Biomedical subjects

A L Hammond

Publications and source records attributed to A L Hammond.

9 recordsLinked to original sources

PubMed2001

Measles virus envelope glycoproteins hetero-oligomerize in the endoplasmic reticulum.

The endoplasmic reticulum (ER) was investigated as the initial oligomerization site for the envelope glycoproteins H and F of measles virus (MV), a clinically relevant member of the Paramyxoviridae family, and consequences of this interaction for viral replication were studied. Both proteins were tagged at their cytosolic tails with RRR and KKXX motifs, respectively, resulting in their efficient retention in the ER. Co-transfection of the retained constructs with transport competent MV glycoproteins revealed a dominant negative effect on their biological activity indicating intracellular complex formation and thus retention. Pulse-chase analysis and co-immunoprecipitation experiments demonstrated that this effect is based on both homo- and hetero-oligomerization in the ER. Recombinant viruses additionally expressing ER-retained F showed an altered cytopathic phenotype accompanied by greatly reduced particle release. Similar mutant viruses additionally expressing ER-retained H could not be rescued indicating an even greater negative effect of this protein on virus viability. Our study suggests that both homo- and hetero-oligomerization of MV glycoproteins occur in the ER and that these events are of significance for early steps of particle assembly.

Animals↗

PubMed2001

Antigenic variation within the CD4 binding site of human immunodeficiency virus type 1 gp120: effects on chemokine receptor utilization.

To assess the antigenicity of envelope glycoproteins derived from primary human immunodeficiency virus type 1 populations, their interactions with the receptor CD4, and their coreceptor usage, we have cloned and expressed multiple gp120 proteins from a number of primary virus isolates. Characterization of these proteins showed a high degree of antigenic polymorphism both within the CD4 binding site and in defined neutralization epitopes, which may partially account for the general resistance of primary isolates to neutralizing agents. Furthermore, chimeric viruses expressing gp120 proteins with reduced CD4 binding abilities are viable, suggesting that primary viruses may require a less avid interaction with the receptor CD4 to initiate infection than do their laboratory-adapted counterparts. The coreceptor usage of chimeric viruses was related to the ability of the virus to bind CD4, with reduced CD4 binding correlating with preferential usage of CXCR4. Changes in coreceptor usage mapped to sequence changes in the C2 and V4 regions, with no changes seen in the V3 region.

Amino Acid Sequence↗

PubMed2001

Single-chain antibody displayed on a recombinant measles virus confers entry through the tumor-associated carcinoembryonic antigen.

To redirect the tropism of the vaccine strain of measles virus (MV), Edmonston B, to a targeted cell population, we displayed on the viral hemagglutinin (H) a single-chain antibody (scAb) specific for the tumor-associated carcinoembryonic antigen (CEA). We generated H fusion proteins with three forms of the scAb appended, differing in the lengths of the linkers separating the VH and VL domains and thus in the oligomerization states of the scAbs. All proteins were stable, appeared properly folded, and were transported to the cell surface, but only H displaying the long-linker form of scAb was functional in supporting cell-cell fusion. This protein induced extensive syncytia in cells expressing the normal virus receptor CD46 and also in CD46-negative cells expressing the targeted receptor, human CEA. Replication-competent MV with H replaced by H displaying the long-linker form of scAb was recovered and replicated efficiently in both CD46-positive and CD46-negative, CEA-positive cells. Thus, MV not only tolerates the addition of a scAb on its H protein but also infects cells via a novel interaction between the scAb and its targeted receptor.

Animals↗

PubMed2000

Characterization of a region of the measles virus hemagglutinin sufficient for its dimerization.

Attachment of measles virus (MV) to its cellular receptor is mediated by the viral envelope glycoprotein hemagglutinin (H). H exists at the viral surface as a disulfide-linked dimer which may associate into a tetramer. We aimed to define regions of H essential for its homo-oligomerization. To delineate these more precisely, we have generated a series of H ectodomain truncation mutants and studied their abilities to form both homotypic complexes and heterotypic complexes with full-length H. We define a "minimal unit" which is sufficient for MV H dimerization as that encompassing residues 1 to 151. This unit forms both homodimers and heterodimers with full-length H protein, although neither is transported to the cell surface even in the presence of other MV proteins. We show that cysteine residues at positions 139 and 154 are both critical in mediating covalent dimerization, not only of the truncated H mutants but also of full-length MV H protein. Even those cysteine mutants unable to form covalent intermolecular interactions are biologically active, mediating the formation of syncytia, albeit at a reduced rate. We demonstrate that this impaired capacity to mediate cell-to-cell fusion is based mainly on a reduced transport rate of the mutant molecules to the cell surface, indicating a role for covalent intermolecular interactions in efficient transport of MV H dimers to the cell surface.

Animals↗

PubMed1973