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Biomedical subjects

David Baltimore

Publications and source records attributed to David Baltimore.

31 records · Page 2Linked to original sources
PubMed2002

CARD11 mediates factor-specific activation of NF-kappaB by the T cell receptor complex.

NF-kappaB is a critical target of signaling downstream of the T cell receptor (TCR) complex, but how TCR signaling activates NF-kappaB is poorly understood. We have developed an expression cloning strategy that can identify catalytic and noncatalytic molecules that participate in different pathways of NF-kappaB activation. Screening of a mouse thymus cDNA library yielded CARD11, a membrane-associated guanylate kinase (MAGUK) family member containing CARD, PDZ, SH3 and GUK domains. Using a CARD-deleted variant of CARD11 and RNA interference (RNAi), we demonstrate that CARD11 mediates NF-kappaB activation by alphaCD3/alphaCD28 cross-linking and PMA/ionomycin treatment, but not by TNFalpha or dsRNA. CARD11 is not required for TCR-mediated induction of NFAT or AP-1. CARD11 functions upstream of the IkappaB-kinase (IKK) complex and cooperates with Bcl10 in a CARD domain-dependent manner. RNAi-rescue experiments suggest that the CARD, coiled-coil, SH3 and GUK domains of CARD11 are critical for its signaling function. These results implicate CARD11 in factor- specific activation of NF-kappaB by the TCR complex and establish a role for a MAGUK family member in antigen receptor signaling.

Amino Acid Sequence↗
PubMed2002

Inhibition of DNA binding by NF-kappa B with pyrrole-imidazole polyamides.

Synthetic ligands that bind to predetermined DNA sequences will offer a chemical approach to gene regulation if inhibition of a broad range of transcription factors can be achieved. NF-kappa B is a transcription factor that regulates a multitude of genes, including those involved in immune, inflammatory, and anti-apoptotic responses. NF-kappa B binds as heterodimer predominantly in the major groove. We report the design of polyamides that bind in the minor groove and target overlapping portions of an NF-kappa B binding site (5'-GGGACTTTCC-3'). We find that compounds that target the 5'-GGGACT-3' portion of the site can inhibit DNA binding by NF-kappa B while those that target the 5'-ACTTTCC-3' portion do not. Addition of NF-kappa B to the list of protein-DNA complexes that can be disrupted by minor groove binding ligands potentially increases the utility of polyamides as regulators of gene expression.

Base Sequence↗
PubMed2002

Generation of functional antigen-specific T cells in defined genetic backgrounds by retrovirus-mediated expression of TCR cDNAs in hematopoietic precursor cells.

We have developed an alternative to transgenesis for producing antigen-specific T cells in vivo. In this system, clonal naive T cells with defined antigen specificity are generated by retrovirus-mediated expression of T cell antigen receptor cDNAs in RAG1-deficient murine hematopoietic precursor cells. These T cells can be stimulated to proliferate and produce cytokines by exposure to antigen in vitro, and they become activated and expand in vivo after immunization. IL-2-deficient T cells generated by this technique show decreased proliferation and cytokine production, both of which can be rescued by exogenous addition of this growth factor. Thus, retrovirus-mediated expression of T cell antigen receptor cDNAs in hematopoietic precursor cells permits the rapid and efficient analysis of the life history of antigen-specific T cells in different genetic backgrounds and may allow for the long-term production of antigen-specific T cells with different functional properties for prophylactic and therapeutic purposes.

Animals↗
PubMed2002

Essential roles of the kappa light chain intronic enhancer and 3' enhancer in kappa rearrangement and demethylation.

The kappa intronic (MiE(kappa)) and 3' (3'E(kappa)) enhancers are both quantitatively important to, but not essential for, immunoglobulin kappa rearrangement. To determine the functional redundancy between these two enhancers, B cells derived from mutant embryonic stem cells--in which both MiE(kappa) and 3'E(kappa) were deleted on both kappa alleles--were analyzed for kappa rearrangement. Our findings indicate that these double-mutant B cells have essentially no kappa rearrangement but do rearrange and express lambda. Therefore, these two kappa enhancers share essential roles in activating V(kappa)J(kappa) rearrangement. Our findings also indicate that the two kappa enhancers play overlapping and distinct roles in the demethylation of kappa in B cells.

3' Flanking Region↗
PubMed2002

The combined absence of NF-kappa B1 and c-Rel reveals that overlapping roles for these transcription factors in the B cell lineage are restricted to the activation and function of mature cells.

Transcription factors NF-kappaB1 and c-Rel, individually dispensable during embryogenesis, serve similar, yet distinct, roles in the function of mature hemopoietic cells. Redundancy among Rel/NF-kappaB family members prompted an examination of the combined roles of c-Rel and NF-kappaB1 by using mice that lack both proteins. Embryonic development and the maturation of hemopoietic progenitors were unaffected in nfkb1(-/-)c-rel(-/-) mice. Peripheral T cell populations developed normally, but follicular, marginal zone, and CD5(+) peritoneal B cell populations all were reduced. In culture, a failure of mitogen-stimulated nfkb1(-/-)c-rel(-/-) B cells to proliferate was caused by a cell cycle defect in early G(1) that prevented growth. In vivo, defects in humoral immunity and splenic architecture seen in nfkb1(-/-) and c-rel(-/-) mice were exacerbated in the double mutant mice. These findings demonstrate that in the B lineage overlapping roles for NF-kappaB1 and c-Rel appear to be restricted to regulating the activation and function of mature cells.

Animals↗
PubMed2002

Cooperation of multiple signaling pathways in CD40-regulated gene expression in B lymphocytes.

CD40/CD40L interaction is essential for multiple biological events in T dependent humoral immune responses, including B cell survival and proliferation, germinal center and memory B cell formation, and antibody isotype switching and affinity maturation. By using high-density microarrays, we examined gene expression in primary mouse B lymphocytes after multiple time points of CD40L stimulation. In addition to genes involved in cell survival and growth, which are also induced by other mitogens such as lipopolysaccharide, CD40L specifically activated genes involved in germinal center formation and T cell costimulatory molecules that facilitate T dependent humoral immunity. Next, by examining the roles of individual CD40-activated signal transduction pathways, we dissected the overall CD40-mediated response into genes independently regulated by the individual pathways or collectively by all pathways. We also found that gene down-regulation is a significant part of the overall response and that the p38 pathway plays an important role in this process, whereas the NF-kappa B pathway is important for the up-regulation of primary response genes. Our finding of overlapping independent control of gene expression modules by different pathways suggests, in principle, that distinct biological behaviors that depend on distinct gene expression subsets can be manipulated by targeting specific signaling pathways.

Animals↗
PubMed2002

Requirement for the NF-kappaB family member RelA in the development of secondary lymphoid organs.

The transcription factor nuclear factor (NF)-kappaB has been suggested to be a key mediator of the development of lymph nodes and Peyer's patches. However, targeted deletion of NF-kappaB/ Rel family members has not yet corroborated such a function. Here we report that when mice lacking the RelA subunit of NF-kappaB are brought to term by breeding onto a tumor necrosis factor receptor (TNFR)1-deficient background, the mice that are born lack lymph nodes, Peyer's patches, and an organized splenic microarchitecture, and have a profound defect in T cell-dependent antigen responses. Analyses of TNFR1/RelA-deficient embryonic tissues and of radiation chimeras suggest that the dependence on RelA is manifest not in hematopoietic cells but rather in radioresistant stromal cells needed for the development of secondary lymphoid organs.

Animals↗
PubMed2002

Germline transmission and tissue-specific expression of transgenes delivered by lentiviral vectors.

Single-cell mouse embryos were infected in vitro with recombinant lentiviral vectors to generate transgenic mice carrying the green fluorescent protein (GFP) gene driven by a ubiquitously expressing promoter. Eighty percent of founder mice carried at least one copy of the transgene, and 90% of these expressed GFP at high levels. Progeny inherited the transgene(s) and displayed green fluorescence. Mice generated using lentiviral vectors with muscle-specific and T lymphocyte-specific promoters expressed high levels of GFP only in the appropriate cell types. We have also generated transgenic rats that express GFP at high levels, suggesting that this technique can be used to produce other transgenic animal species.

Animals↗
PubMed2002

Two pathways to NF-kappaB.

NF-kappaB is a transcription factor that is critical for innate and adaptive immunity. Recently, a noncanonical pathway for NF-kappaB activation has emerged. Four recent papers provide physiological roles for this pathway and expand our understanding of lymphoid development and organogenesis with potential applications in the treatment of autoimmune disease.

Animals↗