Tissue expression and actin binding of a novel N-terminal utrophin isoform.
Utrophin and dystrophin present two large proteins that link the intracellular actin cytoskeleton to the extracellular matrix via the C-terminal-associated protein complex. Here we describe a novel short N-terminal isoform of utrophin and its protein product in various rat tissues (N-utro, 62  kDa, amino acids 1-539, comprising the actin-binding domain plus the first two spectrin repeats). Using different N-terminal recombinant utrophin fragments, we show that actin binding exhibits pronounced negative cooperativity (affinity constants K(1) = -5 × 10(6) and K(2) =-1 × 10(5 )M(-1)) and is Ca(2+)-insensitive. Expression of the different fragments in COS7 cells and in myotubes indicates that the actin-binding domain alone binds exclusively to actin filaments. The recombinant N-utro analogue binds in vitro to actin and in the cells associates to the membranes. The results indicate that N-utro may be responsible for the anchoring of the cortical actin cytoskeleton to the membranes in muscle and other tissues.
Department of Endocrinology and Diabetes, University Hospital Zurich, 8091 Zurich, Switzerland.
Richard A
RA
Zuellig
Beat C
BC
Bornhauser
Ralf
R
Amstutz
Bruno
B
Constantin
Marcus C
MC
Schaub
D000199:Actins / Q000378:metabolism*; D000818:Animals; D001681:Biological Assay; D019556:COS Cells; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D002522:Chlorocebus aethiops; D002942:Circular Dichroism; D016189:Dystrophin / Q000737:chemistry / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D015151:Immunoblotting; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000378:metabolism; D009928:Organ Specificity; D011485:Protein Binding; D020033:Protein Isoforms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D049411:Utrophin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D000199:Actins; D002147:Calmodulin; D016189:Dystrophin; D020033:Protein Isoforms; D012333:RNA, Messenger; D011994:Recombinant Proteins; D049411:Utrophin; D002118:Calcium