ABSTRACT
The SRPK family of kinases regulates pre-mRNA splicing by phosphorylating serine/arginine (SR)-rich splicing factors, signals splicing control in response to extracellular stimuli, and contributes to tumorigenesis, suggesting that these splicing kinases are potential therapeutic targets. Here, we report the development of the first irreversible SRPK inhibitor, SRPKIN-1, which is also the first kinase inhibitor that forms a covalent bond with a tyrosine phenol group in the ATP-binding pocket. Kinome-wide profiling demonstrates its selectivity for SRPK1/2, and SRPKIN-1 attenuates SR protein phosphorylation at submicromolar concentrations. Vascular endothelial growth factor (VEGF) is a known target for SRPK-regulated splicing and, relative to the first-generation SRPK inhibitor SRPIN340 or small interfering RNA-mediated SRPK knockdown, SRPKIN-1 is more potent in converting the pro-angiogenic VEGF-A165a to the anti-angiogenic VEGF-A165b isoform and in blocking laser-induced neovascularization in a murine retinal model. These findings encourage further development of SRPK inhibitors for treatment of age-related macular degeneration.
Subject(s)
Angiogenesis Inhibitors/pharmacology , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Vascular Endothelial Growth Factor A/metabolism , Alternative Splicing/drug effects , Angiogenesis Inhibitors/chemistry , Angiogenesis Inhibitors/therapeutic use , Animals , Cell Line , HeLa Cells , Humans , Mice, Inbred C57BL , Molecular Docking Simulation , Neovascularization, Pathologic/drug therapy , Neovascularization, Pathologic/genetics , Neovascularization, Pathologic/metabolism , Protein Isoforms/antagonists & inhibitors , Protein Isoforms/chemistry , Protein Isoforms/metabolism , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/therapeutic use , Protein Serine-Threonine Kinases/chemistry , Protein Serine-Threonine Kinases/metabolism , Vascular Endothelial Growth Factor A/geneticsABSTRACT
Amyloidogenic processing of APP by ß- and γ-secretases leads to the generation of amyloid-ß peptide (Aß), and the accumulation of Aß in senile plaques is a hallmark of Alzheimer's disease (AD). Understanding the mechanisms of APP processing is therefore paramount. Increasing evidence suggests that APP intracellular domain (AICD) interacting proteins influence APP processing. In this study, we characterized the overexpression of AICD interactor GULP1 in a Drosophila AD model expressing human BACE and APP695. Transgenic GULP1 significantly lowered the levels of both Aß1-40 and Aß1-42 without decreasing the BACE and APP695 levels. Overexpression of GULP1 also reduced APP/BACE-mediated retinal degeneration, rescued motor dysfunction and extended longevity of the flies. Our results indicate that GULP1 regulate APP processing and reduce neurotoxicity in a Drosophila AD model.
