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1.
Biochemistry ; 50(32): 6920-32, 2011 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-21721535

RESUMO

S100A4, a member of the Ca(2+)-activated S100 protein family, regulates the motility and invasiveness of cancer cells. Moreover, high S100A4 expression levels correlate with poor patient survival in several cancers. Although biochemical, biophysical, and structural data indicate that S100A4 is a noncovalent dimer, it is unknown if two functional S100A4 monomers are required for the productive recognition of protein targets and the promotion of cell invasion. To address this question, we created covalently linked S100A4 dimers using a glycine rich flexible linker. The single-chain S100A4 (sc-S100A4) proteins exhibited wild-type affinities for calcium and nonmuscle myosin-IIA, retained the ability to regulate nonmuscle myosin-IIA assembly, and promoted tumor cell invasion when expressed in S100A4-deficient colon carcinoma cells. Mutation of the two calcium-binding EF-hands in one monomer, while leaving the other monomer intact, caused a 30-60-fold reduction in binding affinity for nonmuscle myosin-IIA concomitant with a weakened ability to regulate the monomer-polymer equilibrium of nonmuscle myosin-IIA. Moreover, sc-S100A4 proteins with one monomer deficient in calcium responsiveness did not support S100A4-mediated colon carcinoma cell invasion. Cross-linking and titration data indicate that the S100A4 dimer binds a single myosin-IIA target peptide. These data are consistent with a model in which a single peptide forms interactions in the vicinity of the canonical target binding cleft of each monomer in such a manner that both target binding sites are required for the efficient interaction with myosin-IIA.


Assuntos
Miosina não Muscular Tipo IIA/metabolismo , Proteínas S100/fisiologia , Sequência de Aminoácidos , Western Blotting , Linhagem Celular Tumoral , Cromatografia em Gel , Dicroísmo Circular , Dimerização , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Invasividade Neoplásica , Proteína A4 de Ligação a Cálcio da Família S100 , Proteínas S100/química , Proteínas S100/metabolismo
2.
Mol Biol Cell ; 18(8): 3144-55, 2007 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-17567956

RESUMO

In mammalian nonmuscle cells, the mechanisms controlling the localized formation of myosin-II filaments are not well defined. To investigate the mechanisms mediating filament assembly and disassembly during generalized motility and chemotaxis, we examined the EGF-dependent phosphorylation of the myosin-IIA heavy chain in human breast cancer cells. EGF stimulation of MDA-MB-231 cells resulted in transient increases in both the assembly and phosphorylation of the myosin-IIA heavy chains. In EGF-stimulated cells, the myosin-IIA heavy chain is phosphorylated on the casein kinase 2 site (S1943). Cells expressing green fluorescent protein-myosin-IIA heavy-chain S1943E and S1943D mutants displayed increased migration into a wound and enhanced EGF-stimulated lamellipod extension compared with cells expressing wild-type myosin-IIA. In contrast, cells expressing the S1943A mutant exhibited reduced migration and lamellipod extension. These observations support a direct role for myosin-IIA heavy-chain phosphorylation in mediating motility and chemotaxis.


Assuntos
Movimento Celular , Cadeias Pesadas de Miosina/metabolismo , Miosina não Muscular Tipo IIA/metabolismo , Substituição de Aminoácidos , Caseína Quinase II/metabolismo , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Fator de Crescimento Epidérmico/farmacologia , Adesões Focais/efeitos dos fármacos , Humanos , Proteínas Mutantes/metabolismo , Fosforilação/efeitos dos fármacos , Fosfosserina/metabolismo , Ligação Proteica/efeitos dos fármacos , Isoformas de Proteínas/metabolismo , Transporte Proteico/efeitos dos fármacos , Pseudópodes/efeitos dos fármacos , Solubilidade/efeitos dos fármacos
3.
Mol Cancer Res ; 16(4): 567-579, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29555893

RESUMO

RNA-binding proteins (RBP) and noncoding RNAs (ncRNA), such as long noncoding RNAs (lncRNA) and microRNAs (miRNA), control co- and posttranscriptional gene regulation (PTR). At the PTR level, RBPs and ncRNAs contribute to pre-mRNA processing, mRNA maturation, transport, localization, turnover, and translation. Deregulation of RBPs and ncRNAs promotes the onset of cancer progression and metastasis. Both RBPs and ncRNAs are altered by signaling cascades to cooperate or compete with each other to bind their nucleic acid targets. Most importantly, transforming growth factor-beta (TGFß) signaling plays a significant role in controlling gene expression patterns by targeting RBPs and ncRNAs. Because of TGFß signaling in cancer, RBP-RNA or RNA-RNA interactions are altered and cause enhanced cell growth and tumor cell dissemination. This review focuses on the emerging concepts of TGFß signaling on posttranscriptional gene regulation and highlights the implications of RBPs and ncRNAs in cancer progression and metastasis. Mol Cancer Res; 16(4); 567-79. ©2018 AACR.


Assuntos
MicroRNAs/genética , Neoplasias/genética , RNA Longo não Codificante/genética , Proteínas de Ligação a RNA/genética , Fator de Crescimento Transformador beta/metabolismo , Animais , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Metástase Neoplásica , Neoplasias/metabolismo , Processamento Pós-Transcricional do RNA , Transdução de Sinais
4.
Oncotarget ; 6(41): 43620-34, 2015 Dec 22.
Artigo em Inglês | MEDLINE | ID: mdl-26498364

RESUMO

The RNA binding protein CELF1 (also known as CUGBP1) is emerging as a critical regulator of cancer cell proliferation and apoptosis. Here, to provide a global prospective of CELF1 regulation of oral squamous cell carcinoma, we performed RNA-sequencing in oral cancer cells and CELF1 overexpression analysis in non-malignant human oral keratinocytes. Our approaches identified 1283 mRNAs differentially regulated as a function of CELF1 expression and more importantly CELF1 promoted alternative splicing of several target pre-mRNAs, which are known to be involved in various cancer biological processes. Overexpression of CELF1 in non-malignant human oral keratinocytes protected cells against oxidative damage and altered gene expression patterns. Finally, we provide evidence that reduction of CELF1 protein using a xenograft tumorigenesis mouse model decreased tumor growth. Altogether, these data provided a comprehensive view of the CELF1 mRNA regulatory network in oral cancer and suggests that CELF1 and/or its target mRNAs are viable candidates for therapeutic intervention.


Assuntos
Proteínas CELF1/metabolismo , Carcinoma de Células Escamosas/patologia , Regulação Neoplásica da Expressão Gênica/genética , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias Bucais/patologia , Animais , Western Blotting , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Linhagem Celular , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/metabolismo , Xenoenxertos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imunoprecipitação , Masculino , Camundongos , Camundongos Nus , Neoplasias Bucais/genética , Neoplasias Bucais/metabolismo , Reação em Cadeia da Polimerase , Carcinoma de Células Escamosas de Cabeça e Pescoço , Transcriptoma , Transfecção
5.
Mol Biol Cell ; 21(15): 2598-610, 2010 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-20519440

RESUMO

S100A4, a member of the S100 family of Ca(2+)-binding proteins, is directly involved in tumor metastasis. In addition to its expression in tumor cells, S100A4 is expressed in normal cells and tissues, including fibroblasts and cells of the immune system. To examine the contribution of S100A4 to normal physiology, we established S100A4-deficient mice by gene targeting. Homozygous S100A4(-/-) mice are fertile, grow normally and exhibit no overt abnormalities; however, the loss of S100A4 results in impaired recruitment of macrophages to sites of inflammation in vivo. Consistent with these observations, primary bone marrow macrophages (BMMs) derived from S100A4(-/-) mice display defects in chemotactic motility in vitro. S100A4(-/-) BMMs form unstable protrusions, overassemble myosin-IIA, and exhibit altered colony-stimulating factor-1 receptor signaling. These studies establish S100A4 as a regulator of physiological macrophage motility and demonstrate that S100A4 mediates macrophage recruitment and chemotaxis in vivo.


Assuntos
Quimiotaxia , Macrófagos/citologia , Proteínas S100/metabolismo , Actomiosina/metabolismo , Animais , Células da Medula Óssea/citologia , Contagem de Células , Extensões da Superfície Celular/efeitos dos fármacos , Extensões da Superfície Celular/metabolismo , Quimiotaxia/efeitos dos fármacos , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Humanos , Inflamação/metabolismo , Inflamação/patologia , Fator Estimulador de Colônias de Macrófagos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Knockout , Modelos Biológicos , Receptor de Fator Estimulador de Colônias de Macrófagos/metabolismo , Proteína A4 de Ligação a Cálcio da Família S100 , Proteínas S100/deficiência , Transdução de Sinais/efeitos dos fármacos
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