Search PubMed⌕ Search

Biomedical subjects

I M Verma

Publications and source records attributed to I M Verma.

At least 37 records · Page 2Linked to original sources
PubMed2000

Complete lack of NF-kappaB activity in IKK1 and IKK2 double-deficient mice: additional defect in neurulation.

NF-kappaB activity is induced by cytokines, stress, and pathogens. IKK1 and IKK2 are critical IkappaB kinases in NF-kappaB activation. In this study mice lacking IKK1 and IKK2 died at E12. Additional defect in neurulation associated with enhanced apoptosis in the neuroepithelium was also observed. MEF cells from IKK1(-/-)/IKK2(-/-) embryos did not respond to NF-kappaB inducers. Upon crossing with kappaB-lacZ transgenic mice, double-deficient embryos also lost lacZ transgene expression in vascular endothelial cells during development. Our data suggest that IKK1 and IKK2 are essential for NF-kappaB activation in vivo and have an important role in protecting neurons against excessive apoptosis during development.

Alleles↗
PubMed2000

Signal-dependent and -independent degradation of free and NF-kappa B-bound IkappaBalpha.

A family of inhibitory IkappaB molecules regulates the activation of the transcription factor NF-kappaB. One member of the IkappaB family, IkappaBalpha, plays a major role in the rapid signal-induced activation of NF-kappaB. IkappaBalpha itself is transcriptionally regulated by NF-kappaB allowing for a tight autoregulatory loop that is both sensitive to and rapidly influenced by NF-kappaB activating stimuli. For this pathway to remain primed both for rapid activation of NF-kappaB in the presence of signal and then to suppress NF-kappaB activation once that signal is removed, IkappaBalpha must be exquisitely regulated. The regulation of IkappaBalpha is mainly accomplished through phosphorylation, ubiquitination, and subsequent degradation. The mechanism(s) that regulate IkappaBalpha degradation needs to be able to target IkappaBalpha for degradation in both its NF-kappaB bound and free states in the cell. In this study, we utilize a full-length IkappaBalpha mutant that is unable to associate to RelA/p65. We show that the signal-induced IkappaB kinase (IKK) phosphorylation sites on IkappaBalpha can only significantly influence the regulation of signal-dependent but not signal-independent turnover of IkappaBalpha. We also demonstrate that the constitutive carboxyl-terminal casein kinase II phosphorylation sites are necessary for the proper regulation of both signal-dependent and -independent turnover of IkappaBalpha. These findings further elucidate how the phosphorylation of IkappaBalpha influences the complex regulatory mechanisms involved in maintaining a sensitive NF-kappaB pathway.

Animals↗
PubMed2000

CBP/p300 interact with and function as transcriptional coactivators of BRCA1.

BRCA1 is a breast and ovarian cancer-specific tumor suppressor, with properties of a transcription factor involved in DNA repair. We previously have shown the transactivation of heterologous promoters by the carboxyl terminus of BRCA1. We now describe that BRCA1-mediated transactivation is enhanced by p300/CBP (CREB binding protein) and that this effect was suppressed by the adenovirus E1A oncoprotein. We show a physical association of BRCA1 with the transcriptional coactivators/acetyltransferases p300 and CBP. Endogenous as well as overexpressed BRCA1 and p300 were found to associate in a phosphorylation-independent manner. BRCA1 interacts with the cAMP response element binding protein (CREB) domain of p300/CBP via both its amino and carboxyl termini. Finally, full-length BRCA1 is shown to transcriptionally activate the Rous sarcoma virus-long terminal repeat promoter, which was further stimulated by p300. Immunocolocalization analyses suggest that BRCA1 and p300 associate in a cell cycle-dependent manner. Our results support a role for BRCA1 in transcription.

Avian Sarcoma Viruses↗
PubMed2000

Reciprocal role of ERK and NF-kappaB pathways in survival and activation of osteoclasts.

To examine the role of mitogen-activated protein kinase and nuclear factor kappa B (NF-kappaB) pathways on osteoclast survival and activation, we constructed adenovirus vectors carrying various mutants of signaling molecules: dominant negative Ras (Ras(DN)), constitutively active MEK1 (MEK(CA)), dominant negative IkappaB kinase 2 (IKK(DN)), and constitutively active IKK2 (IKK(CA)). Inhibiting ERK activity by Ras(DN) overexpression rapidly induced the apoptosis of osteoclast-like cells (OCLs) formed in vitro, whereas ERK activation after the introduction of MEK(CA) remarkably lengthened their survival by preventing spontaneous apoptosis. Neither inhibition nor activation of ERK affected the bone-resorbing activity of OCLs. Inhibition of NF-kappaB pathway with IKK(DN) virus suppressed the pit-forming activity of OCLs and NF-kappaB activation by IKK(CA) expression upregulated it without affecting their survival. Interleukin 1alpha (IL-1alpha) strongly induced ERK activation as well as NF-kappaB activation. Ras(DN) virus partially inhibited ERK activation, and OCL survival promoted by IL-1alpha. Inhibiting NF-kappaB activation by IKK(DN) virus significantly suppressed the pit-forming activity enhanced by IL-1alpha. These results indicate that ERK and NF-kappaB regulate different aspects of osteoclast activation: ERK is responsible for osteoclast survival, whereas NF-kappaB regulates osteoclast activation for bone resorption.

Adenoviridae↗
PubMed2000

Sustained expression of therapeutic level of factor IX in hemophilia B dogs by AAV-mediated gene therapy in liver.

We demonstrate that a single intraportal vein injection of a recombinant adeno-associated virus (rAAV) vector encoding canine factor IX (cFIX) cDNA under the control of a liver-specific enhancer/promoter leads to a long-term correction of the bleeding disorder in hemophilia B dogs. Stable expression of the therapeutic level of cFIX (5% of normal level) was detected in the plasma of a dog injected with an AAV vector at a dose of 4.6 x 10(12) particles/kg for over 7 months. Both whole-blood clotting time (WBCT) and activated partial thromboplastin time (aPTT) of the treated dogs have been greatly decreased since the treatment. No anti-canine factor IX antibodies have been detected in the treated animals. Importantly, no bleeding has been observed in the dog that expresses a therapeutic level of cFIX for 7 months following vector administration. Moreover, no persistent significant hepatic enzyme abnormalities were detected in the treated dogs. Thus, a single intraportal injection of a rAAV vector expressing cFIX successfully corrected the bleeding disorder of hemophilia B dogs, supporting the feasibility of using AAV-based vectors for liver-targeted gene therapy of genetic diseases.

Alanine Transaminase↗
PubMed2000

Lentiviral vectors: regulated gene expression.

Lentiviral vectors can deliver and express genes in a wide variety of dividing and nondividing cells. These include terminally differentiated neurons, myotubes, hepatocytes, and hematopoietic stem cells. We now describe the generation of lentiviral vectors in which the expression of the transgene can be regulated. We have developed an inducible lentiviral vector system that contains the entire tetracycline (Tet)-regulated system developed by H. Bujard and colleagues. The novel vector expresses the GFP reporter gene and the tetracycline transactivator under the control of the tetracycline-inducible promoter and the human CMV promoter, respectively. In vitro transduction of human 293 cells resulted in a very low basal expression of GFP in the presence of the effector substance doxycyline. Withdrawal of doxycyline induced a more than 500-fold increase in transgene expression. Switching transgene expression "off and on" did not change either the kinetics or the magnitude of induction. Maximal suppression of GFP mRNA transcription was achieved within 24 h of addition of the drug; however, due to the slow turnover rate of GFP, green fluorescent cells could be detected up to 10 days following doxycyline treatment. Following transduction of rat brain with recombinant lentiviruses, doxycyline-regulated GFP expression could be observed in terminally differentiated neurons. Specifically, by adding or withdrawing doxycyline from the rats' drinking water, induction and suppression of GFP expression could be regulated in vivo. These studies show that an inducible lentiviral vector can deliver and regulate transgene expression in vivo. We believe that regulated gene expression is an essential tool for successful gene therapy approaches.

Animals↗
PubMed2000

Gene therapy: trials and tribulations.

The art and science of gene therapy has received much attention of late. The tragic death of 18-year-old Jesse Gelsinger, a volunteer in a Phase I clinical trial, has overshadowed the successful treatment of three children suffering from a rare but fatal immunological disease. In the light of the success and tragedy, it is timely to consider the challenges faced by gene therapy--a novel form of molecular medicine that may be poised to have an important impact on human health in the new millennium.

Adolescent↗
PubMed1999

Truncated BRCA2 is cytoplasmic: implications for cancer-linked mutations.

BRCA2 mutations predispose carriers mainly to breast cancer. The vast majority of BRCA2 mutations are predicted to result in a truncated protein product. The smallest known cancer-associated deletion removes from the C terminus only 224 of the 3,418 residues constituting BRCA2, suggesting that these terminal amino acids are crucial for BRCA2 function. A series of green fluorescent protein (GFP)-tagged BRCA2 deletion mutants revealed that nuclear localization depends on two nuclear localization signals that reside within the final 156 residues of BRCA2. Consistent with this observation, an endogenous truncated BRCA2 mutant (6174delT) was found to be cytoplasmic. Together, these studies provide a simple explanation for why the vast majority of BRCA2 mutants are nonfunctional: they do not translocate into the nucleus.

BRCA2 Protein↗
PubMed1999

LFG: an anti-apoptotic gene that provides protection from Fas-mediated cell death.

Programmed cell death regulates a number of biological phenomena, and the apoptotic signal must itself be tightly controlled to avoid inappropriate cell death. We established a genetic screen to search for molecules that inhibit the apoptotic signal from the Fas receptor. Here we report the isolation of a gene, LFG, that protects cells uniquely from Fas but not from the mechanistically related tumor necrosis factor alpha death signal. LFG is widely distributed, but remarkably is highly expressed in the hippocampus. LFG can bind to the Fas receptor, but does not regulate Fas expression or interfere with binding of an agonist antibody. Furthermore LFG does not inhibit binding of FADD to Fas.

Amino Acid Sequence↗
PubMed1999

IKK1-deficient mice exhibit abnormal development of skin and skeleton.

IkappaB kinases (IKKs) IKK1 and IKK2 are two putative IkappaBalpha kinases involved in NF-kappaB activation. To examine the in vivo functions of IKK1, we generated IKK1-deficient mice. The mutant mice are perinatally lethal and exhibit a wide range of developmental defects. Newborn mutant mice have shiny, taut, and sticky skin without whiskers. Histological analysis shows thicker epidermis, which is unable to differentiate. Limbs and tail are wrapped inside the skin and do not extend properly out of the body trunk. Skeleton staining reveals a cleft secondary palate, split sternebra 6, and deformed incisors. NF-kappaB activation mediated by TNFalpha and IL-1 is diminished in IKK1-deficient mouse embryonic fibroblast (MEF) cells. The IKK complex in the absence of IKK1 is capable of phosphorylating IkappaBalpha and IkappaBbeta in vitro. Our results support a role for IKK1 in NF-kappaB activation and uncover its involvement in skin and skeleton development. We conclude further that the two related kinases IKK1 and IKK2 have distinct functions and can not be substituted for each other's functions.

Animals↗