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1.
FASEB J ; 38(11): e23702, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38837439

RESUMO

Pyruvate kinase is a glycolytic enzyme that converts phosphoenolpyruvate and ADP into pyruvate and ATP. There are two genes that encode pyruvate kinase in vertebrates; Pkm and Pkl encode muscle- and liver/erythrocyte-specific forms, respectively. Each gene encodes two isoenzymes due to alternative splicing. Both muscle-specific enzymes, PKM1 and PKM2, function in glycolysis, but PKM2 also has been implicated in gene regulation due to its ability to phosphorylate histone 3 threonine 11 (H3T11) in cancer cells. Here, we examined the roles of PKM1 and PKM2 during myoblast differentiation. RNA-seq analysis revealed that PKM2 promotes the expression of Dpf2/Baf45d and Baf250a/Arid1A. DPF2 and BAF250a are subunits that identify a specific sub-family of the mammalian SWI/SNF (mSWI/SNF) of chromatin remodeling enzymes that is required for the activation of myogenic gene expression during differentiation. PKM2 also mediated the incorporation of DPF2 and BAF250a into the regulatory sequences controlling myogenic gene expression. PKM1 did not affect expression but was required for nuclear localization of DPF2. Additionally, PKM2 was required not only for the incorporation of phosphorylated H3T11 in myogenic promoters but also for the incorporation of phosphorylated H3T6 and H3T45 at myogenic promoters via regulation of AKT and protein kinase C isoforms that phosphorylate those amino acids. Our results identify multiple unique roles for PKM2 and a novel function for PKM1 in gene expression and chromatin regulation during myoblast differentiation.


Assuntos
Diferenciação Celular , Histonas , Mioblastos , Piruvato Quinase , Animais , Piruvato Quinase/metabolismo , Piruvato Quinase/genética , Camundongos , Fosforilação , Histonas/metabolismo , Histonas/genética , Mioblastos/metabolismo , Mioblastos/citologia , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Proteínas de Ligação a Hormônio da Tireoide , Humanos , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Cromossômicas não Histona/genética , Hormônios Tireóideos/metabolismo , Hormônios Tireóideos/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Ligação a DNA/genética , Isoenzimas/metabolismo , Isoenzimas/genética
2.
bioRxiv ; 2024 Apr 11.
Artigo em Inglês | MEDLINE | ID: mdl-38645038

RESUMO

Pyruvate kinase is a glycolytic enzyme that converts phosphoenolpyruvate and ADP into pyruvate and ATP. There are two genes that encode pyruvate kinase in vertebrates; Pkm and Pkl encode muscle- and liver/erythrocyte-specific forms, respectively. Each gene encodes two isoenzymes due to alternative splicing. Both muscle-specific enzymes, Pkm1 and Pkm2, function in glycolysis, but Pkm2 also has been implicated in gene regulation due to its ability to phosphorylate histone 3 threonine 11 (H3T11) in cancer cells. Here, we examined the roles of Pkm1 and Pkm2 during myoblast differentiation. RNA-seq analysis revealed that Pkm2 promotes the expression of Dpf2/Baf45d and Baf250a/Arid1A. Dpf2 and Baf250a are subunits that identify a specific sub-family of the mammalian SWI/SNF (mSWI/SNF) of chromatin remodeling enzymes that is required for activation of myogenic gene expression during differentiation. Pkm2 also mediated the incorporation of Dpf2 and Baf250a into the regulatory sequences controlling myogenic gene expression. Pkm1 did not affect expression but was required for nuclear localization of Dpf2. Additionally, Pkm2 was required not only for the incorporation of phosphorylated H3T11 in myogenic promoters, but also for the incorporation of phosphorylated H3T6 and H3T45 at myogenic promoters via regulation of AKT and protein kinase C isoforms that phosphorylate those amino acids. Our results identify multiple unique roles for Pkm2 and a novel function for Pkm1 in gene expression and chromatin regulation during myoblast differentiation.

3.
Front Cell Dev Biol ; 11: 1160227, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37484913

RESUMO

Skeletal muscle differentiation is a tightly regulated process, and the importance of the mammalian SWI/SNF (mSWI/SNF) chromatin remodeling family for regulation of genes involved in skeletal myogenesis is well-established. Our prior work showed that bromodomains of mSWI/SNF ATPases BRG1 and BRM contribute to myogenesis by facilitating the binding of mSWI/SNF enzymes to regulatory regions of myogenic and other target genes. Here, we report that pathway analyses of differentially expressed genes from that study identified an additional role for mSWI/SNF enzymes via the regulation of the Wnt signaling pathway. The Wnt pathway has been previously shown to be important for skeletal muscle development. To investigate the importance of mSWI/SNF enzymes for the regulation of the Wnt pathway, individual and dual knockdowns were performed for BRG1 and BRM followed by RNA-sequencing. The results show that BRG1, but not BRM, is a regulator of Wnt pathway components and downstream genes. Reactivation of Wnt pathway by stabilization of ß-catenin could rescue the defect in myogenic gene expression and differentiation due to BRG1 knockdown or bromodomain inhibition using a specific small molecule inhibitor, PFI-3. These results demonstrate that BRG1 is required upstream of ß-catenin function. Chromatin immunoprecipitation of BRG1, BRM and ß-catenin at promoters of Wnt pathway component genes showed binding of BRG1 and ß-catenin, which provides further mechanistic insight to the transcriptional regulation of these genes.

4.
Int J Mol Sci ; 24(14)2023 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-37511016

RESUMO

Mammalian SWI/SNF (mSWI/SNF) complexes are ATP-dependent chromatin remodeling enzymes that are critical for normal cellular functions. mSWI/SNF enzymes are classified into three sub-families based on the presence of specific subunit proteins. The sub-families are Brm- or Brg1-associated factor (BAF), ncBAF (non-canonical BAF), and polybromo-associated BAF (PBAF). The biological roles for the different enzyme sub-families are poorly described. We knocked down the expression of genes encoding unique subunit proteins for each sub-family, Baf250A, Brd9, and Baf180, which mark the BAF, ncBAF, and PBAF sub-families, respectively, and examined the requirement for each in myoblast differentiation. We found that Baf250A and the BAF complex were required to drive lineage-specific gene expression. KD of Brd9 delayed differentiation. However, while the Baf250A-dependent gene expression profile included myogenic genes, the Brd9-dependent gene expression profile did not, suggesting Brd9 and the ncBAF complex indirectly contributed to differentiation. Baf180 was dispensable for myoblast differentiation. The results distinguish between the roles of the mSWI/SNF enzyme sub-families during myoblast differentiation.


Assuntos
Cromatina , Proteínas Cromossômicas não Histona , Humanos , Animais , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Montagem e Desmontagem da Cromatina/genética , Mioblastos/metabolismo , Mamíferos/metabolismo
5.
bioRxiv ; 2023 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-37398486

RESUMO

Protein arginine methyltransferase 5 (Prmt5) is an essential regulator of embryonic development and adult progenitor cell functions. Prmt5 expression is mis-regulated in many cancers, and the development of Prmt5 inhibitors as cancer therapeutics is an active area of research. Prmt5 functions via effects on gene expression, splicing, DNA repair, and other critical cellular processes. We examined whether Prmt5 functions broadly as a genome-wide regulator of gene transcription and higher-order chromatin interactions during the initial stages of adipogenesis using ChIP-Seq, RNA-seq, and Hi-C using 3T3-L1 cells, a frequently utilized model for adipogenesis. We observed robust genome-wide Prmt5 chromatin-binding at the onset of differentiation. Prmt5 localized to transcriptionally active genomic regions, acting as both a positive and a negative regulator. A subset of Prmt5 binding sites co-localized with mediators of chromatin organization at chromatin loop anchors. Prmt5 knockdown decreased insulation strength at the boundaries of topologically associating domains (TADs) adjacent to sites with Prmt5 and CTCF co-localization. Genes overlapping such weakened TAD boundaries showed transcriptional dysregulation. This study identifies Prmt5 as a broad regulator of gene expression, including regulation of early adipogenic factors, and reveals an unappreciated requirement for Prmt5 in maintaining strong insulation at TAD boundaries and overall chromatin organization.

6.
EMBO Rep ; 24(8): e57306, 2023 08 03.
Artigo em Inglês | MEDLINE | ID: mdl-37334900

RESUMO

Skeletal muscle plays a key role in systemic energy homeostasis besides its contractile function, but what links these functions is poorly defined. Protein Arginine Methyl Transferase 5 (PRMT5) is a well-known oncoprotein but also expressed in healthy tissues with unclear physiological functions. As adult muscles express high levels of Prmt5, we generated skeletal muscle-specific Prmt5 knockout (Prmt5MKO ) mice. We observe reduced muscle mass, oxidative capacity, force production, and exercise performance in Prmt5MKO mice. The motor deficiency is associated with scarce lipid droplets in myofibers due to defects in lipid biosynthesis and accelerated degradation. Specifically, PRMT5 deletion reduces dimethylation and stability of Sterol Regulatory Element-Binding Transcription Factor 1a (SREBP1a), a master regulator of de novo lipogenesis. Moreover, Prmt5MKO impairs the repressive H4R3 symmetric dimethylation at the Pnpla2 promoter, elevating the level of its encoded protein ATGL, the rate-limiting enzyme catalyzing lipolysis. Accordingly, skeletal muscle-specific double knockout of Pnpla2 and Prmt5 normalizes muscle mass and function. Together, our findings delineate a physiological function of PRMT5 in linking lipid metabolism to contractile function of myofibers.


Assuntos
Proteína-Arginina N-Metiltransferases , Transferases , Animais , Camundongos , Arginina/metabolismo , Metabolismo dos Lipídeos/genética , Músculo Esquelético/metabolismo , Proteína-Arginina N-Metiltransferases/genética , Proteína-Arginina N-Metiltransferases/metabolismo , Transferases/metabolismo
7.
Results Probl Cell Differ ; 70: 375-396, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36348115

RESUMO

The cell cycle is governed by stringent epigenetic mechanisms that, in response to intrinsic and extrinsic regulatory cues, support fidelity of DNA replication and cell division. We will focus on (1) the complex and interdependent processes that are obligatory for control of proliferation and compromised in cancer, (2) epigenetic and topological domains that are associated with distinct phases of the cell cycle that may be altered in cancer initiation and progression, and (3) the requirement for mitotic bookmarking to maintain intranuclear localization of transcriptional regulatory machinery to reinforce cell identity throughout the cell cycle to prevent malignant transformation.


Assuntos
Epigênese Genética , Neoplasias , Humanos , Ciclo Celular/genética , Divisão Celular , Neoplasias/genética , Neoplasias/patologia , Cromatina , Regulação da Expressão Gênica
8.
Results Probl Cell Differ ; 70: 339-373, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36348114

RESUMO

Epigenetic gene regulatory mechanisms play a central role in the biological control of cell and tissue structure, function, and phenotype. Identification of epigenetic dysregulation in cancer provides mechanistic into tumor initiation and progression and may prove valuable for a variety of clinical applications. We present an overview of epigenetically driven mechanisms that are obligatory for physiological regulation and parameters of epigenetic control that are modified in tumor cells. The interrelationship between nuclear structure and function is not mutually exclusive but synergistic. We explore concepts influencing the maintenance of chromatin structures, including phase separation, recognition signals, factors that mediate enhancer-promoter looping, and insulation and how these are altered during the cell cycle and in cancer. Understanding how these processes are altered in cancer provides a potential for advancing capabilities for the diagnosis and identification of novel therapeutic targets.


Assuntos
Epigênese Genética , Neoplasias , Humanos , Fenótipo , Neoplasias/genética , Neoplasias/patologia , Regulação da Expressão Gênica , Cromatina
9.
Biochim Biophys Acta Gene Regul Mech ; 1865(2): 194801, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35217218

RESUMO

The mammalian SWItch/Sucrose Non-Fermentable (mSWI/SNF) families of ATP-dependent chromatin remodeling enzymes are established co-regulators of gene expression. mSWI/SNF complexes can be assembled into three major subfamilies: BAF (BRG1 or BRM-Associated Factor), PBAF (Polybromo containing BAF), or ncBAF (non-canonical BAF) that are distinguished by the presence of mutually exclusive subunits. The mechanisms by which each subfamily contributes to the establishment or function of specific cell lineages are poorly understood. Here, we determined the contributions of the BAF, ncBAF, and PBAF complexes to myoblast proliferation via knock down (KD) of distinguishing subunits from each complex. KD of subunits unique to the BAF or the ncBAF complexes reduced myoblast proliferation rate, while KD of PBAF-specific subunits did not affect proliferation. RNA-seq from proliferating KD myoblasts targeting Baf250A (BAF complex), Brd9 (ncBAF complex), or Baf180 (PBAF complex) showed mis-regulation of a limited number of genes. KD of Baf250A specifically reduced the expression of Pax7, which is required for myoblast proliferation, concomitant with decreased binding of Baf250A to and impaired chromatin remodeling at the Pax7 gene promoter. Although Brd9 also bound to the Pax7 promoter, suggesting occupancy by the ncBAF complex, no changes were detected in Pax7 gene expression, Pax7 protein expression or chromatin remodeling at the Pax7 promoter upon Brd9 KD. The data indicate that the BAF subfamily of the mSWI/SNF enzymes is specifically required for myoblast proliferation via regulation of Pax7 expression.


Assuntos
Montagem e Desmontagem da Cromatina , Proteínas Cromossômicas não Histona , Animais , Ciclo Celular , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Mamíferos/genética , Mioblastos/metabolismo , Regiões Promotoras Genéticas
10.
Nucleic Acids Res ; 49(14): 8060-8077, 2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34289068

RESUMO

Skeletal muscle regeneration is mediated by myoblasts that undergo epigenomic changes to establish the gene expression program of differentiated myofibers. mSWI/SNF chromatin remodeling enzymes coordinate with lineage-determining transcription factors to establish the epigenome of differentiated myofibers. Bromodomains bind to acetylated lysines on histone N-terminal tails and other proteins. The mutually exclusive ATPases of mSWI/SNF complexes, BRG1 and BRM, contain bromodomains with undefined functional importance in skeletal muscle differentiation. Pharmacological inhibition of mSWI/SNF bromodomain function using the small molecule PFI-3 reduced differentiation in cell culture and in vivo through decreased myogenic gene expression, while increasing cell cycle-related gene expression and the number of cells remaining in the cell cycle. Comparative gene expression analysis with data from myoblasts depleted of BRG1 or BRM showed that bromodomain function was required for a subset of BRG1- and BRM-dependent gene expression. Reduced binding of BRG1 and BRM after PFI-3 treatment showed that the bromodomain is required for stable chromatin binding at target gene promoters to alter gene expression. Our findings demonstrate that mSWI/SNF ATPase bromodomains permit stable binding of the mSWI/SNF ATPases to promoters required for cell cycle exit and establishment of muscle-specific gene expression.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Cromatina/genética , DNA Helicases/genética , Desenvolvimento Muscular/genética , Proteínas Nucleares/genética , Fatores de Transcrição/genética , Adenosina Trifosfatases/genética , Animais , Compostos Azabicíclicos/farmacologia , Diferenciação Celular/genética , Montagem e Desmontagem da Cromatina/genética , Proteínas de Ligação a DNA/genética , Histonas/genética , Humanos , Músculo Esquelético/citologia , Músculo Esquelético/crescimento & desenvolvimento , Piridinas/farmacologia , Fatores de Transcrição/antagonistas & inibidores
11.
Adv Sci (Weinh) ; 7(23): 2002602, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33304767

RESUMO

The protein arginine methyltransferase 5 (PRMT5) is an emerging regulator of cancer and stem cells including adipogenic progenitors. Here, a new physiological role of PRMT5 in adipocytes and systemic metabolism is reported. Conditional knockout mice were generated to ablate the Prmt5 gene specifically in adipocytes (Prmt5AKO). The Prmt5AKO mice exhibit sex- and depot-dependent progressive lipodystrophy that is more pronounced in females and in visceral (than subcutaneous) white fat. The lipodystrophy and associated energy imbalance, hyperlipidemia, hepatic steatosis, glucose intolerance, and insulin resistance are exacerbated by high-fat-diet. Mechanistically, Prmt5 methylates and releases the transcription elongation factor SPT5 from Berardinelli-Seip congenital lipodystrophy 2 (Bscl2, encoding Seipin) promoter, and Prmt5AKO disrupts Seipin-mediated lipid droplet biogenesis. Prmt5 also methylates Sterol Regulatory Element-Binding Transcription Factor 1a (SREBP1a) and promotes lipogenic gene expression, and Prmt5AKO suppresses SREBP1a-dependent fatty acid metabolic pathways in adipocytes. Thus, PRMT5 plays a critical role in regulating lipid metabolism and lipid droplet biogenesis in adipocytes.

12.
Biology (Basel) ; 9(7)2020 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-32635263

RESUMO

Myogenesis is the biological process by which skeletal muscle tissue forms. Regulation of myogenesis involves a variety of conventional, epigenetic, and epigenomic mechanisms that control chromatin remodeling, DNA methylation, histone modification, and activation of transcription factors. Chromatin remodeling enzymes utilize ATP hydrolysis to alter nucleosome structure and/or positioning. The mammalian SWItch/Sucrose Non-Fermentable (mSWI/SNF) family of chromatin remodeling enzymes is essential for myogenesis. Here we review diverse and novel mechanisms of regulation of mSWI/SNF enzymes by kinases and phosphatases. The integration of classic signaling pathways with chromatin remodeling enzyme function impacts myoblast viability and proliferation as well as differentiation. Regulated processes include the assembly of the mSWI/SNF enzyme complex, choice of subunits to be incorporated into the complex, and sub-nuclear localization of enzyme subunits. Together these processes influence the chromatin remodeling and gene expression events that control myoblast function and the induction of tissue-specific genes during differentiation.

13.
Int J Mol Sci ; 21(3)2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-32019271

RESUMO

Brg1 (Brahma-related gene 1) is one of two mutually exclusive ATPases that can act as the catalytic subunit of mammalian SWI/SNF (mSWI/SfigureNF) chromatin remodeling enzymes that facilitate utilization of the DNA in eukaryotic cells. Brg1 is a phospho-protein, and its activity is regulated by specific kinases and phosphatases. Previously, we showed that Brg1 interacts with and is phosphorylated by casein kinase 2 (CK2) in a manner that regulates myoblast proliferation. Here, we use biochemical and cell and molecular biology approaches to demonstrate that the Brg1-CK2 interaction occurred during mitosis in embryonic mouse somites and in primary myoblasts derived from satellite cells isolated from mouse skeletal muscle tissue. The interaction of CK2 with Brg1 and the incorporation of a number of other subunits into the mSWI/SNF enzyme complex were independent of CK2 enzymatic activity. CK2-mediated hyperphosphorylation of Brg1 was observed in mitotic cells derived from multiple cell types and organisms, suggesting functional conservation across tissues and species. The mitotically hyperphosphorylated form of Brg1 was localized with soluble chromatin, demonstrating that CK2-mediated phosphorylation of Brg1 is associated with specific partitioning of Brg1 within subcellular compartments. Thus, CK2 acts as a mitotic kinase that regulates Brg1 phosphorylation and subcellular localization.


Assuntos
Mama/metabolismo , Caseína Quinase II/metabolismo , DNA Helicases/metabolismo , Células Epiteliais/metabolismo , Mitose , Mioblastos/metabolismo , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Mama/citologia , Montagem e Desmontagem da Cromatina , DNA Helicases/genética , Células Epiteliais/citologia , Feminino , Humanos , Camundongos , Mioblastos/citologia , Proteínas Nucleares/genética , Fosforilação , Fatores de Transcrição/genética
14.
FASEB J ; 33(12): 14556-14574, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31690123

RESUMO

Metal-regulatory transcription factor 1 (MTF1) is a conserved metal-binding transcription factor in eukaryotes that binds to conserved DNA sequence motifs, termed metal response elements. MTF1 responds to both metal excess and deprivation, protects cells from oxidative and hypoxic stresses, and is required for embryonic development in vertebrates. To examine the role for MTF1 in cell differentiation, we use multiple experimental strategies [including gene knockdown (KD) mediated by small hairpin RNA and clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9), immunofluorescence, chromatin immunopreciptation sequencing, subcellular fractionation, and atomic absorbance spectroscopy] and report a previously unappreciated role for MTF1 and copper (Cu) in cell differentiation. Upon initiation of myogenesis from primary myoblasts, both MTF1 expression and nuclear localization increased. Mtf1 KD impaired differentiation, whereas addition of nontoxic concentrations of Cu+-enhanced MTF1 expression and promoted myogenesis. Furthermore, we observed that Cu+ binds stoichiometrically to a C terminus tetra-cysteine of MTF1. MTF1 bound to chromatin at the promoter regions of myogenic genes, and Cu addition stimulated this binding. Of note, MTF1 formed a complex with myogenic differentiation (MYOD)1, the master transcriptional regulator of the myogenic lineage, at myogenic promoters. These findings uncover unexpected mechanisms by which Cu and MTF1 regulate gene expression during myoblast differentiation.-Tavera-Montañez, C., Hainer, S. J., Cangussu, D., Gordon, S. J. V., Xiao, Y., Reyes-Gutierrez, P., Imbalzano, A. N., Navea, J. G., Fazzio, T. G., Padilla-Benavides, T. The classic metal-sensing transcription factor MTF1 promotes myogenesis in response to copper.


Assuntos
Diferenciação Celular , Cobre/farmacologia , Proteínas de Ligação a DNA/metabolismo , Desenvolvimento Muscular , Mioblastos/metabolismo , Fatores de Transcrição/metabolismo , Animais , Células Cultivadas , Camundongos , Camundongos Endogâmicos C57BL , Proteína MyoD/metabolismo , Mioblastos/citologia , Mioblastos/efeitos dos fármacos , Fator MTF-1 de Transcrição
15.
PLoS One ; 14(8): e0216015, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31430278

RESUMO

JMJD6 is a member of the Jumonji C domain containing enzymes that demethylate and/or hydroxylate substrate proteins. It is a multi-functional protein that has been implicated in disparate aspects of transcriptional and post-transcriptional control of gene expression, including but not limited to enhancer and promoter binding, release of paused RNA polymerase II, control of splicing, and interaction with the translation machinery. JMJD6 contributes to multiple aspects of animal development, including adipogenesis modeled in culture. We mutated proposed or characterized domains in the JMJD6 protein to better understand the requirement for JMJD6 in adipogenic differentiation. Mutation of JMJD6 amino acids that mediate binding of iron and 2-oxogluterate, which are required cofactors for enzymatic activity, had no impact on JMJD6 function, showing that catalytic activity is not required for JMJD6 contributions to adipogenic differentiation. In addition, we documented the formation of JMJD6 oligomers and showed that catalytic activity is not required for oligomerization, as has been reported previously. We also observed no effect of mutations in the sumoylation site and in the poly-serine stretch. In contrast, mutation of the AT hook-like structure, which mediates interaction with DNA and/or RNA, compromised JMJD6 function by blocking its ability to interact with chromatin at genes that express regulators of adipogenesis. The ability of JMJD6 to interact with nucleic acids may be a critical requirement for its function in adipogenic differentiation. The requirement for the AT hook-like domain and the lack of requirement for catalytic activity giving rise to the idea that co-activation of transcription by JMJD6 may be functioning as a scaffold protein that supports the interactions of other critical regulators.


Assuntos
Motivos AT-Hook , Adipogenia , Biocatálise , Receptores de Superfície Celular/química , Receptores de Superfície Celular/metabolismo , Animais , Linhagem Celular , Camundongos , Modelos Moleculares , Mutação , Proteínas Nucleares/metabolismo , Domínios Proteicos , Receptores de Superfície Celular/genética , Sumoilação , Fatores de Transcrição/metabolismo
16.
Mol Cell Biol ; 39(19)2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31308130

RESUMO

Calcineurin (Cn) is a calcium-activated serine/threonine protein phosphatase that is broadly implicated in diverse cellular processes, including the regulation of gene expression. During skeletal muscle differentiation, Cn activates the nuclear factor of activated T-cell (NFAT) transcription factor but also promotes differentiation by counteracting the negative influences of protein kinase C beta (PKCß) via dephosphorylation and activation of Brg1, an enzymatic subunit of the mammalian SWI/SNF ATP-dependent chromatin remodeling enzyme. Here we identified four major temporal patterns of Cn-dependent gene expression in differentiating myoblasts and determined that Cn is broadly required for the activation of the myogenic gene expression program. Mechanistically, Cn promotes gene expression through direct binding to myogenic promoter sequences and facilitating the binding of Brg1, other SWI/SNF subunit proteins, and MyoD, a critical lineage determinant for skeletal muscle differentiation. We conclude that the Cn phosphatase directly impacts the expression of myogenic genes by promoting ATP-dependent chromatin remodeling and formation of transcription-competent promoters.


Assuntos
Calcineurina/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Perfilação da Expressão Gênica/métodos , Redes Reguladoras de Genes , Mioblastos/citologia , Fatores de Transcrição/metabolismo , Animais , Diferenciação Celular , Linhagem Celular , Montagem e Desmontagem da Cromatina , DNA Helicases/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Redes Reguladoras de Genes/efeitos dos fármacos , Camundongos , Proteína MyoD , Mioblastos/química , Proteínas Nucleares/metabolismo , Regiões Promotoras Genéticas/efeitos dos fármacos , Tacrolimo/farmacologia
17.
J Cell Physiol ; 234(9): 15194-15205, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30667054

RESUMO

Brahma-related gene 1 (BRG1) is one of two mutually exclusive ATPases that function as the catalytic subunit of human SWItch/Sucrose NonFermentable (SWI/SNF) chromatin remodeling enzymes. BRG1 has been identified as a tumor suppressor in some cancer types but has been shown to be expressed at elevated levels, relative to normal tissue, in other cancers. Using TCGA (The Cancer Genome Atlas) prostate cancer database, we determined that BRG1 mRNA and protein expression is elevated in prostate tumors relative to normal prostate tissue. Only 3 of 491 (0.6%) sequenced tumors showed amplification of the locus or mutation in the protein coding sequence, arguing against the idea that elevated expression due to amplification or expression of a mutant BRG1 protein is associated with prostate cancer. Kaplan-Meier survival curves showed that BRG1 expression in prostate tumors inversely correlated with survival. However, BRG1 expression did not correlate with Gleason score/International Society of Urological Pathology (ISUP) Grade Group, indicating it is an independent predictor of tumor progression/patient outcome. To experimentally assess BRG1 as a possible therapeutic target, we treated prostate cancer cells with a biologic inhibitor called ADAADi (active DNA-dependent ATPase A Domain inhibitor) that targets the activity of the SNF2 family of ATPases in biochemical assays but showed specificity for BRG1 in prior tissue culture experiments. The inhibitor decreased prostate cancer cell proliferation and induced apoptosis. When directly injected into xenografts established by injection of prostate cancer cells in mouse flanks, the inhibitor decreased tumor growth and increased survival. These results indicate the efficacy of pursuing BRG1 as both an indicator of patient outcome and as a therapeutic target.

18.
J Cell Physiol ; 234(6): 8597-8609, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30515788

RESUMO

The RUNX1 transcription factor has recently been shown to be obligatory for normal development. RUNX1 controls the expression of genes essential for proper development in many cell lineages and tissues including blood, bone, cartilage, hair follicles, and mammary glands. Compromised RUNX1 regulation is associated with many cancers. In this review, we highlight evidence for RUNX1 control in both invertebrate and mammalian development and recent novel findings of perturbed RUNX1 control in breast cancer that has implications for other solid tumors. As RUNX1 is essential for definitive hematopoiesis, RUNX1 mutations in hematopoietic lineage cells have been implicated in the etiology of several leukemias. Studies of solid tumors have revealed a context-dependent function for RUNX1 either as an oncogene or a tumor suppressor. These RUNX1 functions have been reported for breast, prostate, lung, and skin cancers that are related to cancer subtypes and different stages of tumor development. Growing evidence suggests that RUNX1 suppresses aggressiveness in most breast cancer subtypes particularly in the early stage of tumorigenesis. Several studies have identified RUNX1 suppression of the breast cancer epithelial-to-mesenchymal transition. Most recently, RUNX1 repression of cancer stem cells and tumorsphere formation was reported for breast cancer. It is anticipated that these new discoveries of the context-dependent diversity of RUNX1 functions will lead to innovative therapeutic strategies for the intervention of cancer and other abnormalities of normal tissues.


Assuntos
Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Neoplasias/metabolismo , Animais , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Mutação , Neoplasias/genética , Neoplasias/patologia , Prognóstico , Transdução de Sinais
19.
J Cell Physiol ; 233(12): 9136-9144, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-29968906

RESUMO

Breast cancer is the most common cancer in women, and accounts for ~30% of new cancer cases and 15% of cancer-related deaths. Tumor relapse and metastasis are primary factors contributing to breast cancer-related deaths. Therefore, the challenge for breast cancer treatment is to sustain remission. A driving force behind tumor relapse is breast cancer heterogeneity (both intertumor, between different patients, and intratumor, within the same tumor). Understanding breast cancer heterogeneity is necessary to develop preventive interventions and targeted therapies. A recently emerging concept is that intratumor heterogeneity is driven by cancer stem cells (CSCs) that are capable of giving rise to a multitude of different cells within a tumor. Studies have highlighted linkage of CSC formation with epithelial-to-mesenchymal transition (EMT). In this review, we summarize the current understanding of breast cancer heterogeneity, links between EMT and CSCs, regulation of EMT by Runx transcription factors, and potential therapeutic strategies targeting these processes.


Assuntos
Neoplasias da Mama/genética , Carcinogênese/genética , Subunidades alfa de Fatores de Ligação ao Core/genética , Transição Epitelial-Mesenquimal/genética , Neoplasias da Mama/patologia , Feminino , Heterogeneidade Genética , Humanos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia
20.
Mol Cancer Res ; 16(11): 1617-1624, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-30002192

RESUMO

Reconfiguration of nuclear structure and function during mitosis presents a significant challenge to resume the next cell cycle in the progeny cells without compromising structural and functional identity of the cells. Equally important is the requirement for cancer cells to retain the transformed phenotype, that is, unrestricted proliferative potential, suppression of cell phenotype, and activation of oncogenic pathways. Mitotic gene bookmarking retention of key regulatory proteins that include sequence-specific transcription factors, chromatin-modifying factors, and components of RNA Pol (RNAP) I and II regulatory machineries at gene loci on mitotic chromosomes plays key roles in coordinate control of cell phenotype, growth, and proliferation postmitotically. There is growing recognition that three distinct protein types, mechanistically, play obligatory roles in mitotic gene bookmarking: (i) Retention of phenotypic transcription factors on mitotic chromosomes is essential to sustain lineage commitment; (ii) Select chromatin modifiers and posttranslational histone modifications/variants retain competency of mitotic chromatin for gene reactivation as cells exit mitosis; and (iii) Functional components of RNAP I and II transcription complexes (e.g., UBF and TBP, respectively) are retained on genes poised for reactivation immediately following mitosis. Importantly, recent findings have identified oncogenes that are associated with target genes on mitotic chromosomes in cancer cells. The current review proposes that mitotic gene bookmarking is an extensively utilized epigenetic mechanism for stringent control of proliferation and identity in normal cells and hypothesizes that bookmarking plays a pivotal role in maintenance of tumor phenotypes, that is, unrestricted proliferation and compromised control of differentiation. Mol Cancer Res; 16(11); 1617-24. ©2018 AACR.


Assuntos
Mitose/genética , Diferenciação Celular , Epigênese Genética , Humanos , Neoplasias/genética , Neoplasias/patologia , Fenótipo
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