title: The ZZ domain of dystrophin in DMD: making sense of missense mutations.
abstract: Duchenne muscular dystrophy (DMD) is associated with the loss of dystrophin, which plays an important role in myofiber integrity via interactions with β-dystroglycan and other members of the transmembrane dystrophin-associated protein complex. The ZZ domain, a cysteine-rich zinc-finger domain near the dystrophin C-terminus, is implicated in forming a stable interaction between dystrophin and β-dystroglycan, but the mechanism of pathogenesis of ZZ missense mutations has remained unclear because not all such mutations have been shown to alter β-dystroglycan binding in previous experimental systems. We engineered three ZZ mutations (p.Cys3313Phe, p.Asp3335His, and p.Cys3340Tyr) into a short construct similar to the Dp71 dystrophin isoform for in vitro and in vivo studies and delineated their effect on protein expression, folding properties, and binding partners. Our results demonstrate two distinct pathogenic mechanisms for ZZ missense mutations. The cysteine mutations result in diminished or absent subsarcolemmal expression because of protein instability, likely due to misfolding. In contrast, the aspartic acid mutation disrupts binding with β-dystroglycan despite an almost normal expression at the membrane, confirming a role for the ZZ domain in β-dystroglycan binding but surprisingly demonstrating that such binding is not required for subsarcolemmal localization of dystrophin, even in the absence of actin binding domains.
authors.affiliation: The Center for Gene Therapy, The Research Institute at Nationwide Children's Hospital, Columbus, Ohio.
authors.forename: Adeline
authors.initials: A
authors.lastname: Vulin
authors.forename: Nicolas
authors.initials: N
authors.lastname: Wein
authors.forename: Dana M
authors.initials: DM
authors.lastname: Strandjord
authors.forename: Eric K
authors.initials: EK
authors.lastname: Johnson
authors.forename: Andrew R
authors.initials: AR
authors.lastname: Findlay
authors.forename: Baijayanta
authors.initials: B
authors.lastname: Maiti
authors.forename: Michael T
authors.initials: MT
authors.lastname: Howard
authors.forename: Yuuki J
authors.initials: YJ
authors.lastname: Kaminoh
authors.forename: Laura E
authors.initials: LE
authors.lastname: Taylor
authors.forename: Tabatha R
authors.initials: TR
authors.lastname: Simmons
authors.forename: Will C
authors.initials: WC
authors.lastname: Ray
authors.forename: Federica
authors.initials: F
authors.lastname: Montanaro
authors.forename: Jim M
authors.initials: JM
authors.lastname: Ervasti
authors.forename: Kevin M
authors.initials: KM
authors.lastname: Flanigan
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D001224:Aspartic Acid / Q000235:genetics; D003545:Cysteine / Q000235:genetics; D049030:Dystroglycans / Q000378:metabolism*; D016189:Dystrophin / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D014644:Genetic Variation; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D020388:Muscular Dystrophy, Duchenne / Q000235:genetics* / Q000473:pathology; D020125:Mutation, Missense; D017510:Protein Folding; D055550:Protein Stability; D016335:Zinc Fingers / Q000235:genetics*
chemical_list: D000199:Actins; D016189:Dystrophin; D049030:Dystroglycans; D001224:Aspartic Acid; D003545:Cysteine
keywords: Duchenne muscular dystrophy; ZZ domain; dystrophin; missense mutation