Sequence analysis and expression of a mouse homolog of human IkappaBL gene.
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The family of transcriptional inhibitors, IkappaBLs, are critical to the regulation of cytokine and chemokine production. We have identified the complete cDNA sequence of the mouse IkappabL gene. The predicted 381-amino-acid sequence showed evidence of two ankyrin repeats characteristic of Ikappab family proteins and 92% identity to the IkappaBL human homolog. Although human IkappaBL has been reported to be ubiquitously expressed, here we show that mouse IkappaBL is transcribed in a more tissue-specific manner. (+info)
Definition of a major p53 binding site on Ad2E1B58K protein and a possible nuclear localization signal on the Ad12E1B54K protein.
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Previous studies have established that adenovirus 2/5 early region 1B (Ad E1B) 58K protein binds p53 strongly and co-localizes with it to cytoplasmic dense bodies whilst the homologous Ad12E1B54K protein binds only weakly and co-localizes primarily to the nucleus in Ad12E1 transformed cells. We have used these properties of the E1B proteins from different viral serotypes to map the p53 binding site on the Ad2/5 protein. A set of chimaeric genes was constructed containing different proportions of the Ad12 and Ad2E1B DNA. These, together with Ad12E1A and E1B19K DNA, were transfected into baby rat kidney cells and transformed lines isolated. From an examination of the properties of these Ad12/Ad2E1B fusion proteins in co-immunoprecipitation and subcellular localization experiments it has been concluded that the p53 binding site on Ad2E1B58K protein lies between amino acids 216 and 235 and that the homologous region on Ad12E1B54K protein also binds p53. In addition, a unique nuclear localization signal is located on Ad12E1B54K between residues 228 and 239. We suggest that primary structure differences in these regions of the Ad2 and Ad12E1B proteins are responsible for the different subcellular localizations in AdE1 transformants. (+info)
Structure of CD94 reveals a novel C-type lectin fold: implications for the NK cell-associated CD94/NKG2 receptors.
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The crystal structure of the extracellular domain of CD94, a component of the CD94/NKG2 NK cell receptor, has been determined to 2.6 A resolution, revealing a unique variation of the C-type lectin fold. In this variation, the second alpha helix, corresponding to residues 102-112, is replaced by a loop, the putative carbohydrate-binding site is significantly altered, and the Ca2+-binding site appears nonfunctional. This structure may serve as a prototype for other NK cell receptors such as Ly-49, NKR-P1, and CD69. The CD94 dimer observed in the crystal has an extensive hydrophobic interface that stabilizes the loop conformation of residues 102-112. The formation of this dimer reveals a putative ligand-binding region for HLA-E and suggests how NKG2 interacts with CD94. (+info)
RFLAT-1: a new zinc finger transcription factor that activates RANTES gene expression in T lymphocytes.
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RANTES (Regulated upon Activation, Normal T cell Expressed and Secreted) is a chemoattractant cytokine (chemokine) important in the generation of inflammatory infiltrate and human immunodeficiency virus entry into immune cells. RANTES is expressed late (3-5 days) after activation in T lymphocytes. Using expression cloning, we identified the first "late" T lymphocyte associated transcription factor and named it "RANTES Factor of Late Activated T Lymphocytes-1" (RFLAT-1). RFLAT-1 is a novel, phosphorylated, zinc finger transcription factor that is expressed in T cells 3 days after activation, coincident with RANTES expression. While Rel proteins play the dominant role in RANTES gene expression in fibroblasts, RFLAT-1 is a strong transactivator for RANTES in T cells. (+info)
BLNK required for coupling Syk to PLC gamma 2 and Rac1-JNK in B cells.
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Signaling through the B cell receptor (BCR) is essential for B cell function and development. Despite the key role of Syk in BCR signaling, little is known about the mechanism by which Syk transmits downstream effectors. BLNK (B cell LiNKer protein), a substrate for Syk, is now shown to be essential in activating phospholipase C (PLC)gamma 2 and JNK. The BCR-induced PLC gamma 2 activation, but not the JNK activation, was restored by introduction of PLC gamma 2 membrane-associated form into BLNK-deficient B cells. As JNK activation requires both Rac1 and PLC gamma 2, our results suggest that BLNK regulates the Rac1-JNK pathway, in addition to modulating PLC gamma 2 localization. (+info)
Interleukin-18 binding protein: a novel modulator of the Th1 cytokine response.
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An interleukin-18 binding protein (IL-18BP) was purified from urine by chromatography on IL-18 beads, sequenced, cloned, and expressed in COS7 cells. IL-18BP abolished IL-18 induction of interferon-gamma (IFNgamma), IL-8, and activation of NF-kappaB in vitro. Administration of IL-18BP to mice abrogated circulating IFNgamma following LPS. Thus, IL-18BP functions as an inhibitor of the early Th1 cytokine response. IL-18BP is constitutively expressed in the spleen, belongs to the immunoglobulin superfamily, and has limited homology to the IL-1 type II receptor. Its gene was localized on human chromosome 11q13, and no exon coding for a transmembrane domain was found in an 8.3 kb genomic sequence. Several Poxviruses encode putative proteins highly homologous to IL-18BP, suggesting that viral products may attenuate IL-18 and interfere with the cytotoxic T cell response. (+info)
Rpp14 and Rpp29, two protein subunits of human ribonuclease P.
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In HeLa cells, the tRNA processing enzyme ribonuclease P (RNase P) consists of an RNA molecule associated with at least eight protein subunits, hPop1, Rpp14, Rpp20, Rpp25, Rpp29, Rpp30, Rpp38, and Rpp40. Five of these proteins (hPop1p, Rpp20, Rpp30, Rpp38, and Rpp40) have been partially characterized. Here we report on the cDNA cloning and immunobiochemical analysis of Rpp14 and Rpp29. Polyclonal rabbit antibodies raised against recombinant Rpp14 and Rpp29 recognize their corresponding antigens in HeLa cells and precipitate catalytically active RNase P. Rpp29 shows 23% identity with Pop4p, a subunit of yeast nuclear RNase P and the ribosomal RNA processing enzyme RNase MRP. Rpp14, by contrast, exhibits no significant homology to any known yeast gene. Thus, human RNase P differs in the details of its protein composition, and perhaps in the functions of some of these proteins, from the yeast enzyme. (+info)
The C-terminal region of hPrp8 interacts with the conserved GU dinucleotide at the 5' splice site.
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A U5 snRNP protein, hPrp8, forms a UV-induced crosslink with the 5' splice site (5'SS) RNA within splicing complex B assembled in trans- as well as in cis-splicing reactions. Both yeast and human Prp8 interact with the 5'SS, branch site, polypyrimidine tract, and 3'SS during splicing. To begin to define functional domains in Prp8 we have mapped the site of the 5'SS crosslink within the hPrp8 protein. Immunoprecipitation analysis limited the site of crosslink to the C-terminal 5060-kDa segment of hPrp8. In addition, size comparison of the crosslink-containing peptides generated with different proteolytic reagents with the pattern of fragments predicted from the hPrp8 sequence allowed for mapping of the crosslink to a stretch of five amino acids in the C-terminal portion of hPrp8 (positions 1894-1898). The site of the 5'SS:hPrp8 crosslink falls within a segment spanning the previously defined polypyrimidine tract recognition domain in yPrp8, suggesting that an overlapping region of Prp8 may be involved both in the 5'SS and polypyrimidine tract recognition events. In the context of other known interactions of Prp8, these results suggest that this protein may participate in formation of the catalytic center of the spliceosome. (+info)