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dc.contributor.authorZhou, Daixing
dc.date.accessioned2012-04-20T18:45:16Z
dc.date.available2012-04-20T18:45:16Z
dc.date.issued1995
dc.identifier.urihttp://hdl.handle.net/10713/1460
dc.descriptionUniversity of Maryland, Baltimore. Physiology. Ph.D. 1995en_US
dc.description.abstractMembers of the families of structural proteins, called spectrins and ankyrins, are present on the cytoplasmic surface of the sarcolemma of vertebrate skeletal muscle fibers. I studied these proteins in order to identify new spectrin binding proteins, to determine how the spectrins and ankyrins are assembled into a subsarcolemmal skeleton during development, and to learn how different alternatively spliced forms are disposed in developing and adult skeletal muscle fibers. To identify {dollar}\beta{dollar}-spectrin-binding proteins, I constructed a cDNA expression library from rat skeletal muscle mRNA. Using I{dollar}\sp{lcub}125{rcub}{dollar}-labeled {dollar}\beta{dollar}-spectrin to screen the library, I identified several clones, all of which appear to derive from the {dollar}\alpha{dollar}-fodrin gene. The sequence obtained from these clones encodes a protein of 2,473 amino acids. When compared to {dollar}\alpha{dollar}-fodrins cloned from chicken brain and human fibroblasts, {dollar}\alpha{dollar}-fodrin was found to be highly conserved, sharing 96% identity at the amino acid level. Bacterial expression of a portion of {dollar}\alpha{dollar}-fodrin and in vitro gel overlay assay confirmed the binding between {dollar}\alpha{dollar}-fodrin and {dollar}\beta{dollar}-spectrin. Skeletal muscle contained no {dollar}\alpha{dollar}-spectrin by Northern blot analysis. {dollar}\beta{dollar}-Fodrin mRNA, which was present, stayed at constant levels in skeletal muscle tissue from embryo to adult. {dollar}\alpha{dollar}-Fodrin expression increased with age, with a significant increase occurring in the early postnatal period. The 11.0 kb transcript of {dollar}\beta{dollar}-spectrin also increased with development, especially after birth. {dollar}\beta{dollar}-fodrin was present at the sarcolemma of muscle cells and assembled into membrane skeletal structures in late embryonic muscle, but decreased in muscle fibers after birth and was absent from the sarcolemma of adult myofibers. {dollar}\beta{dollar}-Spectrin was expressed at the membrane of muscle cells at all ages, but was especially prevalent in adult fibers, where it appeared to replace {dollar}\beta{dollar}-fodrin. It is localized to membrane skeletal structures at all stages of development. {dollar}\alpha{dollar}-Fodrin was also present in the membrane skeleton at all stages of development. In embryonic muscle, which contained both spectrin and fodrin, the three subunits appeared to co-assemble at the membrane, whereas in adult, some regions of the membrane skeleton contained both {dollar}\alpha{dollar}-fodrin and {dollar}\beta{dollar}-spectrin, while others contained only {dollar}\beta{dollar}-spectrin. I also studied ankyrin, a major spectrin binding protein, in skeletal muscle. Northern blot analysis revealed three small ankyrin transcripts besides the 7.5 and 9.0 kb transcripts known to encode full-length ankyrin. The small transcripts in fact were the predominant forms in skeletal muscle. Their sizes were 1.6, 2.0 and 3.5 kb, which are too small to encode full-length ankyrin protein. Cloning work, done by a collaborating laboratory, demonstrated they probably encoded 1-3 proteins of {dollar}<{dollar}22 kDa but differed in their 3{dollar}\sp\prime{dollar}UTR. Antibodies specific for the small ankyrins made by these short transcripts showed them to be located at the Z line and M line of skeletal muscle, in contrast to the full-length ankyrin, which was present only at the sarcolemma. The expression of small ankyrin is unaffected in the nb/nb mouse, which lacks full length ankyrin. The small ankyrins are highly enriched in purified fractions of sarcoplasmic reticulum. (Abstract shortened by UMI.)en_US
dc.language.isoen_USen_US
dc.subjectBiology, Molecularen_US
dc.subjectBiology, Cellen_US
dc.subject.meshAnkyrinsen_US
dc.subject.meshMuscle, Skeletalen_US
dc.subject.meshSpectrinen_US
dc.titleSpectrins and ankyrins in skeletal muscleen_US
dc.typedissertationen_US
dc.contributor.advisorBloch, Robert J.
dc.identifier.ispublishedYes
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