title: Distinct Splice Variants of Dynamin-related Protein 1 Differentially Utilize Mitochondrial Fission Factor as an Effector of Cooperative GTPase Activity.
abstract: Multiple isoforms of the mitochondrial fission GTPase dynamin-related protein 1 (Drp1) arise from the alternative splicing of its single gene-encoded pre-mRNA transcript. Among these, the longer Drp1 isoforms, expressed selectively in neurons, bear unique polypeptide sequences within their GTPase and variable domains, known as the A-insert and the B-insert, respectively. Their functions remain unresolved. A comparison of the various biochemical and biophysical properties of the neuronally expressed isoforms with that of the ubiquitously expressed, and shortest, Drp1 isoform (Drp1-short) has revealed the effect of these inserts on Drp1 function. Utilizing various biochemical, biophysical, and cellular approaches, we find that the A- and B-inserts distinctly alter the oligomerization propensity of Drp1 in solution as well as the preferred curvature of helical Drp1 self-assembly on membranes. Consequently, these sequences also suppress Drp1 cooperative GTPase activity. Mitochondrial fission factor (Mff), a tail-anchored membrane protein of the mitochondrial outer membrane that recruits Drp1 to sites of ensuing fission, differentially stimulates the disparate Drp1 isoforms and alleviates the autoinhibitory effect imposed by these sequences on Drp1 function. Moreover, the differential stimulatory effects of Mff on Drp1 isoforms are dependent on the mitochondrial lipid, cardiolipin (CL). Although Mff stimulation of the intrinsically cooperative Drp1-short isoform is relatively modest, CL-independent, and even counter-productive at high CL concentrations, Mff stimulation of the much less cooperative longest Drp1 isoform (Drp1-long) is robust and occurs synergistically with increasing CL content. Thus, membrane-anchored Mff differentially regulates various Drp1 isoforms by functioning as an allosteric effector of cooperative GTPase activity.
authors.affiliation: From the Department of Physiology and Biophysics.
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Macdonald
authors.affiliation: Department of Pharmacology, Center for Mitochondrial Diseases, and Cleveland Center for Membrane and Structural Biology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Francy
authors.affiliation: From the Department of Physiology and Biophysics.
authors.forename: Natalia
authors.initials: N
authors.lastname: Stepanyants
authors.affiliation: From the Department of Physiology and Biophysics.
authors.forename: Lance
authors.initials: L
authors.lastname: Lehman
authors.affiliation: From the Department of Physiology and Biophysics.
authors.forename: Anthony
authors.initials: A
authors.lastname: Baglio
authors.affiliation: Department of Pharmacology, Center for Mitochondrial Diseases, and Cleveland Center for Membrane and Structural Biology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
authors.forename: Jason A
authors.initials: JA
authors.lastname: Mears
authors.affiliation: From the Department of Physiology and Biophysics, Center for Mitochondrial Diseases, and.
authors.forename: Xin
authors.initials: X
authors.lastname: Qi
authors.affiliation: From the Department of Physiology and Biophysics, Cleveland Center for Membrane and Structural Biology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106 rxr275@case.edu.
authors.forename: Rajesh
authors.initials: R
authors.lastname: Ramachandran
mesh_terms: D000818:Animals; D002308:Cardiolipins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D034281:Dynamins / Q000235:genetics* / Q000378:metabolism; D020558:GTP Phosphohydrolases / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism*; D018345:Mice, Knockout; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D012326:RNA Splicing / Q000235:genetics*; D051381:Rats
chemical_list: D002308:Cardiolipins; D008565:Membrane Proteins; C533267:Mff protein, human; D008869:Microtubule-Associated Proteins; D024101:Mitochondrial Proteins; C000603499:mitochondrial fission factor, mouse; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; C110768:DNM1L protein, human; C486391:Dnm1l protein, mouse; D034281:Dynamins
keywords: GTPase; alternative splicing; cardiolipin; dynamin-related protein 1; fluorescence resonance energy transfer (FRET); isoforms; mitochondria; mitochondrial dynamics; mitochondrial fission factor