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Phosphorylation by protein kinase A disassembles the caspase-9 core

Cell Death & Differentiationvolume 25pages10251039 (2018) | Download Citation


Caspases, the cysteine proteases which facilitate the faithful execution of apoptosis, are tightly regulated by a number of mechanisms including phosphorylation. In response to cAMP, PKA phosphorylates caspase-9 at three sites preventing caspase-9 activation, and suppressing apoptosis progression. Phosphorylation of caspase-9 by PKA at the functionally relevant site Ser-183 acts as an upstream block of the apoptotic cascade, directly inactivating caspase-9 by a two-stage mechanism. First, Ser-183 phosphorylation prevents caspase-9 self-processing and directly blocks substrate binding. In addition, Ser-183 phosphorylation breaks the fundamental interactions within the caspase-9 core, promoting disassembly of the large and small subunits. This occurs despite Ser-183 being a surface residue distal from the interface between the large and small subunits. This phosphorylation-induced disassembly promotes the formation of ordered aggregates around 20 nm in diameter. Similar aggregates of caspase-9 have not been previously reported. This two-stage regulatory mechanism for caspase-9 has likewise not been reported previously but may be conserved across the caspases.

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This work was supported by the National Institutes of Health (GM 080532) to JH. BS was supported in part by the UMass Chemistry-Biology Interface Training Program (National Research Service Award T32 GM 08515 from the National Institutes of Health). We thank Jesse Rinehart (Yale University) for generously providing the E. coli strain C321.ΔA, the pBAD-GST-AmpR and SepOTSλ plasmids and for advice and helpful discussions about phosphoprotein synthesis. We thank Alex Ribbe, Director of the UMass Electron Microscopy facility for the collection of EM images. We thank Tyler Marcinko for his assistance with staining of protein samples and ThT assay. We also thank Scott Eron for providing casp-8 proteins as samples for caspase cleavage assays.

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  1. Department of Chemistry, University of Massachusetts, 104 LGRT, 710 N. Pleasant Street, Amherst, MA, 01003, USA

    • Banyuhay P. Serrano
    •  & Jeanne A. Hardy


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Correspondence to Jeanne A. Hardy.

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