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Molecular cloning of a myoD-like gene from the parasitic nematode, Trichinella spiralis

MOLECULAR AND BIOCHEMICAL PARASITOLOGY 1996年第2期81卷 137-149页

作者： Connolly, B Trenholme, K Smith, DF UNIV LONDON IMPERIAL COLL SCI TECHNOL & MED DEPT BIOCHEM WELLCOME CTR PARASIT INFECT LONDON SW7 2AZ ENGLAND UNIV BERN INST PARASITOL CH-3012 BERN SWITZERLAND

The infective larval stage of the nematode Trichinella spiralis is an intracellular parasite of skeletal muscle cells. Infection with T. spiralis results in dedifferentiation of the host cell and the formation of a host/parasite complex. A gene encoding a T. spiralis helix-loop-helix (HLH) protein with homology to the myogenic transcription factor, MyoD, and to the Caenorhabditis elegans protein, CeMyoD, has been identified and partially characterized. The tsmyd-1 gene is expressed constitutively during the muscle-larval and adult stages. A purified recombinant Tsmyd-1 protein, expressed in Escherichia coli, binds to a high affinity mouse MyoD DNA binding site in vitro. The present study describes the first HLH gene to be identified in Trichinella.

关键词： Trichinella muscle-stage larva basic helix-loop-helix MyoD CeMyoD nau

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Phosphorylation of E47 as a potential determinant of B-cell-specific activity

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MOLECULAR AND CELLULAR BIOLOGY 1996年第12期16卷 6900-6908页

作者： Sloan, SR Shen, CP McCarrickWalmsley, R Kadesch, T UNIV PENN SCH MED HOWARD HUGHES MED INST PHILADELPHIA PA 19104 USA UNIV PENN SCH MED DEPT GENET PHILADELPHIA PA 19104 USA UNIV PENN SCH MED DEPT PATHOL & LAB MED PHILADELPHIA PA 19104 USA

The E2A gene encodes two basic helix-loop-helix proteins designated E12 and E47. Although these proteins are widely expressed, they are required only for the B-lymphocyte lineage where DNA binding is mediated distinctively by E47 homodimers. By studying the properties of Delta E47, an N-terminal truncation of E47, we provide evidence that phosphorylation may contribute to B-cell-specific DNA binding by E47. Two serines N terminal to the Delta E37 basic helix-loop-helix domain were found to be phosphorylated in a variety of cell types bat were hypophosphorylated in B cells, Phosphorylating these serines in vitro inhibited DNA binding by Delta E47 homodimers but not by Delta E47-containing heterodimers, such as Delta E47:MyoD. These results argue that hypophosphorylation may be a prerequisite for activity of E47 homodimers in B cells, suggesting the use of an inductive (nonstochastic) step in early B-cell development.

关键词： .widely argue cell types Howard Hughes Medical DNA binding bHLH cell development MyoD hypophosphorylated heterodimers basic helix-loop-helix serines binding by homodimers

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In vivo interactions of the Drosophila Hairy and Runt transcriptional repressors with target promoters

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EMBO JOURNAL 1996年第24期15卷 7088-7098页

作者： Jimenez, G Pinchin, SM IshHorowicz, D IMPERIAL CANC RES FUND LONDON WC2A 3PXENGLAND

The Hairy and Runt pair-rule proteins regulate Drosophila segmentation by repressing transcription. To explore the ability of these proteins to function as promoter-bound regulators in vivo, we examined the effects of Hairy and Runt derivatives containing heterologous transcriptional activation domains (Hairy(Act) and Run(Act)). Using this approach, we find that Hairy and Runt efficiently target such activation domains to specific segmentation gene promoters, leading to rapid induction of transcription. Our results strongly suggest that Hairy normally acts as a promoter-bound repressor of fushi tarazu, runt and odd-skipped, and that Runt directly represses even-skipped. We also show that expressing Hairy(Act) in early blastoderm embryos causes ectopic Sex-lethal expression and mate-specific lethality, implying that the Hairy-related denominator element Deadpan represses Sex-lethal during sex determination by directly recognizing the early Sex-lethal promoter.

关键词： basic helix-loop-helix embryonic patterning segmentation sex determination transcriptional repression

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Knock-out effects of transcription factor GATA-1 on early erythropoiesis

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MOLECULES AND CELLS 1996年第2期6卷 176-182页

作者： Kim, CG Department of Biology College of Natural Sciences Hanyang University Seoul 133-791 Korea

Previous studies have demonstrated that GATA-1 deficient embryonic stem cells fail to produce mature red blood cells either in vivo or in vitro, though GATA-1 is known to be expressed in other hematopoietic cells. To gain further insight into the role of GATA-1 in early erythropoiesis, characteristics of the differentiating GATA-1-deficient embryonic stem (ES) cells in a simple suspension culture were monitored for their hematopoietic differentiation potential and for expressivity of several erythroid marker genes. The data showed that definitive erythropoietic lineage cells failed to mature beyond the pro-erythroblast stage from the colonies derived from the differentiating GATA-1-deficient ES cells, while all other hematopoietic lineage cell types were generated efficiently. Interestingly, beta H1, the first beta-type globin to appear in the primitive erythroid cells, initiated its expression normally but the expression was not maintained afterwards in the differentiating GATA-1 deficient ES cells, while the other beta-type globins were not expressed at all. All other tested erythroid messages were expressed normally. The data suggests that GATA-1 is critical for the maintenance of relatively early erythropoiesis, but not for the onset of either primitive or definitive erythropoiesis, nor even for the onset of beta h1-globin expression in primitive erythroid cells.

关键词： bHLH basic helix-loop-helix ES embryonic stem EB embryoid body LCR locus control region LIF leukemia-inhibitory factor

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BETA3, a novel helix-loop-helix protein, can act as a negative regulator of BETA2 and MyoD-responsive genes

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MOLECULAR AND CELLULAR BIOLOGY 1996年第2期16卷 626-633页

作者： Peyton, M Stellrecht, CMM Naya, FJ Huang, HP Samora, PJ Tsai, MJ BAYLOR COLL MED DEPT CELL BIOLHOUSTONTX 77030

Using degenerate PCR cloning we have identified a novel basic helix-loop-helix (bHLH) transcription factor, BETA3, from a hamster insulin tumor (HIT) cell cDNA library. Sequence analysis revealed that this factor belongs to the class B bHLH family and has the highest degree of homology with another bHLH transcription factor recently isolated in our laboratory, BETA2 (neuroD) (J. E. Lou, S. M. Hollenberg, L. Snider, D. L. Turner, N. Lipnick, and H. Weintraub, Science 268:836-844, 1995;F. J. Naya, C. M. M. Stellrecht, and M.-J. Tsai, Genes Dev. 8:1009-1019, 1995). BETA2 is a brain- and pancreatic-islet-specific bHLH transcription factor and is largely responsible for the tissue-specific expression of th. insulin gene. BETA3 was found to be tissue restricted, with the highest levels of expression in HIT, lung, kidney, and brain cells. Surprisingly, despite the homology between BETA2 and BETA3 and its intact basic region, BETAS is unable to bind the insulin E box in bandshift analysis as a homodimer or as a heterodimer with the class A bHLH factors E12, E47, or BETA1. Instead, BETA3 inhibited both the E47 homodimer and the E47/BETA2 heterodimer binding to the insulin E box. In addition, BETA3 greatly repressed the BETA2/E47 induction of the insulin enhancer in HIT cells as well as the MyoD/E47 induction of a muscle-specific E box in the myoblast cell line C2C12. In contrast, expression of BETA3 had no significant effect on the GAL4-N P16 transcriptional activity. Immunoprecipitation analysis demonstrates that the mechanism of repression in via direct protein-protein interaction, presumably by heterodimerization between BETA3 and class A bHLH factors.

关键词： .Degenerate regulator negative sequence analysis insulin gene heterodimer NeuroD hit Genes Dev Hamster basic helix-loop-helix basic region homodimer Novel helix-loop-helix Protein degree of homology

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The Drosophila melanogaster similar bHLH-PAS gene encodes a protein related to human hypoxia-inducible factor 1 alpha and Drosophila single-minded

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GENE 1996年第2期172卷 249-254页

作者： Nambu, JR Chen, W Hu, S Crews, ST UNIV N CAROLINA SCH MEDDEPT BIOCHEM & BIOPHYSCHAPEL HILLNC 27599 UNIV CALIF LOS ANGELES INST MOLEC BIOLDEPT BIOLLOS ANGELESCA 90024

The Drosophila melanogaster (Dm) similar (sima) gene was isolated using a low-stringency hybridization screen employing a Dm single-minded gene basic helix-loop-helix (bHLH) DNA probe. sima is a member of the bHLH-PAS gene family and the conceptual protein shares a number of structural features, including a bHLH domain, PAS domain, and homopolymeric amino acid stretches. Sima is most closely related to the human hypoxia-inducible factor 1 alpha bHLH-PAS protein. In situ hybridization experiments reveal that sima is transcribed in most or all cells throughout embryogenesis. It has been cytologically mapped to position 99D on the third chromosome, and is not closely linked to other known bHLH-PAS genes.

关键词： sima basic helix-loop-helix transcription factor embryogenesis

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Differences between MyoD DNA binding and activation site requirements revealed by functional random sequence selection

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MOLECULAR AND CELLULAR BIOLOGY 1996年第7期16卷 3893-3900页

作者： Huang, J Blackwell, TK Kedes, L Weintraub, H UNIV SO CALIF SCH MED DEPT BIOCHEM & MOLEC BIOL LOS ANGELES CA 90033 USA FRED HUTCHINSON CANC RES CTR HOWARD HUGHES MED INST SEATTLE WA 98104 USA UNIV SO CALIF SCH MED INST MED GENET LOS ANGELES CA 90033 USA CTR BLOOD RES BOSTON MA 02115 USA HARVARD UNIV SCH MED BOSTON MA 02115 USA

A method has been developed for selecting functional enhancer/promoter sites from random DNA sequences in higher eukaryotic cells. Of sequences that were thus selected for transcriptional activation by the muscle-specific basic helix-loop-helix protein MyoD, only a subset are similar to the preferred in vitro binding consensus, and in the same promoter context an optimal in vitro binding site was inactive. Other sequences with full transcriptional activity instead exhibit sequence preferences that, remarkably, are generally either identical or very similar to those found in naturally occurring muscle-specific promoters. This first systematic examination of the relation between DNA binding and transcriptional activation by basic helix-loop-helix proteins indicates that binding per se is necessary but not sufficient for transcriptional activation by MyoD and implies a requirement for other DNA sequence-dependent interactions or conformations at its binding site.

关键词： .instead random preferred DNA binding Transcriptional activation Preferences bHLH inactive binding site very similar basic helix-loop-helix vitro binding only subset enhancer/promoter sequence selection Activation Site promoter context helix-loop-helix proteins activation by MyoD

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3 IMMEDIATE-EARLY GENE RESPONSE ELEMENTS IN THE PROXIMAL PREPROTACHYKININ-A PROMOTER IN 2 FUNCTIONALLY DISTINCT DOMAINS

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NEUROSCIENCE 1995年第4期66卷 921-932页

作者： PATERSON, JM MENDELSON, SC MCALLISTER, J MORRISON, CF DOBSON, S GRACE, C QUINN, JP ROYAL EDINBURGH & ASSOCIATED HOSP MRCBRAIN METAB UNITEDINBURGH EH10 5HFMIDLOTHIANSCOTLAND

The preprotachykinin-A promoter contains two blocks of DNA sequence, with a high degree of homology to one another, both containing activator protein 1/cAMP response element-like elements which constitute cis-acting regulatory domains. These two domains are differentially regulated in HeLa cells and primary cultures of dorsal root ganglion neurons when they are placed in the context of a reporter gene driven by the c-fos minimum promoter. One of the domains, corresponding to a region of the preprotachykinin promoter spanning nucleotides -345 to -308, contains two activator protein 1 elements adjacent to an E-box binding protein consensus sequence. Both of the activator protein 1 elements can bind a complex containing c-fos/c-fos related antigen proteins and the adjacent E-box element is specifically recognized by proteins present in HeLa nuclear extract. This domain requires the synergistic action of both activator protein 1 elements to drive expression of the reporter gene in both HeLa and dorsal root ganglion cells. The second or proximal domain spans nucleotides -198 to -155 and contains a previously characterized activator protein 1/cAMP response element/ATF enhancer element which, in contrast to the activator protein 1 elements in the distal domain, functions in both HeLa and dorsal root ganglion cells as one copy. This domain is differentially regulated in HeLa and dorsal root ganglia. The previously characterized enhancer activity is repressed in the context of the extended cis-acting domain in HeLa cells but remains active in dorsal root ganglion, although no further enhancement of activity supported by the single enhancer is observed when in the context of the extended sequence. This proximal domain, in addition to binding the enhancer complex, can be bound by at least two other complexes, one of which binds to an E-box consensus sequence. As the elements corresponding to the E-box consensus in both domains cross-compete for binding of specific complex(es)

关键词： API activator protein 1 bHLH basic helix-loop-helix CAT chloramphenicol acetyltransferase CRE cAMP response element CREB cAMP response element binding protein DRG dorsal root ganglia EMSA electrophoretic mobility shift assay Fra c-fos-related antigen GALV Gibbon Ape leukaemia virus NGF nerve growth factor rPPT rat preprotachykinin-A

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Mechanisms of tax regulation of human T cell leukemia virus type I gene expression

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Journal of Biomedical Science 1995年第1期2卷 17-29页

作者： Franklin, Audrey A. Nyborg, Jennifer K. Department of Biochemistry and Molecular Biology Colorado State University Fort Collins 80523 CO United States

During the last several years, the human T cell leukemia virus type I (HTLV-I) has become recognized as an important cause for public health concern throughout the world. HTLV-I is the causative agent of a variety of clinical diseases, including an aggressive lymphoproliferative disorder named adult T cell leukemia. HTLV-I induces pathogenicity in the infected host cell through the synthesis of a virally encoded protein called Tax. Expression of Tax is critical to the life cycle of the virus, as the protein greatly increases the efficiency of HTLV-I gene transcription and replication. Furthermore, Tax has been shown to deregulate the transcription of many cellular genes, leading to the hypothesis that the presence of Tax promotes unchecked growth in the HTLV-I-infected cell. The mechanism of Tax trans-activation of HTLV-I gene expression is not known. Tax does not bind directly to the Tax-responsive promoter elements of the virus, but appears to function through interaction with certain cellular DNA binding proteins, including activating transcription factor 2 and cAMP response element binding protein that recognize these sequences. This review summarizes some of the recent work in the field aimed at elucidating the mechanism of Tax trans-activation of HTLV-I gene expression. © 1995 National Science Council.

关键词： Activating transcription factor basic helix-loop-helix basic-leucine zipper proteins cAMP response element binding protein NF-κB Trans-activation

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Muscle basic helix-loop-helix proteins and the regulation of myogenesis

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CURRENT OPINION IN GENETICS & DEVELOPMENT 1992年第2期2卷 243-248页

作者： Wright, Woodring E. Univ Texas Southwestern Med Ctr Dallas Dept Cell Biol & Neurosci 5323 Harry Hines Blvd Dallas TX 75235 USA

Significant progress has been made in defining the structural motifs that distinguish the muscle-specific from other basic helix-loop-helix proteins. Evidence is accumulating for multiple levels of regulation of the e... 详细信息

关键词： bHLH basic helix-loop-helix EMSA electrophoretic mobility shift assay Id inhibitor of differentiation IGF insulin-like growth factor LTR long terminal repeat PKC protein kinase C TGF transforming growth factor

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