Membrane Hijacking: Biogenesis and Fate of Quasi-Enveloped Hepatovirus

膜劫持：准包膜肝病毒的生物发生和命运

• 批准号: 10223138
• 负责人: Stanley M. Lemon
• 金额: $ 38.88万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-24 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10223138
• 关键词: ARHGEF5 gene; Autophagocytosis; Biogenesis; Biological Assay; Biology; C-terminal; CRISPR/Cas technology; Capsid; Capsid Proteins; Cell Culture Techniques; Cell membrane; Cells; Complement; Complex; Data; Endocytosis; Endosomes; Enteral; Enterovirus; Enzymes; Extracellular Space; Family; Family Picornaviridae; Galactose Binding Lectin; Glycoproteins; Goals; Grant; Hepatitis A; Hepatitis A Virus; Hepatovirus; Human; Image; In Vitro; Infection; Knock-out; Laboratories; Lectin; Ligase; Lysosomes; Maps; Mediating; Membrane; Molecular; Molecular Genetics; Multivesicular Body; Mus; Nonlytic; Outcome; Pathogenesis; Pathway interactions; Phospholipase; Phospholipases A; Process; Proteins; Proteomics; Reporter; Research; Role; Sampling; Signal Transduction; Site; Sorting - Cell Movement; TSG101 gene; Tertiary Protein Structure; Tissues; Vesicle; Viral; Viral hepatitis; Virion; Virus; Virus Diseases; Work; base; biophysical properties; density; exosome; extracellular vesicles; hepatitis A virus receptor; human model; human pathogen; in vivo; late endosome; lysosomal proteins; lysosome membrane; member; mouse model; novel; receptor; response; tissue tropism; trafficking; ubiquitin-protein ligase; uptake; virology; virus envelope

PROJECT ABSTRACT Hepatitis A virus (HAV), a member of the Picornaviridae (genus Hepatovirus), is an ancient human pathogen and a common cause of enterically-transmitted viral hepatitis globally. Long considered `nonenveloped', we discovered that HAV is released noncytolytically from infected cells both in vitro and in infected humans in vivo within extracellular vesicles that are similar in size and buoyant density to exosomes. The HAV capsid is completely cloaked by the membranes of these small vesicles, which nonetheless possess specific infectivity indistinguishable from naked HAV virions. The goal of this grant is to elucidate mechanisms underlying the biogenesis of these `quasi-enveloped' HAV (eHAV) virions and their role in the pathogenesis of hepatitis A. Extensive proteomics, virologic, and biophysical characterization of eHAV produced in cell culture demonstrate that viral capsids are selected for export in eHAV vesicles via a highly specific sorting process that is dependent on sequence in the pX domain of the VP1 capsid protein. We hypothesize (1) that the quasi- envelopment of eHAV results from budding of assembled VP1pX-containing capsids into endosomes (multivesicular bodies) in an ESCRT-dependent process mediated by specific interactions of capsid proteins with ESCRT complexes and facilitated by NEDD4-family E3 ubiquitin ligases; and (2) that cell entry by eHAV differs from naked virions and occurs within late endosome-lysosomes where membranes are hydrolyzed by lysosomal enzymes, providing capsid access to an unknown cellular receptor. In Specific Aim 1, we will map interactions of ESCRT components with VP1pX and ascertain the functional importance of ESCRT-0 and -I components in CRISPR/Cas9 knockout cells. Specific Aim 2 will determine whether catalytic activity of NEDD4-family ligases is required for eHAV biogenesis and contributes to ubiquitylation of pX or other capsid protein domains, and ascertain the extent to which core components of noncanonical `secretory autophagy' are involved in nonlytic eHAV egress. Specific Aim 3 will contrast mechanisms of eHAV vs. HAV entry, including endocytosis and endocytic transport, and define tissue tropisms of eHAV vs. HAV. Results from in vitro studies will be validated in vivo using a novel murine model of human hepatitis A. The discovery of eHAV has profoundly altered our understanding of the pathogenesis of this hepatotropic human virus and has substantial ramifications for the biology of other noncytopathic `nonenveloped' viruses, making this research of broad relevance to the field of virology.

项目摘要

项目成果

Biliary Secretion of Quasi-Enveloped Human Hepatitis A Virus.

• DOI: 10.1128/mbio.01998-16
• 发表时间: 2016-12-06
• 期刊: mBio
• 影响因子: 6.4
• 作者: Hirai-Yuki A;Hensley L;Whitmire JK;Lemon SM
• 通讯作者: Lemon SM

Peek-a-boo: membrane hijacking and the pathogenesis of viral hepatitis.

• DOI: 10.1016/j.tim.2013.10.005
• 发表时间: 2014-02
• 期刊: TRENDS IN MICROBIOLOGY
• 影响因子: 15.9
• 作者: Feng, Zongdi;Lemon, Stanley M.
• 通讯作者: Lemon, Stanley M.

Reassessing immune control of hepatitis A virus.

• DOI: 10.1016/j.coviro.2015.01.003
• 发表时间: 2015-04
• 期刊: CURRENT OPINION IN VIROLOGY
• 影响因子: 5.9
• 作者: Walker, Christopher M.;Feng, Zongdi;Lemon, Stanley M.
• 通讯作者: Lemon, Stanley M.

MAVS-dependent host species range and pathogenicity of human hepatitis A virus.

• DOI: 10.1126/science.aaf8325
• 发表时间: 2016-09-30
• 期刊: Science (New York, N.Y.)
• 作者: Hirai-Yuki A;Hensley L;McGivern DR;González-López O;Das A;Feng H;Sun L;Wilson JE;Hu F;Feng Z;Lovell W;Misumi I;Ting JP;Montgomery S;Cullen J;Whitmire JK;Lemon SM
• 通讯作者: Lemon SM