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Biomedical subjects

R M Bedgood

Publications and source records attributed to R M Bedgood.

6 recordsLinked to original sources
PubMed1994

Abnormal processing of a recombinant feline leukemia virus envelope polyprotein and its interference with subgroup C virus infection.

Processing of the env polyprotein of a noninfectious feline leukemia virus (FeLV) recombinant, named r6gp, was examined in human-transfected cells. The r6gp provirus was previously generated in the frame of FeLV, subgroup B, GA clone with substitution of all but 40 C-terminal amino acid sequences of the surface glycoprotein (SU) from an endogenous FeLV provirus element (CFE-6). Although r6gp produced a normal size (85 kDa) env glycoprotein precursor, the product, unlike the precursor of the parental virus, was neither additionally glycosylated nor further processed into mature env proteins. Biochemical observations were consistent with the idea that the chimeric env polyprotein was trapped in the endoplasmic reticulum (ER) and were directly supported by immunofluorescence microscopy analyses. Interestingly, the residence of the chimeric protein in the ER specifically interfered with FeLV, subgroup C (Sarma) virus infection but not the parental FeLV-B virus infection. Since FeLV-C provirus sequences could be readily detected in the infected cells, it appeared that r6gp env expression did not block entry of the challenge virus. While FeLV-B and CFE-6 env genes share an extensive overall sequence homology, a variable region (region VI) of CFE-6 near the C-terminus of SU, which was retained in the r6gp construct, exhibits a considerably higher degree of homology to FeLV-C than FeLV-B. Thus, we propose that region VI is involved in conferring specificity for the env polyprotein oligomerization in the ER, and that co-oligomerization of the trapped r6gp env with FeLV-C is the reason for specific interference with FeLV-C infection. The results also demonstrate for the first time a functional abnormality of a recombinant FeLV env gene which is structurally similar to those commonly detected in FeLV-induced feline lymphosarcomas.

Amino Acid Sequence↗
PubMed1992

A novel intermediate in processing of murine leukemia virus envelope glycoproteins. Proteolytic cleavage in the late Golgi region.

The intracellular processing of the murine leukemia virus envelope glycoprotein precursor Pr85 to the mature products gp70 and p15e was analyzed in the mouse T-lymphoma cell line W7MG1. Kinetic (pulse-chase) analysis of synthesis and processing, coupled with endoglycosidase (endo H) and neuraminidase digestions revealed the existence of a novel high molecular weight processing intermediate, gp95, containing endo H-resistant terminally glycosylated oligosaccharide chains. In contrast to previously published conclusions, our data indicate that proteolytic cleavage of the envelope precursor occurs after the acquisition of endo H-resistant chains and terminal glycosylation and thus after the mannosidase II step. In the same W7MG1 cell line, the type and order of murine leukemia virus envelope protein processing events was identical to that for the mouse mammary tumor virus envelope protein. Interestingly, complete mouse mammary tumor virus envelope protein processing requires the addition of glucocorticoid hormone, whereas murine leukemia virus envelope protein processing occurs constitutively in these W7MG1 cells. We propose that all retroviral envelope proteins share a common processing pathway in which proteolytic processing is a late event that follows acquisition of endo H resistance and terminal glycosylation.

Electrophoresis, Polyacrylamide Gel↗
PubMed1992

Maturation of mouse mammary tumor virus envelope protein is blocked by a specific point mutation.

Expression of a mouse mammary tumor virus (MMTV) envelope gene, ENV2, in COS-7 cells resulted in the synthesis of a precursor protein Pr74 that underwent complete proteolytic processing to the mature envelope species gp52 and gp33. In contrast, expression of a second MMTV envelope gene, ENV1, in COS-7 cells resulted in the synthesis of a precursor protein Pr74 that failed to undergo proteolytic processing to the mature products. Expression and sequencing of various chimeric ENV1/ENV2 genes determined that a single glycine-to-glutamic acid mutation at position 54 was responsible for the nonprocessable phenotype of ENV1-encoded Pr74. The significance of this point mutation in relation to the requirements for precursor protein folding and maturation are discussed.

Animals↗
PubMed1992

Two different genes coding for processable and nonprocessable forms of a viral envelope protein can account for the apparent hormonal stimulation of protein processing in W7MG1 lymphoma cells.

In the mouse mammary tumor virus (MMTV)-infected mouse T-lymphoma cell line W7MG1, glucocorticoid hormone regulates two aspects of MMTV gene expression: hormone stimulates MMTV gene transcription and increases the ratio of mature envelope proteins to envelope precursor protein produced. To separate these two effects and determine the mechanism by which hormone regulates the conversion of the envelope precursor Pr74 to the mature cleaved products gp52 and gp33, we constructed expression vectors in which the envelope gene is constitutively transcribed. Surprisingly, the envelope precursor protein Pr74 encoded by two independently isolated, allelic envelope genes behaved differently. Pr74-P (encoded by the ENV/P gene) was processed efficiently to the mature products gp52 and gp33, independently of the level of expression, hormonal induction of cellular genes, or the presence of other MMTV proteins. In contrast, under the same conditions, Pr74-N (encoded by the ENV/N gene) was not processed further despite being relatively stable. In sucrose gradient analyses, Pr74-P sedimented as monomers, whereas Pr74-N was found in high mol wt aggregates of heterogeneous size. Coimmunoprecipitation analysis determined that Pr74-N associated with BiP, whereas Pr74-P did not. This is indicative of improper folding of Pr74-N in the endoplasmic reticulum.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗