit the cell cycle and differentiate can over-proliferate, promoting tumorigenesis. Loss of the pRB tumor suppressor can lead to de-repression of cell cycle genes and abnormal proliferation, expanding the population of cells that can sustain a second genetic mutation. Permanent cell cycle exit and differentiation are tightly coupled. It is known that the retinoblastoma (pRB) tumor suppressor family plays a pivotal role in controlling growth arrest and differentiation, in part through its association with co-repressors such as Sin3, and its ability to epigenetically modify chromatin. This proposal seeks to understand the underlying chromatin modifications and remodeling mechanisms whereby the pRB family and the Sin3 co-repressor regulate gene expression during cell cycle progression and growth arrest
Project start date: 1998-06-19
Project end date: 2014-05-31
Budget start date: 1-JUN-2010
Budget end date: 31-MAY-2011
PFA/PA: PA-07-070
5R01CA077245-13 (2010): $348346
2R01CA077245-12A1 (2009): $314521
THE ROLE OF UBIQUITINATION IN CELL CYCLE EXIT AND CACHEXIA
Brian D Dynlacht, Professor
New York University School Of Medicine, New York, Ny 10016
Grant 5R21CA125734-02 from National Cancer Institute
Abstract: Cachexia, or severe tissue wasting, is an extremely debilitating consequence of cancer that results in nearly thirty percent of cancer deaths. The mechanisms underlying cancer cachexia remain poorly defined, but recent experiments have shed important new light on the role of critical mediators, which appear to alter the balance between protein synthesis and degradation in skeletal muscle. These experiments have demonstrated that genes involved in ubiquitin-mediated proteasomal degradation are dramatically induced in several in vitro models of skeletal muscle atrophy, in rodent models, and in patients. We will use a combination of genomic and proteomic approaches to identify both the key upstream regulators and downstream effectors of skeletal muscle atrophy, in particular in the context of cancer cachexia. This proposal will advance two important goals (1) using genomics and a novel proteomic screen to identify key ubiquitin ligases and the substrates of these enzymes that are activated in both in vitro and in vivo murine models of cancer cachexia, simultaneously exploring the utility of in vitro models for identifying key pathways regulated during cancer cachexia, and (2) initiating an outline for key transcriptional regulators that drive expression of ubiquitin ligases during normal muscle differentiation and muscle wasting. If successful, our proposal will generate detailed information regarding mechanisms whereby ubiquitylation mediates the devastating effects of cachexia and suggest potential upstream regulators and downstream targets for therapeutic intervention. Lay Muscle wasting is an extremely debilitating result of cancer and results in the discomfort, and in many instances, death of the patient. Yet the mechanisms through which skeletal muscle wasting, or cachexia, occur are largely unknown. It is clear that protein modification (through attachment of a small molecule, ubiquitin) and degradation play a major role in this process, and this proposal aims to understand some of the important steps in muscle protein degradation that result in cancer cachexia
Keywords: APF-1; ATP-Dependent Proteolysis Factor 1; Address; Atrophic; Atrophy; Atrophy, Muscle; Binding; Binding (Molecular Function); Biochemical; Body Tissues; Cachexia; Cancer Model; CancerModel; Cancers; Cell Cycle; Cell Division Cycle; Cells; Cessation of life; Classification; Condition; Cues; Death; Disease; Disorder; Equilibrium; Expression Profiling; Expression Signature; Genes; Genomics; Goals; HMG-20; HTRPY; High Mobility Protein 20; Hypertrophy; In Vitro; Ligase; Light; Malignant Neoplasms; Malignant Tumor; Mammals, Mice; Mediating; Mediator; Mediator of Activation; Mediator of activation protein; Mice; Modeling; Modification; Molecular Fingerprinting; Molecular Interaction; Molecular Profiling; Murine; Mus; Muscle; Muscle Fibers; Muscle Proteins; Muscle Tissue; Muscle, Skeletal; Muscle, Voluntary; Muscular Atrophy; Myotubes; Numbers; Pathway interactions; Patients; Peptide Biosynthesis, Ribosomal; Photoradiation; Play; Post-Translational Modifications; Post-Translational Protein Processing; Posttranslational Modifications; Process; Programs (PT); Programs [Publication Type]; Protein Biosynthesis; Protein Biosynthesis, Ribosomal; Protein Degradation, Metabolic; Protein Degradation, Regulatory; Protein Modification; Protein Modification, Post-Translational; Protein Processing, Post-Translational; Protein Processing, Posttranslational; Protein Synthesis, Ribosomal; Protein Turnover; Protein/Amino Acid Biochemistry, Post-Translational Modification; Proteins; Proteomics; Rhabdomyocyte; Rodent Model; Role; Skeletal Fiber; Skeletal Muscle Cell; Skeletal Muscle Fiber; Skeletal Muscle Tissue; Skeletal Myocytes; Skeletal muscle structure; Skeletal system; Synthetases; Systematics; Testing; Therapeutic Intervention; Time; Tissues; Transcription Regulatory Protein; Transcriptional Regulatory Protein; Ubiquitilation; Ubiquitin; Ubiquitination; Ubiquitinoylation; ing; balance; balance function; disease/disorder; enzyme substrate; experiment; experimental research; experimental study; gene product; in vitro Model; in vivo; intervention therapy; malignancy; molecuar profile; molecular signature; neoplasm/cancer; novel; pathway; programs; protein degradation; protein synthesis; research study; response; skeletal; small molecule; social role; ubiquination; ubiquitin conjugation; ubiquitin ligase; wasting
Project start date: 2007-09-01
Project end date: 2010-08-31
Budget start date: 1-SEP-2008
Budget end date: 31-AUG-2010
PFA/PA: PA-03-145
5R21CA125734-02 (2008): $0
1R21CA125734-01 (2007): $236600
Role Of The PRB Family In Quiescence And Differentiation
Brian D Dynlacht, Professor
Pathologynew York University School Of Medicine
Grant 5R01GM067132-07 from National Institute Of General Medical Sciences IRG: ZRG1
Abstract: A major focus in the area of human cancer pertains to the programs involved in cell cycle exit, differentiation, and maintenance of the differentiated state. Research over nearly two decades has suggested that the retinoblastoma tumor suppressor (pRB) and two related proteins (p107 and p130; collectively referred to as pocket proteins) play a fundamental role in regulating the cell cycle, and pRB has been shown to be a prototypical tumor suppressor mutated in a large portion of human tumors. In addition, pRB has been shown to play a pivotal role in differentiation of several tissues, including muscle and bone. Our work during the previous funding period has highlighted unique roles for each of the pocket proteins in responding to growth arrest cues and in both promoting and maintaining differentiation of muscle. In addition, we have succeeded for the first time in purifying pRB complexes from proliferating and differentiated muscle cells. In this grant, we propose the following aims to further dissect the mechanisms underlying pocket protein involvement in cell cycle exit and differentiation. (1) We will characterize pRB complexes in proliferating and differentiated cells and examine the impact of depleting associated proteins on gene expression and differentiation; (2) we will identify targets of the pRB complexes in cultured cells and in muscle tissue and examine the role of pRB complexes in modification of target gene chromatin; and (3) we will examine whether mechanisms analogous to those discovered in Aims 1 and 2 pertain to reversible growth arrest as well and determine if there are mechanisms that distinguish reversible and irreversible cell cycle exit. These studies will enhance our understanding of regulatory controls that are essential for both reversible and permanent withdrawal from the cell cycle and differentiation
Project start date: 2003-02-01
Project end date: 2011-01-31
Transcriptional Regulation By The PRB And E2F Families
Brian D Dynlacht, Professor
Pathologynew York University School Of Medicine
550 1st Ave
new York, Ny 10016
Grant 5R01CA077245-11 from National Cancer Institute IRG: CDF
Abstract: A comprehensive understanding of growth control in normal mammalian cells requires that we know how gene regulatory circuits operate as a function of the cell cycle. The transcription factor, E2F, and pRB, a tumor suppressor known to restrain E2F activity, are among the best-studied regulators of cell cycle dependent gene expression. Pathways that involve pRB are known to be de-regulated in a majority of human cancers. Despite the fact that E2F and pRB have been linked genetically to growth control, the mechanisms by which these factors modulate endogenous gene expression and globally regulate cell cycle progression are not well understood. A complete of their function requires that we understand not only physiological changes that occur at a given cell cycle regulated promoter but also the intricate biochemical mechanisms that switch a gene from the ´OFF´ to ´ON´ state in a reversible manner during the cell cycle. A detailed can be achieved only by studying the regulation of E2F targets in a natural (chromatin) setting under conditions where cells are either stimulated to grow or are continuously cycling. Our hypothesis suggests that discrete mechanisms must exist that distinguish gene expression profiles during each stage of the cell cycle. Given the pivotal role of pRB in tumor suppression, an understanding of these mechanisms will provide crucial clues necessary for understanding the regulation of normal cell growth and the ways in which controls may go awry in human cancer. This Research Proposal takes advantage of multiple, complementary approaches to test this idea. In Aim 1, a combination of genomics and biochemistry will be used to identify targets of E2F and pRB families and chromatin modifying enzymes in proliferating cells at each stage of the cell cycle to determine those factors that regulate genes in both active and inactive states. Using rapidly emerging bioinformatics and computational tools, approaches in Aim 2 will attempt to identify those patterns that link genes co-regulated by E2F, pRB, and chromatin modifying enzymes. Biochemical experiments in Aim 3 will test predictions about these patterns and will reveal those mechanisms that distinguish expression profiles of native E2F target genes at different stages of the cell cycle
Keywords: cell cycle, cell growth regulation, gene expression, genetic transcription, neoplasm /cancer genetics, retinoblastoma protein, transcription factor acyltransferase, cell proliferation, chromatin, enzyme activity, gene induction /repression, genetic promoter element, genetic regulation, histone, neoplastic growth, protein binding, tumor suppressor protein bioinformatics, cell line, chromatin immunoprecipitation, gene expression profiling, genetic mapping, microarray technology
Project start date: 1998-06-19
Project end date: 2009-06-30
5R01CA077245-11 (2008): $304460
5R01CA077245-10 (2007): $304460
5R01CA077245-09 (2006): $313554
5R01CA077245-08 (2005): $321100
Sponsored Links Excellgen http://Excellgen.com
2R01CA077245-07 (2004): $308680
Role Of The PRB Family In Quiescence And Differentiation
Brian D Dynlacht, Professor
Pathologynew York University School Of Medicine, 550 1st Ave, New York, Ny 10016
Grant 3R01GM067132-04S1 from National Institute Of General Medical Sciences IRG: CDF
Project start date: 2003-02-01
Project end date: 2008-01-31
3R01GM067132-04S1 (2007): $118271