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1.
Mol Cell ; 31(2): 255-65, 2008 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-18657507

RESUMO

The antiapoptotic protein Bcl-2 inhibits Ca2+ release from the endoplasmic reticulum (ER). One proposed mechanism involves an interaction of Bcl-2 with the inositol 1,4,5-trisphosphate receptor (IP3R) Ca2+ channel localized with Bcl-2 on the ER. Here we document Bcl-2-IP3R interaction within cells by FRET and identify a Bcl-2 interacting region in the regulatory and coupling domain of the IP3R. A peptide based on this IP3R sequence displaced Bcl-2 from the IP3R and reversed Bcl-2-mediated inhibition of IP3R channel activity in vitro, IP3-induced ER Ca2+ release in permeabilized cells, and cell-permeable IP3 ester-induced Ca2+ elevation in intact cells. This peptide also reversed Bcl-2's inhibition of T cell receptor-induced Ca2+ elevation and apoptosis. Thus, the interaction of Bcl-2 with IP3Rs contributes to the regulation of proapoptotic Ca2+ signals by Bcl-2, suggesting the Bcl-2-IP3R interaction as a potential therapeutic target in diseases associated with Bcl-2's inhibition of cell death.


Assuntos
Apoptose , Sinalização do Cálcio , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Sequência de Aminoácidos , Animais , Apoptose/efeitos dos fármacos , Complexo CD3/metabolismo , Células COS , Sinalização do Cálcio/efeitos dos fármacos , Chlorocebus aethiops , Transferência Ressonante de Energia de Fluorescência , Humanos , Receptores de Inositol 1,4,5-Trifosfato/química , Ativação do Canal Iônico/efeitos dos fármacos , Células Jurkat , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/farmacologia , Ligação Proteica/efeitos dos fármacos , Mapeamento de Interação de Proteínas , Estrutura Terciária de Proteína , Receptores de Antígenos de Linfócitos T/metabolismo
2.
Blood ; 117(10): 2924-34, 2011 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-21193695

RESUMO

Bcl-2 contributes to the pathophysiology and therapeutic resistance of chronic lymphocytic leukemia (CLL). Therefore, developing inhibitors of this protein based on a thorough understanding of its mechanism of action is an active and promising area of inquiry. One approach centers on agents (eg, ABT-737) that compete with proapoptotic members of the Bcl-2 protein family for binding in the hydrophobic groove formed by the BH1-BH3 domains of Bcl-2. Another region of Bcl-2, the BH4 domain, also contributes to the antiapoptotic activity of Bcl-2 by binding to the inositol 1,4,5-trisphosphate receptor (IP3R) Ca²(+) channel, inhibiting IP(3)-dependent Ca²(+) release from the endoplasmic reticulum. We report that a novel synthetic peptide, modeled after the Bcl-2-interacting site on the IP3R, binds to the BH4 domain of Bcl-2 and functions as a competitive inhibitor of the Bcl-2-IP3R interaction. By disrupting the Bcl-2-IP3R interaction, this peptide induces an IP3R-dependent Ca²(+) elevation in lymphoma and leukemia cell lines and in primary CLL cells. The Ca²(+) elevation evoked by this peptide induces apoptosis in CLL cells, but not in normal peripheral blood lymphocytes, suggesting the involvement of the Bcl-2-IP3R interaction in the molecular mechanism of CLL and indicating the potential merit of targeting this interaction therapeutically.


Assuntos
Apoptose/fisiologia , Cálcio/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Leucemia Linfocítica Crônica de Células B/metabolismo , Peptídeos/farmacologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ligação Competitiva , Western Blotting , Linhagem Celular Tumoral , Humanos , Imunoprecipitação
3.
Nat Genet ; 35(2): 139-47, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-12973349

RESUMO

Schistosoma japonicum causes schistosomiasis in humans and livestock in the Asia-Pacific region. Knowledge of the genome of this parasite should improve understanding of schistosome-host interactions, biomedical aspects of schistosomiasis and invertebrate evolution. We assigned 43,707 expressed sequence tags (ESTs) derived from adult S. japonicum and their eggs to 13,131 gene clusters. Of these, 35% shared no similarity with known genes and 75% had not been reported previously in schistosomes. Notably, S. japonicum encoded mammalian-like receptors for insulin, progesterone, cytokines and neuropeptides, suggesting that host hormones, or endogenous parasite homologs, could orchestrate schistosome development and maturation and that schistosomes modulate anti-parasite immune responses through inhibitors, molecular mimicry and other evasion strategies.


Assuntos
DNA de Helmintos/genética , Evolução Molecular , Schistosoma japonicum/genética , Sequência de Aminoácidos , Animais , DNA Complementar/genética , Genes de Helmintos , Interações Hospedeiro-Parasita , Humanos , Mamíferos , Dados de Sequência Molecular , Filogenia , Schistosoma japonicum/classificação , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Especificidade da Espécie
4.
Proc Natl Acad Sci U S A ; 106(34): 14397-402, 2009 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-19706527

RESUMO

Although the presence of a BH4 domain distinguishes the antiapoptotic protein Bcl-2 from its proapoptotic relatives, little is known about its function. BH4 deletion converts Bcl-2 into a proapoptotic protein, whereas a TAT-BH4 fusion peptide inhibits apoptosis and improves survival in models of disease due to accelerated apoptosis. Thus, the BH4 domain has antiapoptotic activity independent of full-length Bcl-2. Here we report that the BH4 domain mediates interaction of Bcl-2 with the inositol 1,4,5-trisphosphate (IP3) receptor, an IP3-gated Ca(2+) channel on the endoplasmic reticulum (ER). BH4 peptide binds to the regulatory and coupling domain of the IP3 receptor and inhibits IP3-dependent channel opening, Ca(2+) release from the ER, and Ca(2+)-mediated apoptosis. A peptide inhibitor of Bcl-2-IP3 receptor interaction prevents these BH4-mediated effects. By inhibiting proapoptotic Ca(2+) signals at their point of origin, the Bcl-2 BH4 domain has the facility to block diverse pathways through which Ca(2+) induces apoptosis.


Assuntos
Apoptose/fisiologia , Cálcio/metabolismo , Retículo Endoplasmático/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/fisiologia , Sequência de Aminoácidos , Animais , Anticorpos/farmacologia , Apoptose/efeitos dos fármacos , Sítios de Ligação , Western Blotting , Complexo CD3/imunologia , Sinalização do Cálcio/efeitos dos fármacos , Linhagem Celular Tumoral , Retículo Endoplasmático/efeitos dos fármacos , Humanos , Imunoprecipitação , Receptores de Inositol 1,4,5-Trifosfato/genética , Células Jurkat , Microscopia de Fluorescência , Dados de Sequência Molecular , Oligopeptídeos/síntese química , Oligopeptídeos/farmacologia , Ligação Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/química , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo
5.
Biochim Biophys Acta ; 1793(6): 971-8, 2009 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19056433

RESUMO

Bcl-2 is the founding member of a large family of apoptosis regulating proteins. Bcl-2 is a prime target for novel therapeutics because it is elevated in many forms of cancer and contributes to cancer progression and therapy resistance based on its ability to inhibit apoptosis. Bcl-2 interacts with proapoptotic members of the Bcl-2 family to inhibit apoptosis and small molecules that disrupt this interaction have already entered the cancer therapy arena. A separate function of Bcl-2 is to inhibit Ca2+ signals that promote apoptosis. This function is mediated through interaction of the Bcl-2 BH4 domain with the inositol 1,4,5-trisphosphate receptor (IP3R) Ca2+ channel. A novel peptide inhibitor of this interaction enhances proapoptotic Ca2+ signals. In preliminary experiments this peptide enhanced ABT-737 induced apoptosis in chronic lymphocytic leukemia cells. These findings draw attention to the BH4 domain as a potential therapeutic target. This review summarizes what is currently known about the BH4 domain of Bcl-2, its interaction with the IP3R and other proteins, and the part it plays in Bcl-2's anti-apoptotic function. In addition, we speculate on how the BH4 domain of Bcl-2 can be targeted therapeutically not only for diseases associated with apoptosis resistance, but also for diseases associated with accelerated cell death.


Assuntos
Apoptose/fisiologia , Sinalização do Cálcio/fisiologia , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular Tumoral , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Neoplasias/metabolismo , Peptídeos/química , Peptídeos/genética , Peptídeos/metabolismo , Conformação Proteica , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/química , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteína bcl-X/química , Proteína bcl-X/genética , Proteína bcl-X/metabolismo
6.
Gene ; 315: 71-8, 2003 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-14557066

RESUMO

A novel human gene, named as hTLP19, was isolated and characterized as secretory protein by combining bioinformatics tools with experiments. The hTLP19 encodes 172 amino acid residues with signal peptide in its N-terminal and a thioredoxin (Trx) domain that is homologous with some genes in Mus musculus, Xenopus laevis, etc. Moreover, the result from insulin reduction assay indicated that the hTLP19 protein has Trx enzymatic activity. By comparing full-length cDNA with human genomic DNA, the hTLP19 gene might have seven coding exons spanning 35 kb of genomic DNA on the human chromosome 1p32.3. Northern blot analysis showed that human hTLP19 is expressed widely in many tissues with 1.6 kb band and extra 1.2 kb band in placenta. Subcellular localization and immunoblotting assays indicated that hTLP19 might be secreted out of cell through trans-Golgi network (TGN).


Assuntos
DNA Complementar/isolamento & purificação , Isomerases de Dissulfetos de Proteínas/genética , Sequência de Aminoácidos , Animais , Northern Blotting , Células COS , Chlorocebus aethiops , Clonagem Molecular , DNA Complementar/química , DNA Complementar/genética , Expressão Gênica , Complexo de Golgi/metabolismo , Humanos , Dados de Sequência Molecular , Filogenia , Proteína Dissulfeto Redutase (Glutationa) , Isomerases de Dissulfetos de Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Tiorredoxinas/metabolismo
7.
Artigo em Inglês | MEDLINE | ID: mdl-12098761

RESUMO

We have cloned a new member of the granin family, termed human secretogranin III (SgIII), that encodes 468 amino acid residues. The human SgIII protein possesses an N-terminal signal peptide, seven dibasic sites, and the repeated DSTK sequences. These structure characteristics are similar to other members of secretogranin family. The human SgIII has homologous proteins in mouse, rat, and Xenopus laevis. Genomic organization shows the gene includes 12 coding exons spanning 39 kb of genomic DNA on the human chromosome 15. Human SgIII is expressed widely as showed in Northern blot and its cDNA hybridizes to 2.2 kb and 1.9 kb bands in many tissues, with two additional 4.5 kb and 3.3 kb bands in brain. Subcelluar localization and immunoblotting indicated SgIII was secreted out of cell through trans-Golgi network (TGN). SgIII may take effect in the biogenesis of secretory granules as a helper protein and be involved in the production or release of peptide hormones in the regulated secretory pathway.


Assuntos
Proteínas/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Células COS , Cromograninas , Clonagem Molecular , DNA Complementar/química , DNA Complementar/genética , Éxons , Feminino , Expressão Gênica , Genes/genética , Humanos , Immunoblotting , Íntrons , Microscopia de Fluorescência , Dados de Sequência Molecular , Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos
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