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Biomedical subjects

D Baltimore

Publications and source records attributed to D Baltimore.

At least 37 records · Page 2Linked to original sources
PubMed1999

Dissecting Fas signaling with an altered-specificity death-domain mutant: requirement of FADD binding for apoptosis but not Jun N-terminal kinase activation.

Fas is a cell surface death receptor that regulates peripheral tolerance and lymphoid homeostasis. In many pathologic conditions, ectopic Fas activation mediates tissue destruction. Several proteins that can bind to the cytoplasmic death domain of Fas have been implicated in Fas signal transduction. Here we show that FADD, which couples Fas to pro-caspase-8, and, Daxx, which couples Fas to the Jun N-terminal kinase pathway, bind independently to the Fas death domain. We have isolated a death domain mutant, termed FasDelta, that selectively binds Daxx but not FADD. In tranfected tissue culture cells, FasDelta activated Jun N-terminal kinase normally but was impaired in cell death induction. These results suggest that FADD and Daxx activate two independent pathways downstream of Fas and confirm the essential role of FADD binding in apoptosis induction.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Specificities of CD40 signaling: involvement of TRAF2 in CD40-induced NF-kappaB activation and intercellular adhesion molecule-1 up-regulation.

Several tumor necrosis factor receptor-associated factor (TRAF) family proteins including TRAF2, TRAF3, TRAF5, and TRAF6, as well as Jak3, have been implicated as potential mediators of CD40 signaling. An extensive in vitro binding study indicated that TRAF2 and TRAF3 bind to the CD40 cytoplasmic tail (CD40ct) with much higher affinity than TRAF5 and TRAF6 and that TRAF2 and TRAF3 bind to different residues of the CD40ct. Using CD40 mutants incapable of binding TRAF2, TRAF3, or Jak3, we found that the TRAF2-binding site of the CD40ct is critical for NF-kappaB and stress-activated protein kinase activation, as well as the up-regulation of the intercellular adhesion molecule-1 (ICAM-1) gene, whereas binding of TRAF3 and Jak3 is dispensable for all of these functions. Overexpression of a dominantly active IkappaBalpha strongly inhibited CD40-induced NF-kappaB activation, ICAM-1 promoter activity, and cell-surface ICAM-1 up-regulation. These studies suggest a potential signal transduction pathway from the CD40 receptor to the transcriptional activation of the ICAM-1 gene.

Amino Acid Substitution↗
PubMed1999

The p65 subunit of NF-kappa B is redundant with p50 during B cell proliferative responses, and is required for germline CH transcription and class switching to IgG3.

B cells lacking individual NF-kappa B/Rel family members exhibit defects in activation programs. We generated small resting B cells lacking p65 or p50 alone, or lacking both p50 and p65, then evaluated the ability of these cells to proliferate, secrete Ig, and undergo Ig class switching. B cells lacking p65 proliferated well in response to all stimuli tested. However, these cells demonstrated an isolated defect in switching to IgG3, which was associated with a decrease in gamma 3 germline CH gene expression. Whereas, previously reported, B cells lacking p50 alone had a severe proliferative defect in response to LPS, a moderate defect in response to CD40 ligand (CD40L), and normal proliferation to Ag receptor cross-linking using dextran-conjugated anti-IgD Abs (alpha delta-dex), B cells lacking both p50 and p65 exhibited severely impaired proliferation in response to LPS, alpha delta-dex, and CD40L. This defect could be overcome by simultaneous administration of alpha delta-dex and CD40L. In response to this latter combination of stimuli, B cells lacking both p50 and p65 secreted Ig and underwent isotype switching to IgG1 as efficiently as B cells lacking p50 alone. These data demonstrate a role for the p65 subunit of NF-kappa B in germline CH gene expression as well as functional redundancy between p50 and p65 during proliferative responses.

Animals↗
PubMed1999

The selective downregulation of class I major histocompatibility complex proteins by HIV-1 protects HIV-infected cells from NK cells.

To avoid detection by CTL, HIV encodes mechanisms for removal of class I MHC proteins from the surface of infected cells. However, class I downregulation potentially exposes the virus-infected cell to attack by NK cells. Human lymphoid cells are protected from NK cell cytotoxicity primarily by HLA-C and HLA-E. We present evidence that HIV-1 selectively downregulates HLA-A and HLA-B but does not significantly affect HLA-C or HLA-E. We then identify the residues in HLA-C and HLA-E that protect them from HIV down-regulation. This selective downregulation allows HIV-infected cells to avoid NK cell-mediated lysis and may represent for HIV a balance between escape from CTL and maintenance of protection from NK cells. These results suggest that subpopulations of CTL and NK cells may be uniquely suited for combating HIV.

Cells, Cultured↗
PubMed1999

Uncoupling IL-2 signals that regulate T cell proliferation, survival, and Fas-mediated activation-induced cell death.

IL-2 is an important growth and survival factor for T lymphocytes but also sensitizes these cells to Fas-mediated activation-induced cell death (AICD). The molecular basis of these different effects of IL-2 was studied by introducing wild-type and mutant forms of the IL-2 receptor beta (IL-2Rbeta) chain that lacked specific signaling capacities into receptor-deficient T cells by retroviral gene transfer. Activation of Stat5 by IL-2 was found to be involved in T cell proliferation and promoted Fas ligand (FasL) expression and AICD. T cell survival was dependent on a receptor region that activated Akt and the expression of Bcl-2. Thus, distinct IL-2Rbeta chain signaling modules regulate T cell fate by stimulating growth and survival or by promoting apoptosis.

Animals↗
PubMed1999

Autoimmunity as a consequence of retrovirus-mediated expression of C-FLIP in lymphocytes.

The induction of apoptosis by death receptors serves to regulate immune responses by eliminating unwanted and harmful cells. Mature lymphocytes express FLICE inhibitory proteins (FLIPs) that block death receptor-induced cell death. Here, we show that both B and T cells downregulate c-FLIP upon activation in vitro. Retrovirus-mediated expression of c-FLIP blocks Fas-induced apoptosis of activated lymphocytes but does not affect cell death resulting from cytokine withdrawal. In vivo, c-FLIP expression results in defective superantigen-mediated elimination of T cells, the accumulation of activated B cells, the production of autoantibodies, and the development of autoimmune disease. No effect was seen on negative selection of thyomocytes. These results suggest that activation-dependent downregulation of c-FLIP renders mature lymphocytes sensitive to death receptor-mediated apoptosis and is required to maintain self-tolerance.

Adaptor Proteins, Signal Transducing↗
PubMed1999

HIV's evasion of the cellular immune response.

Despite a strong cytotoxic T-lymphocyte (CTL) response directed against viral antigens, untreated individuals infected with the human immunodeficiency virus (HIV-1) develop AIDS. We have found that primary T cells infected with HIV-1 downregulate surface MHC class I antigens and are resistant to lysis by HLA-A2-restricted CTL clones. In contrast, cells infected with an HIV-1 in which the nef gene is disrupted are sensitive to CTLs in an MHC and peptide-specific manner. In primary T cells HLA-A2 antigens are downmodulated more dramatically than total MHC class I antigens, suggesting that nef selectively downmodulates certain MHC class I antigens. In support of this, studies on cells expressing individual MHC class I alleles have revealed that nef does not downmodulate HLA-C and HLA-E antigens. This selective downmodulation allows infected cells to maintain resistance to certain natural killer cells that lyse infected cells expressing low levels of MHC class I antigens. Downmodulation of MHC class I HLA-A2 antigens occurs not only in primary T cells, but also in B and astrocytoma cell lines. No effect of other HIV-1 accessory proteins such as vpu and vpr was observed. Thus Nef is a protein that may promote escape of HIV-1 from immune surveillance.

Animals↗
PubMed1998

Efficient and rapid induction of a chronic myelogenous leukemia-like myeloproliferative disease in mice receiving P210 bcr/abl-transduced bone marrow.

Expression of the 210-kD bcr/abl fusion oncoprotein can cause a chronic myelogenous leukemia (CML)-like disease in mice receiving bone marrow cells transduced by bcr/abl-encoding retroviruses. However, previous methods failed to yield this disease at a frequency sufficient enough to allow for its use in the study of CML pathogenesis. To overcome this limitation, we have developed an efficient and reproducible method for inducing a CML-like disease in mice receiving P210 bcr/abl-transduced bone marrow cells. All mice receiving P210 bcr/abl-transduced bone marrow cells succumb to a myeloproliferative disease between 3 and 5 weeks after bone marrow transplantation. The myeloproliferative disease recapitulates many of the hallmarks of human CML and is characterized by high white blood cell counts and extensive extramedullary hematopoiesis in the spleen, liver, bone marrow, and lungs. Use of a retroviral vector coexpressing P210 bcr/abl and green fluorescent protein shows that the vast majority of bcr/abl-expressing cells are myeloid. Analysis of the proviral integration pattern shows that, in some mice, the myeloproliferative disease is clonal. In multiple mice, the CML-like disease has been transplantable, inducing a similar myeloproliferative syndrome within 1 month of transfer to sublethally irradiated syngeneic recipients. The disease in many of these mice has progressed to the development of acute lymphoma/leukemia resembling blast crisis. These results demonstrate that murine CML recapitulates important features of human CML. As such, it should be an excellent model for addressing specific issues relating to the pathogenesis and treatment of this disease.

Animals↗
PubMed1998

Activation of apoptosis signal-regulating kinase 1 (ASK1) by the adapter protein Daxx.

The Fas death receptor can activate the Jun NH2-terminal kinase (JNK) pathway through the receptor-associated protein Daxx. Daxx was found to activate the JNK kinase kinase ASK1, and overexpression of a kinase-deficient ASK1 mutant inhibited Fas- and Daxx-induced apoptosis and JNK activation. Fas activation induced Daxx to interact with ASK1, which consequently relieved an inhibitory intramolecular interaction between the amino- and carboxyl-termini of ASK1, activating its kinase activity. The Daxx-ASK1 connection completes a signaling pathway from a cell surface death receptor to kinase cascades that modulate nuclear transcription factors.

Adaptor Proteins, Signal Transducing↗
PubMed1998

Essential role of CED-4 oligomerization in CED-3 activation and apoptosis.

Control of the activation of apoptosis is important both in development and in protection against cancer. In the classic genetic model Caenorhabditis elegans, the pro-apoptotic protein CED-4 activates the CED-3 caspase and is inhibited by the Bcl-2-like protein CED-9. Both processes are mediated by protein-protein interaction. Facilitating the proximity of CED-3 zymogen molecules was found to induce caspase activation and cell death. CED-4 protein oligomerized in cells and in vitro. This oligomerization induced CED-3 proximity and competed with CED-4:CED-9 interaction. Mutations that abolished CED-4 oligomerization inactivated its ability to activate CED-3. Thus, the mechanism of control is that CED-3 in CED-3:CED-4 complexes is activated by CED-4 oligomerization, which is inhibited by binding of CED-9 to CED-4.

Apoptosis↗
PubMed1998

HIV-1 directly kills CD4+ T cells by a Fas-independent mechanism.

The mechanism by which HIV-1 induces CD4(+) T cell death is not known. A fundamental issue is whether HIV-1 primarily induces direct killing of infected cells or indirectly causes death of uninfected bystander cells. This question was studied using a reporter virus system in which infected cells are marked with the cell surface protein placental alkaline phosphatase (PLAP). Infection by HIV-PLAP of peripheral blood mononuclear cells (PBMCs) and T cell lines leads to rapid depletion of CD4(+) T cells and induction of apoptosis. The great majority of HIV-induced T cell death in vitro involves direct loss of infected cells rather than indirect effects on uninfected bystander cells. Because of its proposed role in HIV-induced cell death, we also examined the Fas (CD95/Apo1) pathway in killing of T cells by HIV-1. Infected PBMCs or CEM cells display no increase in surface Fas relative to uninfected cells. In addition, HIV-1 kills CEM and Jurkat T cells in the presence of a caspase inhibitor that completely blocks Fas-mediated apoptosis. HIV-1 also depletes CD4+ T cells in PBMCs from patients who have a genetically defective Fas pathway. These results suggest that HIV-1 induces direct apoptosis of infected cells and kills T cells by a Fas-independent mechanism.

Alkaline Phosphatase↗
PubMed1998

HIV-1 Nef protein protects infected primary cells against killing by cytotoxic T lymphocytes.

Cytotoxic T lymphocytes (CTLs) lyse virally infected cells that display viral peptide epitopes in association with major histocompatibility complex (MHC) class I molecules on the cell surface. However, despite a strong CTL response directed against viral epitopes, untreated people infected with the human immunodeficiency virus (HIV-1) develop AIDS. To resolve this enigma, we have examined the ability of CTLs to recognize and kill infected primary T lymphocytes. We found that CTLs inefficiently lysed primary cells infected with HIV-1 if the viral nef gene product was expressed. Resistance of infected cells to CTL killing correlated with nef-mediated downregulation of MHC class I and could be overcome by adding an excess of the relevant HIV-1 epitope as soluble peptide. Thus, Nef protected infected cells by reducing the epitope density on their surface. This effect of nef may allow evasion of CTL lysis by HIV-1-infected cells.

Alkaline Phosphatase↗
PubMed1998

Essential roles for the Abl and Arg tyrosine kinases in neurulation.

The Abl and Arg tyrosine kinases play fundamental roles in the development and function of the central nervous system. Arg is most abundant in adult mouse brain, especially in synapse-rich regions. arg(-/-) mice develop normally but exhibit multiple behavioral abnormalities, suggesting that arg(-/-) brains suffer from defects in neuronal function. Embryos deficient in both Abl and Arg suffer from defects in neurulation and die before 11 days postcoitum (dpc). Although they divide normally, abl(-/-)arg(-/-) neuroepithelial cells display gross alterations in their actin cytoskeleton. We find that Abl and Arg colocalize with each other and with actin microfilaments at the apical surface of the developing neuroepithelium. Thus, Abl and Arg play essential roles in neurulation and can regulate the structure of the actin cytoskeleton.

Actins↗
PubMed1998

Autoproteolytic activation of pro-caspases by oligomerization.

Initiation of apopotosis requires the conversion of procaspases to mature caspases. Here we show that oligomerization of pro-caspases is sufficient to induce proteolytic generation of mature caspase subunits and activation of their cell death activity. Deletion of the protein interaction motif DED from pro-caspase-8 greatly suppresses its apoptotic activity. Cell death activity can be restored by oligomerization of pro-caspase-8 protease domains by two heterologous inducible oligomerization systems. Induced oligomerization also activates the apoptotic activity of pro-caspase-1 but not pro-caspase-3. In vitro, oligomerization leads to pro-caspase processing to from the mature caspase subunits; this processing requires the intrinsic caspase activity of zymogens and proceeds via a novel order of cleavage events.

Apoptosis↗
PubMed1998

Involvement of p53 and p21 in cellular defects and tumorigenesis in Atm-/- mice.

Disruption of the mouse Atm gene, whose human counterpart is consistently mutated in ataxia-telangiectasia (A-T) patients, creates an A-T mouse model exhibiting most of the A-T-related systematic and cellular defects. While ATM plays a major role in signaling the p53 response to DNA strand break damage, Atm-/- p53(-/-) mice develop lymphomas earlier than Atm-/- or p53(-/-) mice, indicating that mutations in these two genes lead to synergy in tumorigenesis. The cell cycle G1/S checkpoint is abolished in Atm-/- p53(-/-) mouse embryonic fibroblasts (MEFs) following gamma-irradiation, suggesting that the partial G1 cell cycle arrest in Atm-/- cells following gamma-irradiation is due to the residual p53 response in these cells. In addition, the Atm-/- p21(-/-) MEFs are more severely defective in their cell cycle G1 arrest following gamma-irradiation than Atm-/- and p21(-/-) MEFs. The Atm-/- MEFs exhibit multiple cellular proliferative defects in culture, and an increased constitutive level of p21 in these cells might account for these cellular proliferation defects. Consistent with this notion, Atm-/- p21(-/-) MEFs proliferate similarly to wild-type MEFs and exhibit no premature senescence. These cellular proliferative defects are also rescued in Atm-/- p53(-/-) MEFs and little p21 can be detected in these cells, indicating that the abnormal p21 protein level in Atm-/- cells is also p53 dependent and leads to the cellular proliferative defects in these cells. However, the p21 mRNA level in Atm-/- MEFs is lower than that in Atm+/+ MEFs, suggesting that the higher level of constitutive p21 protein in Atm-/- MEFs is likely due to increased stability of the p21 protein.

Animals↗
PubMed1997

Degeneration of neurons, synapses, and neuropil and glial activation in a murine Atm knockout model of ataxia-telangiectasia.

Neural degeneration is one of the clinical manifestations of ataxia-telangiectasia, a disorder caused by mutations in the Atm protein kinase gene. However, neural degeneration was not detected with general purpose light microscopic methods in previous studies using several different lines of mice with disrupted Atm genes. Here, we show electron microscopic evidence of degeneration of several different types of neurons in the cerebellar cortex of 2-month-old Atm knockout mice, which is accompanied by glial activation, deterioration of neuropil structure, and both pre- and postsynaptic degeneration. These findings are similar to those in patients with ataxia-telangiectasia, indicating that Atm knockout mice are a useful model to elucidate the mechanisms underlying neurodegeneration in this condition and to develop and test strategies to palliate and prevent the disease.

Animals↗