In vivo requirement of the alpha-syntrophin PDZ domain for the sarcolemmal localization of nNOS and aquaporin-4.
alpha-Syntrophin is a scaffolding adapter protein expressed primarily on the sarcolemma of skeletal muscle. The COOH-terminal half of alpha-syntrophin binds to dystrophin and related proteins, leaving the PSD-95, discs-large, ZO-1 (PDZ) domain free to recruit other proteins to the dystrophin complex. We investigated the function of the PDZ domain of alpha-syntrophin in vivo by generating transgenic mouse lines expressing full-length alpha-syntrophin or a mutated alpha-syntrophin lacking the PDZ domain (Delta PDZ). The Delta PDZ alpha-syntrophin displaced endogenous alpha- and beta 1-syntrophin from the sarcolemma and resulted in sarcolemma containing little or no syntrophin PDZ domain. As a consequence, neuronal nitric oxide synthase (nNOS) and aquaporin-4 were absent from the sarcolemma. However, the sarcolemmal expression and distribution of muscle sodium channels, which bind the alpha-syntrophin PDZ domain in vitro, were not altered. Both transgenic mouse lines were bred with an alpha-syntrophin-null mouse which lacks sarcolemmal nNOS and aquaporin-4. The full-length alpha-syntrophin, not the Delta PDZ form, reestablished nNOS and aquaporin-4 at the sarcolemma of these mice. Genetic crosses with the mdx mouse showed that neither transgenic syntrophin could associate with the sarcolemma in the absence of dystrophin. Together, these data show that the sarcolemmal localization of nNOS and aquaporin-4 in vivo depends on the presence of a dystrophin-bound alpha-syntrophin PDZ domain.
Adams
M E
ME
Department of Physiology and Biophysics, University of Washington, Seattle, WA 98195, USA.
Mueller
H A
HA
Froehner
S C
SC
D000818:Animals; D051401:Aquaporin 4; D020346:Aquaporins / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000378:metabolism; D016189:Dystrophin / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D009124:Muscle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D020033:Protein Isoforms / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012508:Sarcolemma / Q000201:enzymology / Q000378:metabolism*; D015222:Sodium Channels / Q000378:metabolism
C496031:Aqp4 protein, mouse; D051401:Aquaporin 4; D020346:Aquaporins; D002135:Calcium-Binding Proteins; D016189:Dystrophin; D008565:Membrane Proteins; D009124:Muscle Proteins; D020033:Protein Isoforms; D015222:Sodium Channels; C091825:syntrophin alpha1; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; C496321:Nos1 protein, mouse