Step 2: Entry and Fusion (Lecture 10) Flashcards
What is the first step in viral entry/fusion?
attachment of viral receptor binding protein to viral receptor molecule on cell surface
What are the 2 roles that viral receptors play in uncoating?
initiate conformational changes to capsid and direct the virus into the endocytic pathway
How are particles taken in via phagocytosis?
actin-mediated event
What are the events in phagocytosis?
actin pushes cell membrane forward around whatever it’s going to take in »_space;> taken to endosome = phagosome »_space;> targeted to lysosome
What is significant about lysosomes? What do they contain?
digestive enzymes and many protons = very acidic = degrades protein
How are particles taken in via receptor-mediated endocytosis?
clathrin-coated vesicles (or calveolin)
What are the events in receptor-mediated endocytosis with clathrin?
clathrin-coated vesicle takes in substance »_space;> becomes early endosome »_space;> H+ gets pumped inside = late endosome (at right pH) »_space;> fuses with lysosome
How do we distinguish between an early and late endosome?
level of pH, early endosome is more basic than late endosome as late endosome is more acidic
What is the benefit/effect to the receptor of the differing levels of pH?
an increase in H+ = increase of removing the viral receptors off and recycle it back to plasma membrane for reuse
What are the 4 triggering events that will cause the nucleocapsid to pop open?
receptor engagement, endosomal acidification, 2-entry method, receptor-mediated signaling event
What is receptor engagement?
basic conformational changes will occur due to the receptor-ligand interaction
What are the 2 ways that receptor engagement triggers viral entry/fusion?
1) fusion peptide is exposed and inserts itself onto host membrane and then delivers nucleocapsid into cytoplasm | 2) receptor-ligand interaction causes a pore formation on host cell membrane and injects nucleic acid into cytoplasm
What is a fusion peptide and how is it exposed?
it is a second viral protein on the surface of the viral envelope; it is initially hidden but is exposed when virus binds to receptor on host cell membrane and causes a conformational change thus exposing the fusion peptide
What is the triggering event in endosomal acidification?
acidity of endosome will expose viral fusion peptide = causes fusion
At what pH does influenza pop open? What stage of endosome is this?
pH = 5 at the late endosome
What are the events that take place in the entry/fusion of influenza?
receptor-ligand interaction causes virus to be uptaken by clathrin-coated vesicles »_space;> then it begins to pump H+ in endosome »_space;> turns into late endosome with acidic pH »_space;> fusion peptide inserts into host membrane = fusion of membranes »_space;> nucleic acids are released and directed to nucleus
What allows influenza’s RNA genome to be directed to nucleus?
ribonucleo proteins (RNPs); genome is segmented
At what pH does the fusion peptide get exposed in flaviviruses?
very very low pH (lysosome or late endosome)
What are the events that take place in the entry/fusion of flaviviruses?
receptor-ligand interaction causes virus to be uptaken via fusion (endocytosis) »_space;> early endosome turns into late endosome with the pumping of H+ »_space;> fusion peptide is exposed and brings the membranes together allowing fusion »_space;> RNA genome is released
What is the 2-entry mechanism of viral entry and fusion?
two distinct and separate entry mechanisms that go hand-in-hand to get inside the host and help with uncoating and delivery; steps vary between viruses
What are the 2 step entry mechanisms for Semliki Forest Viruses? What are they used for/do?
1) endosomal acidification induces membrane fusion but capsid is still bound to the cytoplasmic leaflet of the endosomal bilayer »_space;> 2) ribosomes bind the capsid and disassemble it to release the nucleic acid into cytoplasm
What are the 2 step entry mechanisms for Adenoviruses? What are they used for/do?
1) endosomal acidification destabilizes capsid and releases Protein IV = disrupt endosomal membrane to release capsid to cytoplasm »_space;> 2) microtubules deliver and dock unstable capsid onto nuclear pore complex where the genome practically swims out into nucleus
What are the events that take place in the entry/fusion of Ebola viruses? What are the 2-step entry mechanisms it uses?
Ebola gets taken up via CCV »_space;> early endosome »_space;> late endosome »_space;> lysosome »_space;> proteases cleave off gp protein off of ebola’s receptors = allows it to bind to NPC1 »_space;> causes fusion event and releases nucleocapsid into cytoplasm ||| endosomal acidification and binding of cleaved gp with NPC1
What is so unique about Ebola?
uses a co-receptor only found on the late endosome or lysosome
What is the coreceptor that ebola uses?
NPC1
What is the receptor-mediated signaling method of viral entry/fusion?
signaling cascade allows virus to enter in a unique way
What are the 2 receptors that coxsackie virus requires?
DAF (decay-accelerating factor) and CAR (coxsackievirus-adenovirus receptor)
What is the main receptor for Coxsackie viruses? What is it’s hurdle?
CAR receptor = buried between tight junctions making it unavailable for virus to bind to it
How does Coxsackie virus get to it’s receptor?
Coxsackie does electrostatic interactions on cell surface and will eventually interact with DAF »_space;> signaling cascade remodels actin causing membrane to ruffle = pushes virus to surf close to the tight junction proteins =CAR exposed »_space;> virus binds to CAR and gets taken up via endocytic vesicle
Why does the Coxsackie virus need the DAF receptor?
causes actin remodeling due to signaling event
What are the 5 ways viruses regulate viral entry and fusion?
1) makes sure that genome gets into right cell (cytoplasm can easily degrade RNA) || 2) virus only fuses with right cell || 3) prevents aggregation of newly assembled virions by keeping genome delivery and virion assembly separate areas in cell || 4) maintains cell’s integrity by using cellular processes because it needs to survive || 5) prevents premature activation of fusion peptides during assembly in order to successfully deliver genome onto next cell
What is the role of the nuclear pore complex in viruses?
ensures that viruses (especially DNA viruses) deliver their genome into the nucleus
What are the 4 different strategies to delivering genome into nucleus?
viral ribonucleic proteins direct genome to nucleus, un-dismantled capsid docks onto NPC and injects genome, dismantled capsid docks onto NPC and genome flows through into nucleus, virus disrupts nuclear envelope to go into nucleus
What do the RNPs need to have on them to get the nucleic acid into the nucleus?
specific signal sequence that other proteins recognize to bring into nucleus (ie: importin)
