مجال
التميز
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تميز دراسي وبحثي + إبداع علمي
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البحوث المنشورة
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البحث (1):
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عنوان البحث:
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Factors affecting de novo RNA synthesis and
back-priming by the respiratory syncytial virus polymerase
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رابط إلى البحث:
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Click here
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تاريخ النشر:
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27/08/2014
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موجز عن البحث:
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Respiratory syncytial virus RNA dependent
RNA polymerase (RdRp) initiates RNA synthesis from the leader (le) and
trailer-complement (trc) promoters. The RdRp can also add nucleotides to the
3′ end of the trc promoter by back-priming, but there is no evidence this
occurs at the le promoter in infected cells. We examined how environmental
factors and RNA sequence affect de novo RNA synthesis versus back-priming
using an in vitro assay. We found that replacing Mg2+ with Mn2+ in the
reaction buffer increased de novo initiation relative to back-priming, and
different lengths of trc sequence were required for the two activities.
Experiments with le RNA showed that back-priming occurred with this sequence
in vitro, but less efficiently than with trc RNA. These findings indicate
that during infection, the RdRp is governed between de novo RNA synthesis and
back-priming by RNA sequence and environment, including a factor missing from
the in vitro assay.
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البحث (2):
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عنوان البحث:
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Interactome analysis of the human
respiratory syncytial virus RNA polymerase complex identifies protein
chaperones as important cofactors that promote L-protein stability and RNA
synthesis
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رابط إلى البحث:
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Click here
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تاريخ النشر:
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14/10/2014
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موجز عن البحث:
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The
human respiratory syncytial virus (HRSV) core viral RNA polymerase comprises
the large polymerase protein (L) and its cofactor, the phosphoprotein (P),
which associate with the viral ribonucleoprotein complex to replicate the
genome and, together with the M2-1 protein, transcribe viral mRNAs. While
cellular proteins have long been proposed to be involved in the synthesis of
HRSV RNA by associating with the polymerase complex, their characterization
has been hindered by the difficulty of purifying the viral polymerase from
mammalian cell culture. In this study, enhanced green fluorescent protein
(EGFP)-tagged L- and P-protein expression was coupled with high-affinity
anti-GFP antibody-based immunoprecipitation and quantitative proteomics to
identify cellular proteins that interacted with either the L- or the
P-proteins when expressed as part of a biologically active viral RNP. Several
core groups of cellular proteins were identified that interacted with each
viral protein including, in both cases, protein chaperones. Ablation of
chaperone activity by using small-molecule inhibitors confirmed previously
reported studies which suggested that this class of proteins acted as
positive viral factors. Inhibition of HSP90 chaperone function in the current
study showed that HSP90 is critical for L-protein function and stability,
whether in the presence or absence of the P-protein. Inhibition studies
suggested that HSP70 also disrupts virus biology and might help the
polymerase remodel the nucleocapsid to allow RNA synthesis to occur
efficiently. This indicated a proviral role for protein chaperones in HRSV
replication and demonstrates that the function of cellular proteins can be
targeted as potential therapeutics to disrupt virus replication.
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البحث (3):
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عنوان البحث:
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Investigating the influence of ribavirin
on human respiratory syncytial virus RNA synthesis using a high-resolution
RNAseq approach
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رابط إلى البحث:
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Click here
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تاريخ النشر:
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15/05/2016
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موجز عن البحث:
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Human respiratory syncytial virus (HRSV) is a major
cause of serious respiratory tract infection. Treatment options include
administration of ribavirin, a purine analog, although the mechanism of its
anti-HRSV activity is unknown. We used RNAseq to investigate genome mutation
frequency and viral mRNA accumulation in HRSV-infected cells that had been
untreated or treated with ribavirin. In the absence of ribavirin, HRSV
specific transcripts accounted for up to one third of total RNA reads from
the infected cell RNA population. Ribavirin treatment resulted in a greater
than 90% reduction in abundance of viral mRNA reads, while at the same time
no such reduction was detected for the abundance of cellular transcripts. The
presented data revealed that ribavirin significantly increased the frequency
of HRSV-specific RNA mutations, suggesting direct influence on the fidelity
of the HRSV polymerase. The presented data shows transition and transversion
occurs during HRSV replication, and that these changes occurred in ‘hot
spots’ along the HRSV genome. Examination of nucleotide substitution rates in
the viral genome indicated an increase in the frequency of transition but not
transversion mutations in the presence of ribavirin. In addition, our data
indicated that in the continuous cell types used, and at the time points
analyzed, the abundance of some HRSV mRNAs did not reflect the order in which
the mRNAs were transcribed.
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المؤتمرات العلمية:
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المؤتمر (1):
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عنوان المؤتمر:
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Society
of General Microbiology (SGM)
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تاريخ الإنعقاد:
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14/04/2014
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مكان
الإنعقاد:
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Liverpool, UK
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طبيعة المشاركة:
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Poster presentation
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عنوان المشاركة:
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Using label free proteomics to investigate
human respiratory syncytial virus and the effects of the antiviral ribavirin
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ملخص المشاركة:
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Human respiratory syncytial virus (HRSV)
causes severe lower respiratory tract infection in infants worldwide. Costly
antiviral therapies to HRSV are of limited efficacy. Ribavirin is one of the
most commonly used antiviral drugs licensed for the treatment of many
different virus infections including HRSV. The precise mechanism of how
ribavirin disrupts HRSV biology is unknown and both error catastrophe and
defects in cellular RNA metabolism have been proposed. This project used a
combination of label free quantitative proteomics and deep sequencing to
investigate the potential mechanism of action of ribavirin in the inhibition
of HRSV. Different treatments with ribavirin were analyzed for their effect
on HRSV biology. Selected examples were further characterized using the omic
approaches. Over 1500 cellular proteins were identified and quantified by
LC-MS/MS and pathway analysis showed significant changes in proteins
associated with RNA metabolism. Quasi species analysis of the viral RNA
genome suggested that the frequency of nucleotide changes over the assay
period was no different to when the virus was passaged on a different cell
type or treated with an alternative inhibitor of HRSV polymerase function.
The data suggests that the effect of ribavirin on HRSV is cellular mediated
rather than directly anti-viral.
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المؤتمر (2):
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عنوان المؤتمر:
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24th
European Congress of Clinical Microbiology and Infectious Diseases (ECCMID)
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تاريخ الإنعقاد:
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16/05/2014
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مكان
الإنعقاد:
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Barcelona-Spain
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طبيعة المشاركة:
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Poster presentation
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عنوان المشاركة:
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Using label free proteomics to investigate
human respiratory syncytial virus and the effects of the antiviral ribavirin
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ملخص المشاركة:
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Objectives: Ribavirin
is one of the most commonly used antiviral drugs licensed for the treatment
of many different virus infections including HRSV. The precise mechanism of how ribavirin
disrupts HRSV biology is unknown and both error catastrophe and defects in
cellular RNA metabolism have been proposed. The aim of this study is to
investigate the potential mechanism of ribavirin on the whole host cell
lysate in the absence and presence of HRSV.
Methods: This
project used a combination of label free quantitative proteomics and deep
sequencing to investigate the potential mechanism of action of ribavirin in
the inhibition of HRSV. Different
treatments with ribavirin were analyzed for their effect on HRSV
biology. Selected examples were
further characterized using the omic approaches.
Results: Over 1500 cellular proteins were identified
and quantified by LC-MS/MS and pathway analysis showed significant changes in
proteins associated with RNA metabolism.
Quasi species analysis of the viral RNA genome suggested that the
frequency of nucleotide changes over the assay period was no different to
when the virus was passaged on a different cell type or treated with an
alternative inhibitor of HRSV polymerase function.
Conclusions: The data suggests that the effect of
ribavirin on HRSV is cellular mediated rather than directly anti-viral.
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المؤتمر (3):
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عنوان المؤتمر:
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8th
Saudi Student conference
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تاريخ الإنعقاد:
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31/01-01/02/2015
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مكان
الإنعقاد:
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London,
UK
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طبيعة المشاركة:
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Poster
presentation
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عنوان المشاركة:
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Using
label free proteomics and RNA sequence to investigate human respiratory
syncytial virus and the effects of the antiviral ribavirin
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ملخص المشاركة:
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Different
treatment regimes with ribavirin were analyzed for their effect on HRSV
biology and selected examples were further characterized using the deep
discovery approaches. Pathway analysis
showed significant changes in the abundance of proteins associated with RNA
metabolism. Quasi species analysis of the viral RNA genome sequence suggested
that when ribavirin was given at an early time point post-infection then
nucleotide frequency changes increased, however at a later time point no
evidence of this could be found. Taken together the data suggested that the
effect of ribavirin on HRSV maybe biphasic and further work is ongoing to
investigate this.
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جوائز التكريم:
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الجائزة (1):
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مسمى الجائزة:
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Award of best poster
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الجهة المانحة:
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8th
Saudi Student Conference
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تاريخ الجائزة:
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01/02/2015
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مجال التكريم:
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Medical and Health Sciences
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