title: Actin filaments as dynamic reservoirs for Drp1 recruitment.
abstract: Drp1 is a dynamin-family GTPase recruited to mitochondria and peroxisomes, where it oligomerizes and drives membrane fission. Regulation of mitochondrial Drp1 recruitment is not fully understood. We previously showed that Drp1 binds actin filaments directly, and actin polymerization is necessary for mitochondrial Drp1 oligomerization in mammals. Here we show the Drp1/actin interaction displays unusual properties that are influenced by several factors. At saturation, only a fraction Drp1 binds actin filaments, and the off-rate of actin-bound Drp1 is significantly increased by unbound Drp1. GDP and GTP accelerate and decelerate Drp1/actin binding dynamics, respectively. Actin has a biphasic effect on Drp1 GTP hydrolysis, increasing at low actin:Drp1 ratio but returning to baseline at high ratio. Drp1 also bundles filaments. Bundles have reduced dynamics but follow the same trends as single filaments. Drp1 preferentially incorporates into bundles at higher ionic strength. We measure Drp1 concentration to be ∼0.5 μM in U2OS cell cytosol, suggesting the actin-binding affinity measured here (Kd= 0.6 μM) is in the physiologically relevant range. The ability of Drp1 to bind actin filaments in a highly dynamic manner provides potential for actin filaments to serve as reservoirs of oligomerization-competent Drp1 that can be accessed for mitochondrial fission.
authors: Department of Biochemistry and Cell Biology, Geisel School of Medicine at Dartmouth College, Hanover, NH 03755.
authors: Anna L
authors: AL
authors: Hatch
authors: Department of Biochemistry and Cell Biology, Geisel School of Medicine at Dartmouth College, Hanover, NH 03755.
authors: Wei-Ke
authors: WK
authors: Ji
authors: Department of Pharmacology, Carver School of Medicine, University of Iowa, Iowa City, IA 52242.
authors: Ronald A
authors: RA
authors: Merrill
authors: Department of Pharmacology, Carver School of Medicine, University of Iowa, Iowa City, IA 52242.
authors: Stefan
authors: S
authors: Strack
authors: Department of Biochemistry and Cell Biology, Geisel School of Medicine at Dartmouth College, Hanover, NH 03755 henry.higgs@dartmouth.edu.
authors: Henry N
authors: HN
authors: Higgs
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000818:Animals; D018929:Cell Culture Techniques; D003600:Cytosol / Q000378:metabolism; D034281:Dynamins; D020558:GTP Phosphohydrolases / Q000378:metabolism* / Q000493:pharmacokinetics / Q000502:physiology; D006801:Humans; D006868:Hydrolysis; D008869:Microtubule-Associated Proteins / Q000378:metabolism* / Q000493:pharmacokinetics / Q000502:physiology; D008928:Mitochondria / Q000378:metabolism; D063154:Mitochondrial Dynamics / Q000502:physiology; D024101:Mitochondrial Proteins / Q000378:metabolism* / Q000493:pharmacokinetics / Q000502:physiology; D020675:Peroxisomes / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization
chemical_list: D000199:Actins; D008869:Microtubule-Associated Proteins; D024101:Mitochondrial Proteins; D020558:GTP Phosphohydrolases; C110768:DNM1L protein, human; D034281:Dynamins