Global Human-Kinase Screening Identifies Therapeutic Host Targets against Influenza
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Global Human-Kinase Screening Identifies Therapeutic Host Targets against Influenza. / Atkins, Colm; Evans, Carrie W; Nordin, Brian; Patricelli, Matthew P; Reynolds, Robert; Wennerberg, Krister; Noah, James W.
In: Journal of Biomolecular Screening, Vol. 19, No. 6, 07.2014, p. 936-46.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - Global Human-Kinase Screening Identifies Therapeutic Host Targets against Influenza
AU - Atkins, Colm
AU - Evans, Carrie W
AU - Nordin, Brian
AU - Patricelli, Matthew P
AU - Reynolds, Robert
AU - Wennerberg, Krister
AU - Noah, James W
N1 - © 2014 Society for Laboratory Automation and Screening.
PY - 2014/7
Y1 - 2014/7
N2 - During viral infection of human cells, host kinases mediate signaling activities that are used by all viruses for replication; therefore, targeting of host kinases is of broad therapeutic interest. Here, host kinases were globally screened during human influenza virus (H1N1) infection to determine the time-dependent effects of virus infection and replication on kinase function. Desthiobiotin-labeled analogs of adenosine triphosphate and adenosine diphosphate were used to probe and covalently label host kinases in infected cell lysates, and probe affinity was determined. Using infected human A549 cells, we screened for time-dependent signal changes and identified host kinases whose probe affinities differed significantly when compared to uninfected cells. Our screen identified 10 novel host kinases that have not been previously shown to be involved with influenza virus replication, and we validated the functional importance of these novel kinases during infection using targeted small interfering RNAs (siRNAs). The effects of kinase-targeted siRNA knockdowns on replicating virus levels were measured by quantitative reverse-transcription PCR and cytoprotection assays. We identified several novel host kinases that, when knocked down, enhanced or reduced the viral load in cell culture. This preliminary work represents the first screen of the changing host kinome in influenza virus-infected human cells.
AB - During viral infection of human cells, host kinases mediate signaling activities that are used by all viruses for replication; therefore, targeting of host kinases is of broad therapeutic interest. Here, host kinases were globally screened during human influenza virus (H1N1) infection to determine the time-dependent effects of virus infection and replication on kinase function. Desthiobiotin-labeled analogs of adenosine triphosphate and adenosine diphosphate were used to probe and covalently label host kinases in infected cell lysates, and probe affinity was determined. Using infected human A549 cells, we screened for time-dependent signal changes and identified host kinases whose probe affinities differed significantly when compared to uninfected cells. Our screen identified 10 novel host kinases that have not been previously shown to be involved with influenza virus replication, and we validated the functional importance of these novel kinases during infection using targeted small interfering RNAs (siRNAs). The effects of kinase-targeted siRNA knockdowns on replicating virus levels were measured by quantitative reverse-transcription PCR and cytoprotection assays. We identified several novel host kinases that, when knocked down, enhanced or reduced the viral load in cell culture. This preliminary work represents the first screen of the changing host kinome in influenza virus-infected human cells.
KW - A549 Cells
KW - Adenosine Diphosphate/chemistry
KW - Adenosine Triphosphate/chemistry
KW - Apoptosis
KW - Biotin/analogs & derivatives
KW - Cell Survival
KW - Chromatography, Liquid
KW - Drug Discovery
KW - Humans
KW - Influenza A Virus, H1N1 Subtype/physiology
KW - Influenza, Human/enzymology
KW - Mass Spectrometry
KW - NIMA-Related Kinase 1/chemistry
KW - Peptides/chemistry
KW - Protein-Serine-Threonine Kinases/chemistry
KW - RNA, Small Interfering/genetics
KW - Reverse Transcriptase Polymerase Chain Reaction
KW - Viral Load
KW - Virus Replication
U2 - 10.1177/1087057113518068
DO - 10.1177/1087057113518068
M3 - Journal article
C2 - 24464431
VL - 19
SP - 936
EP - 946
JO - SLAS Discovery
JF - SLAS Discovery
SN - 2472-5552
IS - 6
ER -
ID: 199431250