DAPK-1 Bbnding to a linear peptide motif in MAP1B stimulates autophagy and membrane blebbing

Harrison, Ben; Kraus, Michaela; Burch, Lindsay; Stevens, Craig; Craig, Ashley; Gordon-Weeks, Phillip; Hupp, Ted R.

Abstract

DAPK-1 (death-activated protein kinase) has wide ranging functions in cell growth control; however, DAPK-1 interacting proteins that mediate these effects are not well defined. Protein-protein interactions are driven in part by linear interaction motifs, and combinatorial peptide libraries were used to identify peptide interfaces for the kinase domain of DAPK-1. Peptides bound to DAPK-1core kinase domain fragments had homology to the N-terminal domain of the microtubule-associated protein MAP1B. Immunobinding assays demonstrated that DAPK-1 can bind to the full-length human MAP1B, a smaller N-terminal miniprotein containing amino acids 1-126 and the 12-amino acid polypeptides acquired by peptide selection. Amino acid starvation of cells induced a stable immune complex between MAP1B and DAPK-1, identifying a signal that forms the endogenous complex in cells. DAPK-1 and MAP1B co-expression form a synthetic lethal interaction as they cooperate to induce growth inhibition in a clonogenic assay. In cells, two co-localizing populations of DAPK-1 and MAP1B were observed using confocal microscopy; one pool co-localized with MAP1B plus tubulin, and a second pool co-localized with MAP1B plus cortical F-actin. Reduction of MAP1B protein using short interfering RNA attenuated DAPK-1-stimulated autophagy. Transfected MAP1B can synergize with DAPK-1 to stimulate membrane blebbing, whereas reduction of MAP1B using short interfering RNA attenuates DAPK-1 membrane blebbing activity. The autophagy inhibitor 3-methyladenine inhibits the DAPK-1 plus MAP1B-mediated membrane blebbing. These data highlight the utility of peptide aptamers to identify novel binding interfaces and highlight a role for MAP1B in DAPK-1-dependent signaling in autophagy and membrane blebbing.

Citation

Harrison, B., Kraus, M., Burch, L., Stevens, C., Craig, A., Gordon-Weeks, P., & Hupp, T. R. (2008). DAPK-1 Bbnding to a linear peptide motif in MAP1B stimulates autophagy and membrane blebbing. Journal of Biological Chemistry, 283(15), 9999-10014. https://doi.org/10.1074/jbc.M706040200

• Journal Article Type: Article
• Acceptance Date: Jan 2, 2008
• Online Publication Date: Jan 14, 2008
• Publication Date: Apr 11, 2008
• Deposit Date: Aug 2, 2016
• Journal: Journal of Biological Chemistry
• Print ISSN: 0021-9258
• Electronic ISSN: 1083-351X
• Publisher: American Society for Biochemistry and Molecular Biology
• Peer Reviewed: Peer Reviewed
• Volume: 283
• Issue: 15
• Pages: 9999-10014
• DOI: https://doi.org/10.1074/jbc.M706040200
• Keywords: DAPK-1 (death-activated protein kinase), cell growth, Protein-protein interactions,
• Public URL: http://researchrepository.napier.ac.uk/Output/322312