title: The Andes Virus Nucleocapsid Protein Directs Basal Endothelial Cell Permeability by Activating RhoA.
abstract: Andes virus (ANDV) predominantly infects microvascular endothelial cells (MECs) and nonlytically causes an acute pulmonary edema termed hantavirus pulmonary syndrome (HPS). In HPS patients, virtually every pulmonary MEC is infected, MECs are enlarged, and infection results in vascular leakage and highly lethal pulmonary edema. We observed that MECs infected with the ANDV hantavirus or expressing the ANDV nucleocapsid (N) protein showed increased size and permeability by activating the Rheb and RhoA GTPases. Expression of ANDV N in MECs increased cell size by preventing tuberous sclerosis complex (TSC) repression of Rheb-mTOR-pS6K. N selectively bound the TSC2 N terminus (1 to 1403) within a complex containing TSC2/TSC1/TBC1D7, and endogenous TSC2 reciprocally coprecipitated N protein from ANDV-infected MECs. TSCs normally restrict RhoA-induced MEC permeability, and we found that ANDV infection or N protein expression constitutively activated RhoA. This suggests that the ANDV N protein alone is sufficient to activate signaling pathways that control MEC size and permeability. Further, RhoA small interfering RNA, dominant-negative RhoA(N19), and the RhoA/Rho kinase inhibitors fasudil and Y27632 dramatically reduced the permeability of ANDV-infected MECs by 80 to 90%. Fasudil also reduced the bradykinin-directed permeability of ANDV and Hantaan virus-infected MECs to control levels. These findings demonstrate that ANDV activation of RhoA causes MEC permeability and reveal a potential edemagenic mechanism for ANDV to constitutively inhibit the basal barrier integrity of infected MECs. The central importance of RhoA activation in MEC permeability further suggests therapeutically targeting RhoA, TSCs, and Rac1 as potential means of resolving capillary leakage during hantavirus infections.
  
  
  
  HPS is hallmarked by acute pulmonary edema, hypoxia, respiratory distress, and the ubiquitous infection of pulmonary MECs that occurs without disrupting the endothelium. Mechanisms of MEC permeability and targets for resolving lethal pulmonary edema during HPS remain enigmatic. Our findings suggest a novel underlying mechanism of MEC dysfunction resulting from ANDV activation of the Rheb and RhoA GTPases that, respectively, control MEC size and permeability. Our studies show that inhibition of RhoA blocks ANDV-directed permeability and implicate RhoA as a potential therapeutic target for restoring capillary barrier function to the ANDV-infected endothelium. Since RhoA activation forms a downstream nexus for factors that cause capillary leakage, blocking RhoA activation is liable to restore basal capillary integrity and prevent edema amplified by tissue hypoxia and respiratory distress. Targeting the endothelium has the potential to resolve disease during symptomatic stages, when replication inhibitors lack efficacy, and to be broadly applicable to other hemorrhagic and edematous viral diseases.
authors: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, USA.
authors: Elena E
authors: EE
authors: Gorbunova
authors: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, USA.|Molecular and Cell Biology Program, Stony Brook University, Stony Brook, New York, USA.
authors: Matthew J
authors: MJ
authors: Simons
authors: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, USA.
authors: Irina N
authors: IN
authors: Gavrilovskaya
authors: Department of Molecular Genetics and Microbiology, Stony Brook University, Stony Brook, New York, USA Erich.Mackow@stonybrook.edu.|Molecular and Cell Biology Program, Stony Brook University, Stony Brook, New York, USA.
authors: Erich R
authors: ER
authors: Mackow
mesh_terms: D042783:Endothelial Cells / Q000502:physiology* / Q000821:virology*; D006476:Orthohantavirus / Q000502:physiology*; D054884:Host-Pathogen Interactions; D019590:Nucleocapsid Proteins / Q000378:metabolism*; D010539:Permeability; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
chemical_list: D019590:Nucleocapsid Proteins; D020742:rhoA GTP-Binding Protein