Biomedical subjects

Abhijit Raha

Publications and source records attributed to Abhijit Raha.

5 recordsLinked to original sources

PubMed2005

IFN-gamma-stimulated transcriptional activation by IFN-gamma-activated transcriptional element-binding factor 1 occurs via an inducible interaction with CAAAT/enhancer-binding protein-beta.

IFN-gamma-activated transcriptional element (GATE)-binding factor 1 (GBF1) was identified as a transactivator that induces gene expression through GATE, a novel IFN-inducible element. Although it can induce gene expression, it is an extremely weak DNA-binding protein on its own. GATE also binds another transcription factor, C/EBP-beta. Therefore, we explored whether GBF1 physically interacts with C/EBP-beta to induce IFN-gamma-regulated transcription. In response to IFN-gamma, C/EBP-beta undergoes phosphorylation at a critical ERK1/2 phosphorylation motif. Mutational inactivation of this motif and/or interference with the ERK1/2 activation prevented the IFN-gamma-induced interactions between GBF1 and C/EBP-beta. A 37-aa long peptide derived from the GBF1 protein can associate with C/EBP-beta in an IFN-inducible manner. These results identify a converging point for two transactivators that exert their effects through a single response element. Together, our studies identify a novel regulatory mechanism that controls IFN-induced transcription.

Amino Acid Motifs↗

PubMed2005

Partial truncation of the NH2-terminus affects physical characteristics and membrane binding, aggregation, and fusion properties of annexin A7.

Annexin A7 (synexin, annexin VII), a member of the annexin family of proteins, causes aggregation of membranes in a Ca2+-dependent manner and has been suggested to promote membrane fusion during exocytosis of lung surfactant, catecholamines, and insulin. Although annexin A7 (A7) was one of the first annexin proteins described, limited studies of its physical characteristics or of structural domains affecting any of its proposed functions have been conducted. As postulated for other annexin proteins, the unique NH2-domain possibly determines the functional specificity of A7. Therefore, we evaluated the effects of segmental deletions in the NH2-terminus on several characteristics associated with the COOH-terminus of A7. The COOH-terminus contains the only tryptophan residue, and all potential trypsin sites, and the Ca2+ and phospholipid binding sites. Recombinant rat A7 and its deletion mutants were expressed using constructs based on the cDNA sequence obtained by screening a rat lung cDNA library. Ca2+ increased the tryptophan fluorescence of A7 and caused a small red shift in the emission maximum (lambdamax), which was further increased in presence of phospholipid vesicles (PLV). NH2-terminal deletions of 29, 51, and 109 residues affected the peak width of fluorescence and lambdamax, surface-exposure of tryptophan residue, and caused a smaller Ca2+-dependent red shift in lambdamax of membrane-bound protein in comparison to A7. Limited proteolysis with trypsin showed that Ca2+ increased the proteolysis of all proteins, but the deletions also affected the pattern of proteolysis. The presence of PLV protected against Ca2+-dependent increase in proteolysis of all proteins. The deletion of first 29 residues also caused decreased membrane binding, aggregation, and fusion, when compared with A7. Collectively, these results suggest that specific NH2-terminus domains can alter those properties of A7 that are normally associated with the COOH-terminus. We speculate that interactions between the NH2- and COOH-termini are required for membrane binding, and aggregation and fusion properties of annexin A7.

Amino Acid Sequence↗

PubMed2002

A novel transactivating factor that regulates interferon-gamma-dependent gene expression.

We have previously identified a novel interferon (IFN)-stimulated cis-acting enhancer element, gamma-IFN-activated transcriptional element (GATE). GATE differs from the known IFN-stimulated elements in its primary sequence. Preliminary analysis has indicated that the GATE-dependent transcriptional response requires the binding of novel transacting factors. A cDNA expression library derived from an IFN-gamma-stimulated murine macrophage cell line was screened with a (32)P-labeled GATE probe to identify the potential GATE-binding factors. A cDNA coding for a novel transcription-activating factor was identified. Based on its discovery, we named it as GATE-binding factor-1 (GBF-1). GBF-1 homologs are present in mouse, human, monkey, and Drosophila. It activates transcription from reporter genes carrying GATE. It possesses a strong transactivating activity but has a weak DNA binding property. GBF-1 is expressed in most tissues with relatively higher steady-state levels in heart, liver, kidney, and brain. Its expression is induced by IFN-gamma treatment. GBF-1 is present in both cytosolic and nuclear compartments. These studies thus identify a novel transactivating factor in IFN signaling pathways.

Amino Acid Sequence↗

PubMed2002

Defective NF-kappaB activation in virus-infected neuronal cells is restored by genetic complementation.

The interferon-beta (IFNbeta) gene is not inducible in neuronal cells in response to measles virus (MV) due to lack of nuclear factor kappa B (NF-kappaB) activation. NF-kappaB is normally sequestered in the cytoplasm by an inhibitor (IkappaBalpha). Previously, the authors demonstrated that the failure to activate neuronal NF-kappaB by MV was due to the inability to phosphorylate and degrade its inhibitor, IkappaBalpha. Here the authors demonstrate that transient transfection of a brain cDNA library into neuronal cells restores the ability of MV to activate NF-kappaB. In addition, tumor necrosis factor-alpha (TNFalpha), but not interleukin-1 (IL-1) or lipopolysaccharide (LPS), stimulation resulted in IkappaBalpha phosphorylation and degradation in two neuronal cell lines. These results indicate that failure of MV to activate neuronal NF-kappaB is due to a signaling defect and that MV utilizes an NF-kappaB signaling pathway distinct from that of TNFalpha, but may overlap with that for IL-1 and LPS.

Cell Line↗

PubMed2002

Characterization of monoclonal antibodies against GRIM-19, a novel IFN-beta and retinoic acid-activated regulator of cell death.

A combination of interferon-beta (IFN-beta) and all-trans retinoic acid (IFN/RA) induces tumor cell apoptosis via some unknown mechanisms. Apoptosis is a gene-directed process that limits the proliferation of undesired cells. Several genes are required to regulate cell death in the higher-order animals. Earlier, we employed a gene expression knockout technique to isolate cell death-related genes. A novel gene, the gene associated with retinoid-interferon-induced mortality-19 (GRIM-19), was found to be essential for tumor cell death induced by IFN/RA. Here, we describe the development and characterization of three monoclonal antibodies (mAbs) against GRIM-19. GRIM-19 is present in the nucleus and cytoplasm. Its expression is induced by the IFN/RA combination. We also show that GRIM-19 inhibits the cell-transforming property of viral oncogenic protein viral IFN regulatory factor-1 (vIRF-1) via a physical interaction. mAbs developed in this study should be useful for studying the other physiologic roles of GRIM-19 and serve as a potent tool for studying tumor responses to IFN/RA therapy.

3T3 Cells↗