HCV part 1 Flashcards
How did people discover hep c
In the 1980s large numbers of hepatitis patients began to appear, apparently with a virally caused disease
* When examined, patients tested negative for both hepatitis A and B
* HCV, the etiologic agent of non-A, non-B hepatitis, was discovered by Michael Houghton and investigators at Chiron, Inc.
true or false: it was hard to discover hep c and which techniques were used
-true
- cdna, phages for e.coli, expressed the modified phage that had the cdna in them
which virus is responsible for 40-60% of the chronic liver disease
Hep C
True or false: Hep C is the leading cause of liver transfer
treu
how do you get hep c
- Blood transfusion
- Injecting drug use
- High-risk sexual activity
- Other
- Health-care workers
- Hemodialysis
- Mother-to-child (low)
- Unknown
- Household exposures/accidental injuries
- Cocaine use
Diagnosis of hcv
- Not part of a standard STD panel
- Serology (enzyme immunosorbent assay)
- Look for anti-HCV antibodies in the blood (once
exposed to infection) - > 95% of chronically infected patients
- Viral genome copies (qPCR)
- Look for viral RNA in the blood (active infection)
- genotyping
- Assess degree of liver damage
- Serum liver enzyme elevation (liver health);
Fibroscan (Liver inflammation/fibrosis)
what is the incubation pereod of acude hepatitis
6-10 weeks
True or false: 80% people with acute HCV will have no symptoms
true
symptoms of acute hepatitis, clearance rate
Symptoms can include:
- Pain in the upper right quadrant,
anorexia, abdominal pain, nausea,
vomiting, fever, fatigue, and jaundice
(25%)
* Approximately 15% will clear the infection
* 85% will develop chronic HCV
True or false: chronic hepatitis can last your whole life
true
what happens to your liver when you have chronic hepatitis
- Cirrhosis (scar tissue) and liver failure (10-20%)
- Hepatocellular Carcinoma (liver cancer, 3-5%)
- Typically clinical symptoms appear during liver failure
- 20 yrs may elapse between infection and the development of serious complications
HCV genome
- 9.6 kb, (+) sense, ssRNA virus
- Single long open reading frame (~3000 aa polyprotein)
- cleaved by host and viral proteases into the 10 mature viral proteins
-has IRES, kissing loop and a pseudi knot
What are the 2 models for LVPs of HCV
-lvp: lipoviral particles
-two particle model: it interacts with more than one apo protein: LDL and HDL
-single particles: the most accurate, interacts with one lipoprotein
HCV entry
Receptors: CD81, CLDN-1, OCLN
* Co-receptors/co-factors: GAGs, LDLR, SR-BI
* Lipoproteins play a prominent role in entry (i.e. LVPs)
* Clathrin-mediated endocytosis
true or false: HCV is capped
false it is not
translation of HCV
Cap-independent translation is
accomplished by the Internal
Ribosome Entry Site (IRES)
element
- SLII-IV of the 5’ NCR
- Binds to the 40S subunit of the
ribosome (uses only eIF2 & 3)
- Places the start codon (AUG)
in the P site of the ribosome
(for initiation of translation)
what do NS3 and Ns5B do in hcv
they form the replication complex
* Replication Complex:
* NS3 (helicase) + NS4A (NS3 cofactor)
* NS4B = membrane reorganization
* NS5A = phosphoprotein, RNA replication and assembly?
* NS5B = RNA-dependent RNA polymerase (RdRp)
HCV rna replication occurs on///
membranous webs
* Vesicles contain pores that
may allow transport of viral
genomic RNA in and out
* The pore stains with nuclear
pore complex-specific
antibodies
* Nuclear localization
sequences can direct
proteins to the inside of these
vesicles
true or false: membranous webs are uncommon with +rna viruses
false they are common
what is the use of membranous webs
- Physical support?
- Compartmentalization
- Protection
- Tethering of RNA from unwinding
- Retention of the negative strand
membranous webs are derived from which membrane and are induced by what?
- Derived from the ER
- Lipid constituents (lipid droplets, LDs)
- Double Membrane Vesicles (DMVs)
- Multi-Membrane Vesicles (MMVs)
- Induced by NS4B
what is NS5B responsible for in HCV
- RNA-dependent RNA polymerase
- ‘right-hand’ topology
- initiation is de novo
- GTP is generally accepted as the
initiating NTP (in vitro) - requires divalent metal ions
true or false: HCV rdrp is error prone af
true
Error prone (replication rate = 1012
viral particles/day)
- RESULT: spectrum of genetic
variants (genotypes/subtypes;
quasispecies)
true or false: we still don’t know all the tea about hcv assembly
true
virion assembly hcv
-Both structural proteins (E1/E2, Core) and nonstructural proteins
-(p7, NS2, NS3/4A, NS5A) participate in assembly and packaging
what does p7 do? in hcv
Viroporin = ion channel; promotes pH-mediated maturation
and release of infectious viral particles from the cell
how do we study hcv
-hcvpp
-replicons
-hcvcc
HCVpp
-pseudo particles: basically faje outside particles
* Pros:
- easy to perform
- entry inhibitors
- glycoprotein mutagenesis
* Cons:
- particles made in non-hepatic cells
‣ glycosylation?
‣ misfolding (E1/E2)?
- inefficient
Replicons
-basically NS proteins with reporter genes
* Pros:
- non-infectious
- isolates the replication step
‣ protease, polymerase, and NS5A
inhibitors
- multiple genotypes can be studied
* Cons:
- Ignores contributions from the structural
proteins
- Results might not translate to infectious
cycle
HCVcc
-cell culture derived hcv; discovered by a japanase dude. Works really well
- JFH-1 produces infectious virus in cell culture (10^2-10^3 ffu/
ml)
* Huh-7.5 cells
- sub-clone of Huh-7 with an inactivating mutation in RIG-I
* Chimeric viruses now exist for all 7 genotypes
- produce varying levels of infectious virus
* Pros:
- Structural genes from all genotypes can be studied
* Cons:
- completely artificial
- all contain JFH-1 nonstructural genes
