Jc. Ayoob et al., Targeting of cardiac muscle titin fragments to the Z-bands and dense bodies of living muscle and non-muscle cells, CELL MOTIL, 45(1), 2000, pp. 67-82
A 6.5-kb N-terminal region of embryonic chick cardiac titin, including the
region previously reported as part of the protein zeugmatin, has been seque
nced, further demonstrating that zeugmatin is part of the N-terminal region
of titin, and not a separate Z-band protein. This Z-band region of cardiac
titin, from both 7- and 19-day embryos as well as from adult animals, was
found to contain six different small motifs, termed 2-repeats [Gautel et al
., 1996: J. Cell Sci. 109:2747-2754], of approximately 45 amino acids each
sandwiched between flanking regions containing Ig domains. Fragments of Z-b
and titin, linked to GFP, were expressed in cultured cardiomyocytes to dete
rmine which regions were responsible for Z-band targeting. Transfections of
primary cultures of embryonic chick cardiomyocytes demonstrated that the z
-repeats play the major role in: targeting titin fragments to the Z-band. S
imilar transfections of skeletal myotubes and non-muscle cells lead to the
localization of these cardiac z-repeats in the Z-bands of the myofibrils an
d the dense bodies of the stress fibers. Over-expression of these z-repeat
constructs in either muscle or non-muscle cells lead to the loss of the myo
fibrils or stress fibers, respectively. The transfection experiments also i
ndicated that small domains of a protein, 40 to 50 amino acids, can be stud
ied for their localization properties in living cells if a suitable linker
is placed between these small domains and the much larger 28 kDa GFP protei
n. Cell,Motil.: Cytoskeleton 45.67-82, 2000. (C) 2000 Wiley-Liss, Inc.