Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UMLS:C0023418 (leukemia)
 93,477 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Among murine leukemia viruses (MuLV) present in the LP-BM5 virus mixture, the agent etiologic for an acquired immunodeficiency syndrome (MAIDS) is replication defective, containing only a single open reading frame which includes all of gag. The Gag polyprotein encoded by the defective virus, termed BM5def, differs most in p12 from that of nonpathogenic ecotropic virus (BM5eco). As one approach to examining the role of p12 in disease, the ecotropic and defective virus forms of the protein, synthesized in bacteria, were used to immunize three strains of mice differing in their sensitivity to MAIDS. In each strain, both proteins elicited substantial antibody responses that were cross-reactive with either p12 and recognized the proteins as part of intact viral Gag polyproteins. Immunization with either p12 before infection with LP-BM5 viruses had no effect on the sensitivity or resistance of mice to MAIDS or on the extent of helper virus spread. The variant p12 of BM5def, when presented on its own, is thus not a crucial antigenic determinant of disease. Alternative mechanisms by which BM5def may contribute to MAIDS are discussed.
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PMID:Effects of immunization with the p12 proteins of LP-BM5 defective and ecotropic viruses on development of MAIDS. 838 12

The retroviral Gag polyprotein is necessary and sufficient for assembly and budding of viral particles. However, the exact inter- and intramolecular interactions of the Gag polyproteins during this process are not known. To locate functional domains within Gag, we generated chimeric proviruses between human immunodeficiency virus type 1 (HIV-1) and murine leukemia virus (MuLV). In these chimeric proviruses, the matrix or capsid proteins of MuLV were precisely replaced with the matrix or capsid proteins of HIV-1. Although the chimeric proviruses were unable to efficiently assemble into mature viral particles by themselves, coexpression of wild-type MuLV Gag rescued the HIV proteins into virions. The specificity of the rescue of HIV proteins into MuLV virions shows that specific interactions involving homologous matrix or capsid regions of Gag are necessary for retroviral particle formation.
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PMID:Incorporation of human immunodeficiency virus type 1 Gag proteins into murine leukemia virus virions. 841 53

For study of the pol gene expression of human T-cell leukemia virus type I (HTLV-I), RNA was transcribed in vitro from proviral DNA and translated in rabbit reticulocyte lysates. This cell-free translation resulted in two major translation products representing the Gag and Gag-Pro polyproteins. By contrast, the Gag-Pro-Pol polyprotein could be readily observed only when translation was performed with mutant mRNA in which the protease (pro) reading frame was aligned to gag to eliminate the frameshifting event in the gag-pro overlap. The results indicated that two independent ribosomal frameshifting events are required for expression of the HTLV-I pol gene product. Studies with mutant DNAs facilitated the characterization of the primary structure of the HTLV-I mRNA responsible for the ribosomal frameshift in the pro-pol overlap and demonstrated that the frameshift occurs at the signal sequence UUUAAAC. Direct amino acid sequencing of the transframe protein localized the site of the frameshift to the asparagine codon AAC.
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PMID:Characterization of ribosomal frameshifting for expression of pol gene products of human T-cell leukemia virus type I. 841 68

The Rex protein of the type I human T-cell leukemia virus (HTLV-I) is essential for viral replication, acting post-transcriptionally to enhance the expression of unspliced and singly spliced viral mRNAs that encode the Gag, Pol, and Env virion proteins. Rex function involves its direct interaction with a complex stem-loop structure termed the Rex RNA response element (RexRE), which is located within the 3' retroviral long terminal repeat. Binding of Rex to the RexRE involves a positively charged arginine-rich domain located near the N-terminus which also functions as a nuclear localization signal. Strikingly, substitution of all seven of the arginine residues present within this domain with positively charged lysine residues exerted no adverse effect on the nuclear targeting of Rex. However, these lysine substitutions completely abrogated both Rex binding to the RexRE and Rex function. Reversion of multiple substituted lysines to arginines at specific locations within this domain was required to restore both RexRE binding and biological function to the Rex protein. Thus, while the presence of positive charge alone in this domain appears sufficient for nuclear localization of Rex, multiple arginine residues at specific sites are essential for the full expression of RNA binding and functional activity of this retroviral trans-regulatory protein.
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PMID:Multiple arginine residues within the basic domain of HTLV-I Rex are required for specific RNA binding and function. 843 77

Murine leukemia viruses (MuLV) have been adapted for use as gene transfer vectors for experimental and human gene therapy applications. Their utility for these purposes has been circumscribed by the limited host range and relatively low titer of available producer clones. Pseudotyping of MuLV particles with the vesicular stomatitis virus envelope protein (VSV-G), expressed transiently in cells producing MuLV Gag and Pol proteins, has yielded vector preparations with a broader host range that can be concentrated by ultracentrifugation. We have explored the use of steroid-inducible and tetracycline-modulated promoter systems (necessary because the VSV-G protein is toxic to cells when constitutively expressed) to derive stable producer cell lines capable of substantial production of VSV-G pseudotyped MuLV particles. A packaging cell line and producer clones capable of expressing a chimeric transcription factor, composed of the tetracycline repressor (tetR) and the VP16 trans-activating sequences of herpes simplex virus VP16 gene and containing the VSV-G coding sequences linked to a minimal promoter having seven tandem copies of the tetracycline responsive operator (tetO), exhibited high levels of VSV-G protein expression when cultured in the absence of tetracycline. Vector particles, produced at titers of 10(5)-10(6) infectious colony forming units per ml (cfu/ml), could be concentrated effectively by ultracentrifugation yielding vector preparations having a titer of 10(9) cfu/ml. These cell lines grew normally when VSV-G protein expression was repressed by tetracycline. Such producer clones hold promise for future human gene therapy applications.
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PMID:Inducible, high-level production of infectious murine leukemia retroviral vector particles pseudotyped with vesicular stomatitis virus G envelope protein. 852 79

The activity of avian sarcoma leukemia virus (ASLV) protease (PR) prior to its release from the precursor protein was determined by introducing mutations at the cleavage site between PR and the adjacent upstream nucleocapsid (NC) protein. Gag DNA fragments containing these mutations were cloned into expression vectors and introduced into Escherichia coli in which the ASLV proteins were expressed. The dipeptide NC-PR containing these mutations did not undergo autoprocessing when expressed in bacterial cells and the fused proteins were devoid of enzymatic activity. However, when the whole Gag polyprotein containing these mutations was expressed in bacterial cells, other PR cleavage sites in the viral Gag polyprotein underwent normal cleavage, indicating that the release of free PR is not a prerequisite for correct processing of the ASLV Gag precursor.
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PMID:Avian sarcoma leukemia virus protease linked to the adjacent Gag polyprotein is enzymatically active. 855 45

The yeast two-hybrid system was used to test for interactions among the Gag precursor proteins of three members of the murine leukemia virus family. These Gag proteins all interact with each other in all combinations, but do not interact with the distantly related HIV-1 Gag. A series of deletion mutants of Moloney MuLV were examined to determine the minimal interaction domain. Either one of two regions of Gag was independently sufficient to mediate homodimerization, suggesting multiple points of contact in the precursor. Analysis of a set of point mutations in the CA region revealed a complex pattern of effects on Gag-Gag interactions.
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PMID:Mutational analysis of interactions between the Gag precursor proteins of murine leukemia viruses. 860 72

Successful retroviral-mediated gene therapy will depend on safe, efficient packaging cell lines for vector particle production. Existing packaging lines for murine leukemia virus (MLV)-based vectors are predominantly derived from NIH/3T3 cells which carry endogenous MLV sequences that could participate in recombination to form replication-competent retrovirus (RCR). To identify cells devoid of such sequences, we screened genomic DNA from eight cell lines. DNA from the human 293 cell line did not cross-hybridize with MLV sequences, and these cells were able to secrete Gag particles after transfection. We derived a stable amphotropic packaging cell line (called ProPak-A) in 293 cells in which the Gag-Pol and Env (packaging) functions are expressed separately from a heterologous (non-MLV) promoter, to maximally reduce homology between packaging and vector sequences. ProPak-A-based producer cells are efficient, yielding higher stable titers than PA317-based producers. In addition, a vector that consistently gave rise to RCR in PA317 cells never resulted in detectable RCR in ProPak-A-based producer cultures. We have also shown that ProPak-A-packaged particles are not inactivated by human serum. Thus, the packaging cells we describe are as efficient and safer than the amphotropic packaging cells most commonly used in clinical gene therapy work and are also more appropriate for in vivo gene delivery.
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PMID:A novel human amphotropic packaging cell line: high titer, complement resistance, and improved safety. 861 38

A necessary downstream element of Abelson murine leukemia virus (Ab-MLV)-mediated transformation is Ras, which can be activated by the phosphotyrosine-dependent association of Shc with the Grb2-mSos complex. Here we show that Shc is tyrosine-phosphorylated and associates with Grb2 in v-Abl-transformed cells, whereas Shc in NIH3T3 cells is phosphorylated solely on serine and is not Grb2-associated. In addition, Shc coprecipitates with P120 v-Abl and P70 v-Abl, which lacks the carboxyl terminus. Surprisingly, a kinase-defective mutant of P120 also binds Shc, demonstrating that Shc/v-Abl association is a phosphotyrosine-independent interaction. Glutathione S-transferase fusion proteins were used to map the interacting domains and showed that Shc from both NIH3T3 and v-Abl-transformed cells binds to the Abl SH2 domain and that P120 v-Abl binds to a region in the amino terminus of Shc. Consistent with these data, a v-Abl mutant encoding only the Gag and SH2 regions was able to bind Shc in vivo. The unique non-phosphotyrosine-mediated binding of Shc may allow direct tyrosine phosphorylation of Shc by v-Abl and subsequent activation of the Ras pathway through assembly of a signaling complex with Grb2-mSos.
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PMID:In vivo association of v-Abl with Shc mediated by a non-phosphotyrosine-dependent SH2 interaction. 861 26

The Gag proteins of replication-competent retroviruses direct budding at the plasma membrane and are cleaved by the viral protease (PR) just before or very soon after particle release. In contrast, defective retroviruses that bud into the endoplasmic reticulum (ER) have been found, and morphologically these appear to contain uncleaved Gag proteins. From this, it has been proposed that activation of PR may depend upon a host factor found only at the plasma membrane. However, if Gag proteins were cleaved by PR before the particle could pinch off the ER membrane, then the only particles that would remain visible are those that packaged smaller-than-normal amounts of PR, and these would have an immature morphology. To distinguish between these two hypotheses, we made use of the Rous sarcoma virus (RSV) Gag protein, the PR of RSV IS included on each Gag molecule. To target Gag to the ER, a signal peptide was installed at its amino terminus in place of the plasma membrane-binding domain. An intervening, hydrophobic, transmembrane anchor was included to keep Gag extended into the cytoplasm. We found that PR-mediated processing occurred, although the cleavage products were rapidly degraded. When the anchor was removed, allowing the entire protein to be inserted into the lumen of the ER, Gag processing occurred with a high level of efficiency, and the cleavage products were quite stable. Thus, PR activation does not require targeting of Gag molecules to the plasma membrane. Unexpectedly, molecules lacking the transmembrane anchor were rapidly secreted from the cell in a nonmembrane-enclosed form and in a manner that was very sensitive to brefeldin A and monensin. In contrast, the wild-type RSV and Moloney murine leukemia virus Gag proteins were completely insensitive to these inhibitors, suggesting that the normal mechanism of transport to the plasma membrane does not require interactions with the secretory pathway.
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PMID:Transport and processing of the Rous sarcoma virus Gag protein in the endoplasmic reticulum. 862 76


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