Title: Pathogenic and attenuated rabies viruses induces differential host protein expression in the central nervous system: Implication of neuronal dysfunction
1Pathogenic and attenuated rabies viruses induces
differential host protein expression in the
central nervous system Implication of neuronal
dysfunction
- Zhen F. Fu
- Department of Pathology
- University of Georgia
2Rabies Pathogenesis
Robert Hurt-USC
Patients die of circulatory insufficiency,
cardiac arrest and respiratory failure.
3Despite extensive research in the past 100 years,
we still know very little about the pathogenic
mechanism by which rabies virus infection of
neurons causes rabies.There are scarce
neuropathology with mild inflammation and little
neuronal loss, which cannot explain the lethality
of the disease.It has been hypothesized that
rabies results from neuronal dysfunction rather
than structural damage. However, it is not known
how RV infection leads to neuronal
dysfunction.To better understand rabies
pathogenesis, we initiate a project to determine
how the host responds to rabies virus infections
using one street and one fixed virus. This is
accomplished by using proteomics technologies.
4Two viruses were used in this studySHBRV Wt
virus, normally circulating in silver-haired bats
and responsible for most of the human rabies in
the US.CVS-B2C Lab-adapted attenuated virus
derived from CVS-24 by passaging in BHK cells.
5Survival curve
6Detection of Differential Protein Levels in the
Proteome
7mutant/Infected
Wild type
In-gel digestion with protease Identify the
protein by Mass Spectrometry
8Identification of gel-separated proteins by mass
spectrometry
Gygi et al.
9The situation to avoid
Software is essential !
10(No Transcript)
11Proteins differentially expressed in response to
SHBRV infection
Category Protein PI Molecular weight (kDa) Up/down-regulation Fold change
Ion Homeostasis Na/K ATPase 5.3 110 ? 3.0
H ATPase subunit a isoform 1 6.0 96 ? 2.6
Synaptic Physiology TRIM 9 6.6 90 ? -3.0
?-SNAP 5.3 33 ? -3.0
Pallidin (Syntaxin 13) 5.8 20 ? -2.2
Synexin 5.9 50 ? -2.0
Syntaxin 18 5.2 35 ? -1.8
12Proteins differentially expressed in response to
B2C infection in mice
Category Protein PI Molecular weight (kDa) Up/down regulation Fold change
Innate immunity G protein coupled receptor 44 4.8 88 ? 3
Hsp 60 5.9 60 ? 3.2
Ion Homeostasis Villin 1 5.1 89 ? 2
Calretinin 4.9 31 ? 2.2
H ATPase synthase subunit b 5.5 58 ? 2.8
Apoptosis APAF 1 6.0 143 ? 2.4
SH3 domain binding protein 2 7.6 62 ? 2.6
Neuro-Physiology Neurofilament protein NF 66 5.1 60 ? 3
CRMP-2 6.6 62 ? 3.6
13Western blotting of proteins involved in ion
homeostasis and synaptic physiology
14(No Transcript)
15(No Transcript)
16(No Transcript)
17Accumulation of vesicles
No Docking
No release of Neurotransmitters
18SHBRV B2C Control
19Conclusions
Proteomics data indicate that wt RV infection
resulted in up-regulation of proteins involved in
ion homeostasis and down-regulation of synaptic
proteins. The altered protein expression as
detected by 2D-gel analysis is confirmed by
Western blotting in animals infected either ic or
im as well as in primary neuron. Up-regulation
of Na/K-ATPase leads to decrease in Na
concentrations in infected cells. Likewise,
down-regulation of Ca-ATPase resulted in decrease
of Ca concentration in infected cells. Changes
in Na/Ca concentration affects membrane potential
and thus leading to alteration of neuronal
transmission. Synaptic proteins such as
syntaxin, a-SANP, and TRIM9 play important roles
in synaptic-vesicle fusion and docking of
synaptic vesicles. Down-regulation of these
proteins prevented the docking and fusion of
synaptic vesicles with presynaptic membrane, thus
resulting in accumulation of synaptic vesicles in
the presynapses. Thus our data may provide
structural and metabolic basis by which RV
infection causes neuronal dysfunction.
20Acknowledgements Vikas Dhingra Xia-qin
g Li Luciana Sarmento UGA
Proteomics Facility Tracy Andachtc
21Thank you!!