cAMP-stimulated phosphorylation of diaphanous 1 regulates protein stability and interaction with binding partners in adrenocortical cells

• Type of Material: Article
• Authors: Donghui Li, University of California San Diego; Eric Dammer, Emory University; Natasha C. Lucki, University of California San Diego; Marion B. Sewer, University of California San Diego
• Language: English
• Date: 2013-03-15
• Publisher: American Society for Cell Biology
• Publication Version: Final Published Version
• Copyright Statement: © 2013 Li et al.
• License: Creative Commons Attribution-NonCommercial-ShareAlike 3.0 Unported
• Final Published Version (URL): http://dx.doi.org/10.1091/mbc.E12-08-0597
• Title of Journal or Parent Work: Molecular Biology of the Cell
• ISSN: 1059-1524
• Volume: 24
• Issue: 6
• Start Page: 848
• End Page: 857
• Grant/Funding Information: This work was supported by National Institutes of Health Grant DK094151.
• Keywords: MICROTUBULE STABILIZATION; Science & Technology; TUMOR-CELLS; MIGRATING CELLS; KINASE-A; RHO-MDIA1 PATHWAY; Life Sciences & Biomedicine; FORMIN MDIA2; SMALL GTPASE; Cell Biology; CELL BIOLOGY; LYSOPHOSPHATIDIC ACID; SERUM RESPONSE FACTOR; MTOC REORIENTATION
• Research Categories: Health Sciences, Pharmacology; Health Sciences, Pharmacy

Abstract

Diaphanous homologue 1 (DIAPH1) is a Rho effector protein that coordinates cellular dynamics by regulating microfilament and microtubule function. We previously showed that DIAPH1 plays an integral role in regulating the production of cortisol by controlling the rate of mitochondrial movement, by which activation of the adrenocorticotropin (ACTH)/cAMP signaling pathway stimulates mitochondrial trafficking and promotes the interaction between RhoA and DIAPH1. In the present study we use mass spectrometry to identify DIAPH1 binding partners and find that DIAPH1 interacts with several proteins, including RhoA, dynamin-1, kinesin, β-tubulin, β-actin, oxysterol-binding protein (OSBP)-related protein 2(ORP2), and ORP10. Moreover, DIAPH1 is phosphorylated in response to dibutyryl cAMP (Bt2cAMP) at Thr-759 via a pathway that requires extracellular signal-related kinase (ERK). Alanine substitution of Thr-759 renders DIAPH1 more stable and attenuates the interaction between DIAPH1 and kinesin, ORP2, and actin but has no effect on the ability of the protein to interact with RhoA or β-tubulin. Finally, overexpression of a DIAPH1 T759A mutant significantly decreases the rate of Bt2cAMP-stimulated mitochondrial movement. Taken together, our findings establish a key role for phosphorylation in regulating the stability and function of DIAPH1.