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Frederick W Alt

Publications and source records attributed to Frederick W Alt.

At least 55 records · Page 3Linked to original sources
PubMed2004

Stress-dependent regulation of FOXO transcription factors by the SIRT1 deacetylase.

The Sir2 deacetylase modulates organismal life-span in various species. However, the molecular mechanisms by which Sir2 increases longevity are largely unknown. We show that in mammalian cells, the Sir2 homolog SIRT1 appears to control the cellular response to stress by regulating the FOXO family of Forkhead transcription factors, a family of proteins that function as sensors of the insulin signaling pathway and as regulators of organismal longevity. SIRT1 and the FOXO transcription factor FOXO3 formed a complex in cells in response to oxidative stress, and SIRT1 deacetylated FOXO3 in vitro and within cells. SIRT1 had a dual effect on FOXO3 function: SIRT1 increased FOXO3's ability to induce cell cycle arrest and resistance to oxidative stress but inhibited FOXO3's ability to induce cell death. Thus, one way in which members of the Sir2 family of proteins may increase organismal longevity is by tipping FOXO-dependent responses away from apoptosis and toward stress resistance.

Acetylation↗
PubMed2004

Unraveling V(D)J recombination; insights into gene regulation.

V(D)J recombination assembles antigen receptor genes from component gene segments. We review findings that have shaped our current understanding of this remarkable mechanism, with a focus on two major reports--the first detailed comparison of germline and rearranged antigen receptor loci and the discovery of the recombination activating gene-1.

Animals↗
PubMed2004

p63 and p73 are not required for the development and p53-dependent apoptosis of T cells.

The recent discoveries of p63 and p73, homologs of the tumor suppressor p53, raised the possibility of a network of these family members governing cell cycle arrest and apoptosis in response to stress. However, mice lacking p73 show no tendency for spontaneous tumors, and mutations in p63 or p73 are rare in human tumors, rendering any obligate role of these genes in cell death and tumor suppression unclear. In an effort to reconcile these incongruent data, we examined the genetic interactions between p53, p63, and p73 in well-established paradigms of p53-dependent and -independent T cell death using primary, genetically defined lymphocytes. Our findings challenge the generality of the notion that p63 and p73 are required for p53 function or for apoptosis in T cells.

Animals↗
PubMed2004

Receptor revision in T cells: an open question?

During lymphocyte development, both B and T cells assemble antigen receptor variable region genes from germline gene segments, allowing the expression of unique receptors in each clonally derived lymphocyte. Previously, it was shown that in certain cases, progenitor and immature B cells are capable of editing their receptors to a new specificity on encounter with self-antigens. Although the existence of such a process in T cells remains controversial, recent studies suggest that mature T cells are able to similarly revise their receptors in the periphery.

Animals↗
PubMed2004

Induction of activation-induced cytidine deaminase gene expression by IL-4 and CD40 ligation is dependent on STAT6 and NFkappaB.

Activation-induced cytidine deaminase (AID) is an inducible gene that plays an important role in class switch recombination, somatic hypermutation and gene conversion in B cells. We examined the regulation of AID gene expression in human and mouse B cells by IL-4 and CD40 ligation. IL-4 by itself and, to a much lesser extent, CD40 ligation induced AID mRNA expression in primary B cells. The two stimuli strongly synergized in inducing AID mRNA and protein expression. IL-4 induced STAT6 binding to a site in the 5' upstream region of the AID gene, while CD40 ligation induced NFkappaB binding to two sites in that region. B cells from STAT6-/- mice failed to up-regulate AID in response to IL-4, while B cells from p50-/- mice were impaired in their ability to up-regulate AID in response to CD40 ligation and IL-4. These results suggest that signals delivered via CD40 that activate NFkappaB synergize with signals delivered via the IL-4 receptor that activate STAT6 to induce optimal AID gene expression.

Animals↗
PubMed2004

The role of the non-homologous end-joining pathway in lymphocyte development.

One of the most toxic insults a cell can incur is a disruption of its linear DNA in the form of a double-strand break (DSB). Left unrepaired, or repaired improperly, these lesions can result in cell death or neoplastic transformation. Despite these dangers, lymphoid cells purposely introduce DSBs into their genome to maximize the diversity and effector functions of their antigen receptor genes. While the generation of breaks requires distinct lymphoid-specific factors, their resolution requires various ubiquitously expressed DNA-repair proteins, known collectively as the non-homologous end-joining pathway. In this review, we discuss the factors that constitute this pathway as well as the evidence of their involvement in two lymphoid-specific DNA recombination events.

Animals↗
PubMed2004

Growth retardation, early death, and DNA repair defects in mice deficient for the nucleotide excision repair enzyme XPF.

Xeroderma pigmentosum (XP) is a human genetic disease which is caused by defects in nucleotide excision repair. Since this repair pathway is responsible for removing UV irradiation-induced damage to DNA, XP patients are hypersensitive to sunlight and are prone to develop skin cancer. Based on the underlying genetic defect, the disease can be divided into the seven complementation groups XPA through XPG. XPF, in association with ERCC1, constitutes a structure-specific endonuclease that makes an incision 5' to the photodamage. XPF-ERCC1 has also been implicated in both removal of interstrand DNA cross-links and homology-mediated recombination and in immunoglobulin class switch recombination (CSR). To study the function of XPF in vivo, we inactivated the XPF gene in mice. XPF-deficient mice showed a severe postnatal growth defect and died approximately 3 weeks after birth. Histological examination revealed that the liver of mutant animals contained abnormal cells with enlarged nuclei. Furthermore, embryonic fibroblasts defective in XPF are hypersensitive to UV irradiation and mitomycin C treatment. No defect in CSR was detected, suggesting that the nuclease is dispensable for this recombination process. These phenotypes are identical to those exhibited by the ERCC1-deficient mice, consistent with the functional association of the two proteins. The complex phenotype suggests that XPF-ERCC1 is involved in multiple DNA repair processes.

Animals↗
PubMed2004

Deficiency in the nuclease activity of xeroderma pigmentosum G in mice leads to hypersensitivity to UV irradiation.

Xeroderma pigmentosum (XP) is a human disorder which is characterized by hypersensitivity to sunlight and elevated incidence of skin cancer. The disease is caused by mutations in genes that encode components of the nucleotide excision repair pathway. The gene product of XP complementation group G (XPG) is a structure-specific endonuclease which makes an incision 3' to DNA photoproducts and other helix-distorting DNA adducts. In addition, the XPG protein has been implicated in transcription and repair of oxidative DNA damage. Moreover, XPG is capable of cleaving R loops in vitro, a potential intermediate during immunoglobulin heavy-chain class switch recombination. Due to its multiple functions, complete elimination of XPG in mice results in severe postnatal growth defects and premature death. To understand the contribution of the XPG nuclease activity to its function in vivo, we introduced a point mutation into the mouse XPG gene which inactivates the nuclease catalytic site but leaves the remainder of the protein intact. The XPG nuclease-deficient animals develop normally and exhibit no obvious defect in class switch recombination. However, the mutant mice are hypersensitive to UV irradiation. This phenotype suggests that the nuclease activity of XPG is required only for nucleotide excision repair and that other regions of the protein perform independent functions.

Animals↗
PubMed2004

H2AX may function as an anchor to hold broken chromosomal DNA ends in close proximity.

The histone H2A variant, H2AX, is a core component of chromatin that is phosphorylated in chromatin flanking DNA double strand breaks (DSBs). Here, we summarize H2AX functions and outline a specific "anchoring" model, that can explain the translocation prone phenotype of H2AX-deficient and H2AX/p53-deficient mice. We also discuss how this model of H2AX function could account for some aspects of the genomic instability and cancer prone human phenotypes associated with Ataxia Telangiectasia (AT), Nijmegen Breakage Syndrome (NBS), Ataxia Telangiectasia Like Disorder (ATLD), and Bloom's Syndrome (BS).

Animals↗
PubMed2003

Vav1/2/3-null mice define an essential role for Vav family proteins in lymphocyte development and activation but a differential requirement in MAPK signaling in T and B cells.

The Vav family of Rho guanine nucleotide exchange factors is thought to orchestrate signaling events downstream of lymphocyte antigen receptors. Elucidation of Vav function has been obscured thus far by the expression of three highly related family members. We generated mice lacking all Vav family proteins and show that Vav-null mice produce no functional T or B cells and completely fail to mount both T-dependent and T-independent humoral responses. Whereas T cell development is blocked at an early stage in the thymus, immature B lineage cells accumulate in the periphery but arrest at a late "transitional" stage. Mechanistically, we show that the Vav family is crucial for both TCR and B cell receptor (BCR)-induced Ca2+ signaling and, surprisingly, is only required for mitogen-activated protein kinase (MAPK) activation in developing and mature T cells but not in B cells. Thus, the abundance of immature B cells generated in Vav-null mice may be due to intact Ras/MAPK signaling in this lineage. Although the expression of Vav1 alone is sufficient for normal lymphocyte development, our data also reveal lineage-specific roles for Vav2 and Vav3, with the first demonstration that Vav3 plays a critical compensatory function in T cells. Together, we define an essential role for the entire Vav protein family in lymphocyte development and activation and establish the limits of functional redundancy both within this family and between Vav and other Rho-guanine nucleotide exchange factors.

Animals↗
PubMed2003

Impaired V(D)J recombination and lymphocyte development in core RAG1-expressing mice.

RAG1 and RAG2 are the lymphocyte-specific components of the V(D)J recombinase. In vitro analyses of RAG function have relied on soluble, highly truncated "core" RAG proteins. To identify potential functions for noncore regions and assess functionality of core RAG1 in vivo, we generated core RAG1 knockin (RAG1(c/c)) mice. Significant B and T cell numbers are generated in RAG1(c/c) mice, showing that core RAG1, despite missing approximately 40% of the RAG1 sequence, retains significant in vivo function. However, lymphocyte development and the overall level of V(D)J recombination are impaired at the progenitor stage in RAG1(c/c) mice. Correspondingly, there are reduced numbers of peripheral RAG1(c/c) B and T lymphocytes. Whereas normal B lymphocytes undergo rearrangement of both JH loci, substantial levels of germline JH loci persist in mature B cells of RAG1(c/c) mice, demonstrating that DJH rearrangement on both IgH alleles is not required for developmental progression to the stage of VH to DJH recombination. Whereas VH to DJH rearrangements occur, albeit at reduced levels, on the nonselected alleles of RAG1(c/c) B cells that have undergone D to JH rearrangements, we do not detect VH to DH rearrangements in RAG1(c/c) B cells that retain germline JH alleles. We discuss the potential implications of these findings for noncore RAG1 functions and for the ordered assembly of VH, DH, and JH segments.

Animals↗
PubMed2003

Developmental defects and p53 hyperacetylation in Sir2 homolog (SIRT1)-deficient mice.

SIRT1 is a mammalian homolog of the Saccharomyces cerevisiae chromatin silencing factor Sir2. Dominant-negative and overexpression studies have implicated a role for SIRT1 in deacetylating the p53 tumor suppressor protein to dampen apoptotic and cellular senescence pathways. To elucidate SIRT1 function in normal cells, we used gene-targeted mutation to generate mice that express either a mutant SIRT1 protein that lacks part of the catalytic domain or has no detectable SIRT1 protein at all. Both types of SIRT1 mutant mice and cells had essentially the same phenotypes. SIRT1 mutant mice were small, and exhibited notable developmental defects of the retina and heart, and only infrequently survived postnatally. Moreover, SIRT1-deficient cells exhibited p53 hyperacetylation after DNA damage and increased ionizing radiation-induced thymocyte apoptosis. In SIRT1-deficient embryonic fibroblasts, however, p53 hyperacetylation after DNA damage was not accompanied by increased p21 protein induction or DNA damage sensitivity. Together, our observations provide direct evidence that endogenous SIRT1 protein regulates p53 acetylation and p53-dependent apoptosis, and show that the function of this enzyme is required for specific developmental processes.

Animals↗
PubMed2003

Histone H2AX: a dosage-dependent suppressor of oncogenic translocations and tumors.

We employed gene targeting to study H2AX, a histone variant phosphorylated in chromatin surrounding DNA double-strand breaks. Mice deficient for both H2AX and p53 (H(delta/delta)P(-/-)) rapidly developed immature T and B lymphomas and solid tumors. Moreover, H2AX haploinsufficiency caused genomic instability in normal cells and, on a p53-deficient background, early onset of various tumors including more mature B lymphomas. Most H2AX(delta/delta)p53(-/-) or H2AX(+/delta)p53(-/-) B lineage lymphomas harbored chromosome 12 (IgH)/15 (c-myc) translocations with hallmarks of either aberrant V(D)J or class switch recombination. In contrast, H2AX(delta/delta)p53(-/-) thymic lymphomas had clonal translocations that did not involve antigen receptor loci and which likely occurred during cellular expansion. Thus, H2AX helps prevent aberrant repair of both programmed and general DNA breakage and, thereby, functions as a dosage-dependent suppressor of genomic instability and tumors in mice. Notably, H2AX maps to a cytogenetic region frequently altered in human cancers, possibly implicating similar functions in man.

Animals↗
PubMed2003

WAVE2 deficiency reveals distinct roles in embryogenesis and Rac-mediated actin-based motility.

The Wiskott-Aldrich syndrome related protein WAVE2 is implicated in the regulation of actin-cytoskeletal reorganization downstream of the small Rho GTPase, Rac. We inactivated the WAVE2 gene by gene-targeted mutation to examine its role in murine development and in actin assembly. WAVE2-deficient embryos survived until approximately embryonic day 12.5 and displayed growth retardation and certain morphological defects, including malformations of the ventricles in the developing brain. WAVE2-deficient embryonic stem cells displayed normal proliferation, whereas WAVE2-deficient embryonic fibroblasts exhibited severe growth defects, as well as defective cell motility in response to PDGF, lamellipodium formation and Rac-mediated actin polymerization. These results imply a non-redundant role for WAVE2 in murine embryogenesis and a critical role for WAVE2 in actin-based processes downstream of Rac that are essential for cell movement.

Actins↗
PubMed2003

Deficiencies of GM-CSF and interferon gamma link inflammation and cancer.

Chronic inflammation contributes to carcinogenesis, but the underlying mechanisms are poorly understood. We report that aged granulocyte-macrophage colony stimulating factor (GM-CSF)-deficient mice develop a systemic lupus erythematosis (SLE)-like disorder associated with the impaired phagocytosis of apoptotic cells. Concurrent deficiency of interferon (IFN)-gamma attenuates the SLE, but promotes the formation of diverse hematologic and solid neoplasms within a background of persistent infection and inflammation. Whereas activated B cells show a resistance to fas-induced apoptosis, antimicrobial therapy prevents lymphomagenesis and solid tumor development. These findings demonstrate that the interplay of infectious agents with cytokine-mediated regulation of immune homeostasis is a critical determinant of cancer susceptibility.

Animals↗