Dataset Information

Role for Gag-CA Interdomain Linker in Primate Lentiviral Replication.

ABSTRACT: Gag proteins underlie retroviral replication by fulfilling numerous functional roles at various stages during viral life cycle. Out of the four mature proteins, Gag-capsid (CA) is a major component of viral particles, and has been most well studied biogenetically, biochemically and structurally. Gag-CA is composed of two structured domains, and also of a short stretch of disordered and flexible interdomain linker. While the two domains, namely, N-terminal and C-terminal domains (NTD and CTD), have been the central target for Gag research, the linker region connecting the two has been poorly studied. We recently have performed systemic mutational analyses on the Gag-CA linker region of HIV-1 by various experimental and in silico systems. In total, we have demonstrated that the linker region acts as a cis-modulator to optimize the Gag-related viral replication process. We also have noted, during the course of conducting the research project, that HIV-1 and SIVmac, belonging to distinct primate lentiviral lineages, share a similarly biologically active linker region with each other. In this brief article, we summarize and report the results obtained by mutational studies that are relevant to the functional significance of the interdomain linker of HIV/SIV Gag-CA. Based on this investigation, we discuss about the future directions of the research in this line.

SUBMITTER: Doi N

PROVIDER: S-EPMC6694209 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Role for Gag-CA Interdomain Linker in Primate Lentiviral Replication.

Doi Naoya N Koma Takaaki T Adachi Akio A Nomaguchi Masako M

Frontiers in microbiology 20190807

Gag proteins underlie retroviral replication by fulfilling numerous functional roles at various stages during viral life cycle. Out of the four mature proteins, Gag-capsid (CA) is a major component of viral particles, and has been most well studied biogenetically, biochemically and structurally. Gag-CA is composed of two structured domains, and also of a short stretch of disordered and flexible interdomain linker. While the two domains, namely, N-terminal and C-terminal domains (NTD and CTD), ha ...[more]

PMID: 31440231

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Project description:The retroviral Gag capsid (Gag-CA) interdomain linker is an unstructured peptide segment connecting structured N-terminal and C-terminal domains. Although the region is reported to play roles in virion morphogenesis and infectivity, underlying molecular mechanisms remain unexplored. To address this issue, we determined biological and molecular phenotypes of HIV-1 CA linker mutants by experimental and in silico approaches. Among the nine linker mutants tested, eight exhibited attenuation of viral particle production to various extents mostly in parallel with a reduction in viral infectivity. Sucrose density gradient, confocal microscopy, and live-cell protein interaction analyses indicated that the defect is accompanied by attenuation of Gag-Gag interactions following Gag plasma membrane targeting in the cells. In silico analyses revealed distinct distributions of interaction-prone hydrophobic patches between immature and mature CA proteins. Molecular dynamics simulations predicted that the linker mutations can allosterically alter structural fluctuations, including the interaction surfaces apart from the mutation sites in both the immature and mature CA proteins. These results suggest that the HIV-1 CA interdomain linker is a cis-modulator of the CA interaction surfaces to optimize efficiency of Gag assembly, virion production, and viral infectivity.IMPORTANCE HIV-1 particle production and infection are highly ordered processes. Viral Gag proteins play a central role in the assembly and disassembly of viral molecules. Of these, capsid protein (CA) is a major contributor to the Gag-Gag interactions. CA consists of two structured domains, i.e., N-terminal (NTD) and C-terminal (CTD) domains, connected by an unstructured domain named the interdomain linker. While multiple regions in the NTD and CTD are reported to play roles in virion morphogenesis and infectivity, the roles of the linker region in Gag assembly and virus particle formation remain elusive. In this study, we showed by biological and molecular analyses that the linker region functions as an intramolecular modulator to tune Gag assembly, virion production, and viral infectivity. Our study thus illustrates a hitherto-unrecognized mechanism, an allosteric regulation of CA structure by the disordered protein element, for HIV-1 replication.

Project description:The histidine kinase CheA plays a central role in signal integration, conversion, and amplification in the bacterial chemotaxis signal transduction pathway. The kinase activity is regulated in chemotaxis signaling complexes formed via the interactions among CheA's regulatory domain (P5), the coupling protein CheW, and transmembrane chemoreceptors. Despite recent advancements in the understanding of the architecture of the signaling complex, the molecular mechanism underlying this regulation remains elusive. An interdomain linker that connects the catalytic (P4) and regulatory domains of CheA may mediate regulatory signals from the P5-CheW-receptor interactions to the catalytic domain. To investigate whether this interdomain linker is capable of both activating and inhibiting CheA, we performed in vivo screens to search for P4-P5 linker mutations that result in different CheA autokinase activities. Several CheA variants were identified with kinase activities ranging from 30% to 670% of the activity of wild-type CheA. All of these CheA variants were defective in receptor-mediated kinase activation, indicating that the natural receptor-mediated signal transmission pathway was simultaneously affected by these mutations. The altered P4-P5 linkers were sufficient for making significant changes in the kinase activity even in the absence of the P5 domain. Therefore, the interdomain linker is an active module that has the ability to impose regulatory effects on the catalytic activity of the P4 domain. These results suggest that chemoreceptors may manipulate the conformation of the P4-P5 linker to achieve CheA regulation in the platform of the signaling complex.IMPORTANCE The molecular mechanism underlying kinase regulation in bacterial chemotaxis signaling complexes formed by the regulatory domain of the histidine kinase CheA, the coupling protein CheW, and chemoreceptors is still unknown. We isolated and characterized mutations in the interdomain linker that connects the catalytic and regulatory domains of CheA and found that the linker mutations resulted in different CheA autokinase activities in the absence and presence of the regulatory domain as well as a defect in receptor-mediated kinase activation. These results demonstrate that the interdomain linker is an active module that has the ability to impose regulatory effects on CheA activity. Chemoreceptors may manipulate the conformation of this interdomain linker to achieve CheA regulation in the platform of the signaling complex.

Dataset Information

Role for Gag-CA Interdomain Linker in Primate Lentiviral Replication.

Publications

Role for Gag-CA Interdomain Linker in Primate Lentiviral Replication.

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