Fluorescent Biosensor of CDK5 Kinase Activity in Glioblastoma Cell Extracts and Living Cells. Issue 9 (3rd June 2020) - Peyressatre, Marion
Abstract: CDK5 plays a major role in neuronal functions, and is hyperactivated in neurodegenerative pathologies as well as in glioblastoma and neuroblastoma. Although this kinase constitutes an established biomarker and pharmacological target, there are few means of probing its activity in cell extracts or in living cells. To this aim a fluorescent peptide reporter of CDK5 kinase activity, derived from a library of CDK5‐specific substrates, is engineered and its ability to respond to recombinant CDK5/p25 is established and CDK5 activity in glioblastoma cell extracts is reported on through sensitive changes in fluorescence intensity. A cell‐penetrating variant of this biosensor which can be implemented to image CDK5 activation dynamics in space and in time is further implemented. This original biosensor constitutes a potent tool for quantifying differences in CDK5 activity following treatment with selective inhibitors and for monitoring CDK5 activation, following inhibition or stimulation, in a physiologically relevant environment. As such it offers attractive opportunities to develop a diagnostic assay for neuronal pathologies associated with hyperactivated CDK5, as well as a companion assay to evaluate response to new therapies targeting this kinase. Abstract : CDK5 kinase is hyperactivated in neurodegenerative pathologies and glioblastoma. In this study, a fluorescent peptide biosensor that reports on CDK5 activity in vitro and a self‐cell‐penetrating derivative for imaging CDK5 activation in living cells are engineered. This original biosensor constitutes a potent tool for quantifying differences in CDK5 activity, thereby offering attractive opportunities to monitor hyperactivation of this kinase in neuronal pathologies and cancers.