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D Baltimore

Publications and source records attributed to D Baltimore.

At least 19 recordsLinked to original sources
PubMed1992

Secondary structure of Src homology 2 domain of c-Abl by heteronuclear NMR spectroscopy in solution.

The Src homology 2 (SH2) domain is a recognition motif thought to mediate the association of the cytoplasmic proteins involved in signal transduction by binding to phosphotyrosyl-containing sequences in proteins. Assignments of nearly all 1H and 15N resonances of the SH2 domain from the c-Abl protein-tyrosine kinase have been obtained from homonuclear and heteronuclear NMR experiments. The secondary structure has been elucidated from the pattern of nuclear Overhauser effects, from vicinal coupling constants, and from observation of slowly exchanging amino hydrogens. The secondary structure contains two alpha-helices and eight beta-strands, six of which are arranged in two contiguous, antiparallel beta-sheets. Residues believed to be involved in phosphotyrosyl ligand binding are on a face of one beta-sheet. The alignment of homologous sequences on the basis of secondary structure suggests a conserved global fold in a family of SH2 domains.

Amino Acid Sequence↗
PubMed1992

Blocked early-stage latency in the peripheral blood cells of certain individuals infected with human immunodeficiency virus type 1.

Human immunodeficiency virus type 1 (HIV-1) infections of humans have a natural history characterized by a variable but usually slow progression to an immunodeficient state. We have described a molecular model of HIV-1 proviral latency in certain cell lines, characterized by extremely low or undetectable levels of unspliced genomic HIV-1-specific RNA but significant levels of multiply spliced HIV-1-specific RNA. We have utilized a quantitative reverse transcriptase-initiated polymerase chain reaction to measure the levels of various HIV-1 RNA species in peripheral blood mononuclear cells. The median level of multiply spliced HIV-1 RNA was dramatically higher than the median level of unspliced viral RNA in asymptomatic individuals. In addition, HIV-1 RNA patterns characterized by at least a 10-fold excess of multiply spliced to unspliced viral RNA were significantly more common in asymptomatic individuals than in patients with the acquired immunodeficiency syndrome. We suggest that asymptomatic clinical HIV-1 infection is characterized by a preponderance of HIV-1-infected peripheral blood cells blocked at an early stage of HIV-1 infection. This viral expression pattern, which we have called blocked early-stage latency, may constitute a reservoir of latently infected cells in certain HIV-1-infected persons.

HIV Seropositivity↗
PubMed1992

Crystal structure of the phosphotyrosine recognition domain SH2 of v-src complexed with tyrosine-phosphorylated peptides.

Three-dimensional structures of complexes of the SH2 domain of the v-src oncogene product with two phosphotyrosyl peptides have been determined by X-ray crystallography at resolutions of 1.5 and 2.0 A, respectively. A central antiparallel beta-sheet in the structure is flanked by two alpha-helices, with peptide binding mediated by the sheet, intervening loops and one of the helices. The specific recognition of phosphotyrosine involves amino-aromatic interactions between lysine and arginine side chains and the ring system in addition to hydrogen-bonding interactions with the phosphate.

Amino Acid Sequence↗
PubMed1992

Identification of a protein that binds to the SH3 region of Abl and is similar to Bcr and GAP-rho.

A Src homology 3 (SH3) region is a sequence of approximately 50 amino acids found in many nonreceptor tyrosine kinases and other proteins. Deletion of the SH3 region from the protein encoded by the c-abl proto-oncogene activates the protein's transforming capacity, thereby suggesting the participation of the SH3 region in the negative regulation of transformation. A complementary DNA was isolated that encoded a protein, 3BP-1, to which the SH3 region of Abl bound with high specificity and to which SH3 regions from other proteins bound differentially. The sequence of the 3BP-1 protein is similar to that of a COOH-terminal segment of Bcr and to guanosine triphosphatase-activating protein (GAP)-rho, which suggests that it might have GAP activity for Ras-related proteins. The 3BP-1 protein may therefore be a mediator of SH3 function in transformation inhibition and may link tyrosine kinases to Ras-related proteins.

3T3 Cells↗
PubMed1992

Different molecular consequences of the 1;19 chromosomal translocation in childhood B-cell precursor acute lymphoblastic leukemia.

The prognostically important 1;19 chromosomal translocation can alter the E2A gene on chromosome 19p13 in childhood B-cell precursor acute lymphoblastic leukemia (ALL), leading to formation of a fusion gene (E2A-PBX1) that encodes a hybrid transcription factor with oncogenic potential. It is not known whether this molecular alteration is a uniform consequence of the t(1;19) or is restricted to translocation events within specific immunologic subtypes of the disease. Therefore, we studied leukemic cells from 25 cases of B-cell precursor ALL, with or without evidence of cytoplasmic Ig mu heavy chains (cIg); 17 cases had the t(1;19) by cytogenetic analysis. Leukemic cell DNA samples were analyzed by Southern blotting to detect alterations within the E2A genomic locus; a polymerase chain reaction assay was used to identify expression of chimeric E2A-pbx1 transcripts in leukemic cell RNA; and immunoblotting with anti-Pbx1 antibodies was used to detect hybrid E2A-Pbx1 proteins. Of 11 cases of cIg+ ALL with the t(1;19), 10 had E2A-pbx1 chimeric transcripts with identical junctions and a characteristic set of E2A-Pbx1 hybrid proteins. Each of these cases had E2A gene rearrangements, including the one in which fusion transcripts were not detected. By contrast, none of the six cases of t(1;19)-positive, cIg- ALL had evidence of rearranged E2A genomic restriction fragments, detectable E2A-pbx1 chimeric transcripts, or hybrid E2A-Pbx1 proteins. Typical chimeric E2A-pbx1 transcripts and proteins were detected in one of eight cIg+ leukemias in which the t(1;19) was not identified by cytogenetic analysis, emphasizing the increased sensitivity of molecular analysis for detection of this abnormality. We conclude that the molecular breakpoints in cases of cIg- B-cell precursor ALL with the t(1;19) differ from those in cIg+ cases with this translocation. Leukemias that express hybrid oncoproteins such as E2A-Pbx1 or Bcr-Abl have had a poor prognosis in most studies. Thus, molecular techniques to detect fusion genes and their aberrant products should allow more timely and appropriate treatment of these aggressive subtypes of the disease.

Adolescent↗
PubMed1992

Cellular latency of human immunodeficiency virus type 1.

The infection of humans by human immunodeficiency virus type 1 is characterized by a prolonged stage of clinical quiescence. This clinically asymptomatic period may be based, in part, on the development of cell populations within the body that maintain human immunodeficiency virus type 1 in a state of latency. Recent advances in the understanding of the molecular mechanisms involved in various forms of cellular latency of human immunodeficiency virus type 1 have begun to shed light on the variable period of asymptomatic infection. The elucidation of cellular retroviral latency, in vivo, will also be critical to the design of novel therapeutic approaches with which to combat human immunodeficiency virus type 1 infections.

DNA Replication↗
PubMed1992

The inhibitory ankyrin and activator Rel proteins.

The gene families encoding the proteins NF-kappa B, c-Rel and Dorsal, in conjunction with their respective inhibitors l kappa B, pp40, and Cactus, achieve specificity in gene regulation by means of common principles. The related activities of NF-kappa B and Dorsal are mediated by heterodimeric or homodimeric complexes of proteins containing the conserved dimerization and DNA-binding domain termed Rel. The l kappa Bs and Cactus, which share a core series of structural repeats termed ankyrin, inhibit cognate activators through differential interactions with the Rel-homology domain. Together, the inhibitory ankyrin proteins and their cognate Rel dimers probably define specific signalling pathways able to activate specific gene expression. Both gene families include proto-oncogenes, thus broadly implicating Rel/l kappa B in the control of both normal gene expression and the aberrant gene expression that makes cells cancerous.

Amino Acid Sequence↗
PubMed1992

Point mutations in the abl SH2 domain coordinately impair phosphotyrosine binding in vitro and transforming activity in vivo.

We have constructed a series of point mutations in the highly conserved FLVRES motif of the src homology 2 (SH2) domain of the abl tyrosine kinase. Mutant SH2 domains were expressed in bacteria, and their ability to bind to tyrosine-phosphorylated proteins was examined in vitro. Three mutants were greatly reduced in their ability to bind both phosphotyrosine itself and tyrosine-phosphorylated cellular proteins. All of the mutants that retained activity bound to the same set of tyrosine-phosphorylated proteins as did the wild type, suggesting that binding specificity was unaffected. These results implicate the FLVRES motif in direct binding to phosphotyrosine. When the mutant SH2 domains were inserted into an activated abl kinase and expressed in murine fibroblasts, decreased in vitro phosphotyrosine binding correlated with decreased transforming ability. This finding implies that SH2-phosphotyrosine interactions are involved in transmission of positive growth signals by the nonreceptor tyrosine kinases, most likely via the assembly of multiprotein complexes with other tyrosine-phosphorylated proteins.

Amino Acid Sequence↗
PubMed1979

Synthetic phospholipid vesicles containing a purified viral antigen and cell membrane proteins stimulate the development of cytotoxic T lymphocytes.

Synthetic phospholipid vesicles (liposomes) containing the purified glycoprotein (G) of vesicular stomatitis virus (VSV) and solubilized membrane proteins from cells of the appropriate H-2 haplotype elicited H-2-restricted cytotoxic T lymphocytes (CTL) that lysed VSV-infected target cells. The CTL were elicited by intact liposomes, not by released components. Thus, when spleen cells from VSV-primed H-2d X H-2b hybrid mice were stimulated with liposomes having G protein + membrane proteins from cells with one of the parental H-2 haplotypes, the resulting CTL lysed only VSV-infected target cells with that parent's H-2 type. This result argues against the view that T cells in general recognize only processed antigenic fragments on macrophages. Moreover, liposomes were only effective when G protein and cell membrane proteins were included in the same vesicles. This result suggests that for effective interaction with CTL precursors the antigen (G protein) and products of the H-2 complex must be closer to each other than 600--1,000 angstrom, the diameter of the lipid vesicles used in this study.

Animals↗
PubMed1979

Immunoglobulin synthesis by lymphoid cells transformed in vitro by Abelson murine leukemia virus.

The majority of cell lines derived by infection of murine bone marrow cells with Abelson murine leukemia virus (A-MuLV) synthesize a mu chain but no detectable light chain. Aside from this mu-only phenotype, lines that make only light chain, both chains or no immunoglobulin-related polypeptides have also been found. Two lines have been studied in detail: one that makes only mu chain and one that makes only kappa light chain. Synthesis of both polypeptides can be increased by modifying the culture conditions so as to decrease the growth rate of the cells. Although some kappa chain secretion was observed, neither secreted nor surface mu was detected. We suggest that the mu- only phenotype may be an early normal step in the pathway of B lymphocyte maturation.

Animals↗
PubMed1979

In vitro synthesis of a 9 kbp terminally redundant DNA carrying the infectivity of Moloney murine leukemia virus.

Detergent-disrupted virions of Moloney murine leukemia virus synthesize a 9 kbp double-stranded infectious DNA. It contains mainly full-length, single-stranded DNA, and its infectivity and size are insensitive to digestion by the single-strand-specific S1 nuclease. Analysis of fragmentation of the DNA using restriction endonucleases has shown that it is indistinguishable from the linear double-stranded DNA synthesized in infected cells. On the basis of the positions of the cleavage sites for a number of enzymes, the 9 kbp DNA has a 575 base direct terminal repetition. It is longer than the viral RNA at both ends, evidently due to repetitive copying of segments of the RNA. Virions also synthesize an 8.4 kbp double-stranded circular DNA that lacks one copy of the terminal repetition, as well as viral DNA longer than 9 kbp. The enzymatic machinery in the virions of retroviruses therefore appears to be responsible for all the steps involved in making fully double-stranded linear and one form of circular DNA.

Base Sequence↗
PubMed1979

Structure of the Abelson murine leukemia virus genome.

Virions produced from cells transformed by A-MuLV contain a 30S, 5.6 kb RNA that can be translated in a cell-free system to form the characteristic A-MuLV protein. This RNA was mapped by heteroduplex methods using DNA probes from M-MuLV, the presumed parent of A-MuLV. The overall organization of the RNA was determined by using full-length M-MuLV reverse transcribed DNA and visualizing the heteroduplexes in the electron microscope. This showed that A-MuLV and M-MuLV have homologous sequences at both ends of their RNAs but that the central portion of the A-MuLV genome is not homologous to sequences in M-MuLV RNA. A precise measure of the lengths of the shared regions was obtained by using S1 nuclease to digest hybrids between 32P-labeled M-MuLV DNA and A-MuLV RNA; the resulting fragments were analyzed for their length by electrophoresis. The regions of homology were shown to be 1320 nucleotides long at the 5' end and 730 nucleotides long at the 3' end. Thus approximately 6200 nucleotides of the approximately 8300 in M-MuLV RNA were deleted when the A-MuLV genome was formed, but an insert of 3600 nucleotides, presumably derived from the normal murine genome, was inserted in place of the deleted region.

Abelson murine leukemia virus↗
PubMed1979

A detailed model of reverse transcription and tests of crucial aspects.

A model of reverse transcription has been devised by which the detailed architecture of ten molecular structures is predicted. The model includes a number of novel features for which experimental evidence is presented. First, growing minus DNA strand is copied from the viral RNA only up to a position about 150 nucleotides from the 5' end of the RNA. Second, plus-strand DNA, after being copied from approximately 600 nucleotides at the 5' end of the minus-strand DNA, then transcribes the first approximately 20 nucleotides of the tRNApro primer (which is covalently attaced to the 5' end of the minus DNA strand). The 3' ends of the minus and plus DNA probably form a hybrid through the homology conferred by the primer binding site sequences. Third, the minus and plus DNA strands are elongated in a continuous fashion resulting in a linear double-stranded DNA molecule containing a 600 nucleotide direct repeat at both ends. The most of the features of the model have experimental support, and it appears to provide a credible description of reverse transcription.

DNA, Viral↗
PubMed1979

Poliovirus replicase: a soluble enzyme able to initiate copying of poliovirus RNA.

The soluble phase of the cytoplasm of poliovirus-infected cells contains an enzymatic activity able to copy RNA without an added primer. This replicase activity has been purified 60-fold; it is absent from uninfected cells. Poly(U) polymerase activity copurifies with replicase activity. Although less pure replicase fractions copy a variety of RNAs, purer fractions respond better to poliovirus RNA than to other viral RNAs. Even the less pure fractions make a specific copy of the added template, as shown by hybridization of the product to its template RNA but not to other RNAs. Among homopolymers only poly(A)-oligo(U) was copied by the replicase; other primed homopolymer templates were inactive.

HeLa Cells↗
PubMed1979

Synthesis of a 600-nucleotide-long plus-strand DNA by virions of Moloney murine leukemia virus.

A discrete, 600-nucleotide-long plus-strand DNA has been identified among the products of reverse transcription by virions of Moloney murine leukemia virus. Its polarity was shown by hybridization to minus-strand DNA. It appears to be copied from the right end of minus-strand DNA because (i) its restriction endonuclease cleavage pattern corresponds to the redundant 600-base segment found at either end of the ultimate double-stranded reverse transcription products, (ii) its synthesis is actinomycin D sensitive, and (iii) its synthesis begins during the first hour of a reverse transcription reaction when only the right-hand end of minus-strand DNA is available as template. We therefore call this DNA plus-strong-stop DNA by analogy with the minus-strong-stop DNA copied from the left end of the viral RNA. Both strong-stop DNAs are made early during in vitro reactions and decline in concentration later, consistent with postulated roles as initiators of long minus- and plus-strand DNA. Unlike minus-strong-stop DNA, plus-strong-stop DNA remains as a double-stranded nucleic acid after its synthesis, as shown by S1 nuclease resistance. A primer to initiate plus-strong-stop DNA synthesis has not been identified; the product found thus far has no detectable RNA attached to it.

Chromosome Mapping↗