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Interferon-induced HERC5 protein inhibits HIV-1
replication
byistwo
novel
mechanisms
and is evolving
Interferon-induced
HERC5
protein
inhibits
replication
by two novel
mechanisms
and
evolving
under
positiveHIV-1
selection
under positive selection
Matthew Woods (Western University, Canada: Barr Lab)
Session Title: Restriction Factors and HIV-1 Replication
21/07/2014
Interferon Inhibits HIV-1 Replication
Mature Virion
Release
Maturation
Tetherin
Budding
Binding
U
U
U
Fusion
Viperin
Assembly
Uncoating
U
APOBEC
Gag
TRIM22
U
Trafficking
Nuclear Import
Translation
U
U
Integration
UU
U U U
U U U
U
UU
U U U
ISG20
U
U U U
PKR
Nuclear
Export
U
Transcription
& Splicing
U
SAMHD1
U
T
T
TRIM5α
Reverse
Transcription
OAS1/RNaseL
PKR
Identifying new interferon induced anti-HIV-1 factors
• Our lab is interested in characterizing and identifying
novel interferon-induced proteins capable of restricting
HIV-1 particle production
• Surveyed IFN-induced genes for potential HIV-1
restriction factor candidates and identified HERC5.
HERC5 is a novel interferon-induced antiviral protein
•
HERC5 expression is up-regulated in response to IFN and both in vitro and in vivo virus infection.
•
By virtue of its HECT domain, HERC5 is the main cellular E3 ligase that conjugates ISG15 to
specific target proteins (Proc Natl Acad Sci U S A. 2006; J Biol Chem. 2006 Feb 17;281(7):4334-8)
•
Tang and colleagues showed that HERC5 inhibits influenza A virus replication by catalyzing the
conjugation of ISG15 to the viral NS1 protein (J Immunol. 2010 May 15;184(10):5777-90).
•
Durfee and colleagues showed that HERC5 conjugates ISG15 to the human papillomavirus (HPV)
L1 capsid protein, conferring a dominant-inhibitory effect on the infectivity of HPV16 pseudoviruses
(Mol Cell. 2010 Jun 11;38(5):722-32).
HERC5 is multifunctional
• We previously showed that HERC5 inhibits an early step of HIV-1 Gag
assembly via the modification of Gag with ISG15 (Retrovirology. 2011 Nov 17;8:95)
1. E3 ligase-dependent
ISG15
X
HERC5 is multifunctional
• We previously showed that HERC5 inhibits an early step of HIV-1 Gag
assembly via the modification of Gag with ISG15 (Retrovirology. 2011 Nov 17;8:95)
1. E3 ligase-dependent
ISG15
2. E3 ligase-independent
X
cytoplasm
nucleus
HERC5
X
Does HERC5 restrict infectious HIV-1 production?
HIV only
Cell
HIV
HERC5
HIV + HERC5
HERC5 inhibits infectious HIV-1 particle production
A
B
Do biological levels of HERC5 reduce Gag production?
293T Cells
Human MDM
What is causing the decrease of Gag within the cell
pellet?
Is Gag degraded in the presence of HERC5?
Does HERC5 interfere with the nuclear export of
Gag-containing mRNA?
MS2 System To Localize
HIV RNA Genomes Inside Cells
GFP
MS2 Stem Loops
HIV-1 (Stem Loop)
MS2-GFP
MS2-GFP
HIV-1 (Stem Loop) + MS2-GFP
HERC5 interferes with export of genomic RNA
HIV-1 and REV
Late Proteins (Ex. Gag, Env, and Genome)
REV
Early Proteins (Ex. Tat, Rev Nef)
REV
REV
REV
REV
Unspliced
Fully spliced
REV
Rev dependant
Rev independent
HERC5 interferes with export of unspliced RNA
Rev dependent
Rev independent
Viral mRNA nuclear export
HERC5 inhibits export of Rev/RRE-dependent RNA
Rev/RRE-dependent
Rev/RRE-independent
HERC5 interferes with Rev localization
Which domain is required for inhibition?
RLD domain is required for the loss of Gag accumulation
E3 ligase-independent
Evolutionary Arms Race Between
Host and Pathogen
VS
Neighbor-joining phylogenetic tree for progressive
alignment of 13 HERC5 orthologs using COBALT
Strong positive selection is operating on
the first 100 amino acids of the RLD
Strong positive selection is operating on
the first 100 amino acids of the RLD
Conclusions
1. E3 ligase-independent
cytoplasm
nucleus
HERC5
2. E3 ligase-dependent
ISG15
X
3. HERC5 is evolving under positive selection
X
Acknowledgements
Supervisor
Past Lab Members
Dr. Stephen Barr
Clayton Hattlmann
Jess Tong
Lab Members
Sherry Xu
Jenna Kelly
Macon Coleman
Sean Tom
Divy Kochar
Collaborators
Johnny Zhang
Dr. Dikeakos
Mike Ding
Craig Cavanagh
Dr. Haeryfar
Peter Szabo
Funding Agencies