title: Dysferlin forms a dimer mediated by the C2 domains and the transmembrane domain in vitro and in living cells.
abstract: Dysferlin was previously identified as a key player in muscle membrane repair and its deficiency leads to the development of muscular dystrophy and cardiomyopathy. However, little is known about the oligomerization of this protein in the plasma membrane. Here we report for the first time that dysferlin forms a dimer in vitro and in living adult skeletal muscle fibers isolated from mice. Endogenous dysferlin from rabbit skeletal muscle exists primarily as a ∼460 kDa species in detergent-solubilized muscle homogenate, as shown by sucrose gradient fractionation, gel filtration and cross-linking assays. Fluorescent protein (YFP) labeled human dysferlin forms a dimer in vitro, as demonstrated by fluorescence correlation spectroscopy (FCS) and photon counting histogram (PCH) analyses. Dysferlin also dimerizes in living cells, as probed by fluorescence resonance energy transfer (FRET). Domain mapping FRET experiments showed that dysferlin dimerization is mediated by its transmembrane domain and by multiple C2 domains. However, C2A did not significantly contribute to dimerization; notably, this is the only C2 domain in dysferlin known to engage in a Ca-dependent interaction with cell membranes. Taken together, the data suggest that Ca-insensitive C2 domains mediate high affinity self-association of dysferlin in a parallel homodimer, leaving the Ca-sensitive C2A domain free to interact with membranes.
authors: Department of Cell and Molecular Physiology, Stritch School of Medicine, Loyola University Medical Center, Maywood, Illinois, United States of America.
authors: Li
authors: L
authors: Xu
authors: Sandeep
authors: S
authors: Pallikkuth
authors: Zhanjia
authors: Z
authors: Hou
authors: Gregory A
authors: GA
authors: Mignery
authors: Seth L
authors: SL
authors: Robia
authors: Renzhi
authors: R
authors: Han
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism*; D002470:Cell Survival; D000073939:Dysferlin; D057809:HEK293 Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism*; D051379:Mice; D008970:Molecular Weight; D018485:Muscle Fibers, Skeletal / Q000378:metabolism; D009124:Muscle Proteins / Q000737:chemistry* / Q000378:metabolism*; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D011817:Rabbits
chemical_list: C114839:DYSF protein, human; D000073939:Dysferlin; D008565:Membrane Proteins; D009124:Muscle Proteins