title: Stimulated Emission Depletion Nanoscopy Reveals Time-Course of Human Immunodeficiency Virus Proteolytic Maturation.
abstract: Concomitant with human immunodeficiency virus type 1 (HIV-1) budding from a host cell, cleavage of the structural Gag polyproteins by the viral protease (PR) triggers complete remodeling of virion architecture. This maturation process is essential for virus infectivity. Electron tomography provided structures of immature and mature HIV-1 with a diameter of 120-140 nm, but information about the sequence and dynamics of structural rearrangements is lacking. Here, we employed super-resolution STED (stimulated emission depletion) fluorescence nanoscopy of HIV-1 carrying labeled Gag to visualize the virion architecture. The incomplete Gag lattice of immature virions was clearly distinguishable from the condensed distribution of mature protein subunits. Synchronized activation of PR within purified particles by photocleavage of a caged PR inhibitor enabled time-resolved in situ observation of the induction of proteolysis and maturation by super-resolution microscopy. This study shows the rearrangement of subviral structures in a super-resolution light microscope over time, outwitting phototoxicity and fluorophore bleaching through synchronization of a biological process by an optical switch.
authors: Department of Infectious Diseases, Virology, Heidelberg University , Im Neuenheimer Feld 324, 69120 Heidelberg, Germany.|Optical Nanoscopy Division, German Cancer Research Center (DKFZ) , Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.
authors: Janina
authors: J
authors: Hanne
authors: Department of NanoBiophotonics, Max Planck Institute for Biophysical Chemistry , Am Fassberg 11, 37077 Göttingen, Germany.
authors: Fabian
authors: F
authors: Göttfert
authors: Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Gilead Sciences and IOCB Research Center , Flemingovo n.2, 166 10 Prague 6, Czech Republic.|Department of Biochemistry, Faculty of Science, Charles University in Prague , Hlavova 8, 12843 Prague 2, Czech Republic.
authors: Jiří
authors: J
authors: Schimer
authors: Department of Infectious Diseases, Virology, Heidelberg University , Im Neuenheimer Feld 324, 69120 Heidelberg, Germany.
authors: Maria
authors: M
authors: Anders-Össwein
authors: Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Gilead Sciences and IOCB Research Center , Flemingovo n.2, 166 10 Prague 6, Czech Republic.|Department of Biochemistry, Faculty of Science, Charles University in Prague , Hlavova 8, 12843 Prague 2, Czech Republic.
authors: Jan
authors: J
authors: Konvalinka
authors: Optical Nanoscopy Division, German Cancer Research Center (DKFZ) , Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.
authors: Johann
authors: J
authors: Engelhardt
authors: Department of Infectious Diseases, Virology, Heidelberg University , Im Neuenheimer Feld 324, 69120 Heidelberg, Germany.|Molecular Medicine Partnership Unit , Meyerhofstrasse 1, 69117 Heidelberg, Germany.
authors: Barbara
authors: B
authors: Müller
authors: Optical Nanoscopy Division, German Cancer Research Center (DKFZ) , Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.|Department of NanoBiophotonics, Max Planck Institute for Biophysical Chemistry , Am Fassberg 11, 37077 Göttingen, Germany.
authors: Stefan W
authors: SW
authors: Hell
authors: Department of Infectious Diseases, Virology, Heidelberg University , Im Neuenheimer Feld 324, 69120 Heidelberg, Germany.|Molecular Medicine Partnership Unit , Meyerhofstrasse 1, 69117 Heidelberg, Germany.
authors: Hans-Georg
authors: HG
authors: Kräusslich
mesh_terms: D055032:Electron Microscope Tomography; D015658:HIV Infections; D015497:HIV-1; D006801:Humans; D010455:Peptides; D059748:Proteolysis; D014771:Virion; D054301:gag Gene Products, Human Immunodeficiency Virus / Q000737:chemistry*
chemical_list: C524345:Nano-1 peptide; D010455:Peptides; D054301:gag Gene Products, Human Immunodeficiency Virus
keywords: HIV-1 maturation; STED nanoscopy; native virus imaging; super-resolution microscopy