Initiation and maturation of I-Z-I bodies in the growth tips of transfected myotubes.
While over a dozen I-Z-I proteins are expressed in postmitotic myoblasts and myotubes it is unclear how, when, or where these first assemble into transitory I-Z-I bodies (thin filament/Z-band precursors) and, a short time later, into definitive I-Z-I bands. By double-staining the growth tips of transfected myotubes expressing (a) MYC-tagged s-alpha-actinins (MYC/s-alpha-actinins) or (b) green fluorescent protein-tagged titin cap (GFP/T-cap) with antibodies against MYC and I-Z-I band proteins, we found that the de novo assembly of I-Z-I bodies and their maturation into I-Z-I bands involved relatively concurrent, cooperative binding and reconfiguration of, at a minimum, 5 integral Z-band molecules. These included s-alpha-actinin, nebulin, titin, T-cap and alpha-actin. Resolution of the approximately 1.0 microm polarized alpha-actin/nebulin/tropomyosin/troponin thin filament complexes occurred subsequent to the maturation of Z-bands into a dense tetragonal configuration. Of particular interest is finding that mutant MYC/s-alpha-actinin peptides (a) lacking spectrin-like repeats 1-4, or consisting of spectrin-like repeats 1-4 only, as well as (b) mutants/fragments lacking titin or alpha-actin binding sites, were promptly and exclusively incorporated into de novo assembling I-Z-I bodies and definitive I-Z-I bands as was exogenous full length MYC/s-alpha-actinin or GFP/T-cap.
Ojima
K
K
Department of Physiology and Cell and Developmental Biology, The School of Medicine, University of Pennsylvania, Philadelphia, PA l9l04, USA.
Lin
Z X
ZX
Zhang
Z Q
ZQ
Hijikata
T
T
Holtzer
S
S
Labeit
S
S
Sweeney
H L
HL
Holtzer
H
H
D000185:Actinin / Q000235:genetics / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D002642:Chick Embryo; D064211:Connectin; D018613:Microscopy, Confocal; D018715:Microscopy, Video; D009124:Muscle Proteins / Q000378:metabolism; D018482:Muscle, Skeletal / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D009210:Myofibrils / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D011494:Protein Kinases / Q000378:metabolism; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012518:Sarcomeres / Q000502:physiology* / Q000648:ultrastructure; D013997:Time Factors; D014335:Tropomyosin / Q000378:metabolism; D019210:Troponin I / Q000378:metabolism
D000199:Actins; D064211:Connectin; D009124:Muscle Proteins; D016271:Proto-Oncogene Proteins c-myc; D011993:Recombinant Fusion Proteins; D014335:Tropomyosin; D019210:Troponin I; C035620:nebulin; D000185:Actinin; D011494:Protein Kinases