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© The Rockefeller University Press,
0021-9525/2000//653 $5.00
The Journal of Cell Biology, Volume 148, Number 4,
, 2000 653-664
Original Article |
Assembly of Smooth Muscle Myosin by the 38k Protein, a Homologue of a Subunit of Pre-mRNA Splicing Factor-2
gacho{at}sb.gunma-u.ac.jp
Smooth muscle myosin in the dephosphorylated state does not form filaments in vitro. However, thick filaments, which are composed of myosin and myosin-binding protein(s), persist in smooth muscle cells, even if myosin is subjected to the phosphorylation– dephosphorylation cycle. The characterization of telokin as a myosin-assembling protein successfully explained the discrepancy. However, smooth muscle cells that are devoid of telokin have been observed. We expected to find another ubiquitous protein with a similar role, and attempted to purify it from chicken gizzard. The 38k protein bound to both phosphorylated and dephosphorylated myosin to a similar extent. The effect of the myosin-binding activity was to assemble dephosphorylated myosin into filaments, although it had no effect on the phosphorylated myosin. The 38k protein bound to myosin with both COOH-terminal 20 and NH2-terminal 28 residues of the 38k protein being essential for myosin binding. The amino acid sequence of the 38k protein was not homologous to telokin, but to human p32, which was originally found in nuclei as a subunit of pre-mRNA splicing factor-2. Western blotting showed that the protein was expressed in various smooth muscles. Immunofluorescence microscopy with cultured smooth muscle cells revealed colocalization of the 38k protein with myosin and with other cytoskeletal elements. The absence of nuclear immunostaining was discussed in relation to smooth muscle differentiation.
Key Words: myosin assembly • myosin binding • myosin filament • human p32 • smooth muscle
© 2000 The Rockefeller University Press
T. Okagaki's present address is Laboratories of Marine Food Science, Faculty of Bioresources, Mie University, Tsu-city, Mie 514-0102, Japan.Abbreviations used in this paper: C-protein, myosin-binding protein-C; HMM, heavy meromyosin; LMM, light meromyosin; MHC, myosin heavy chain; MLCK, myosin light chain kinase.
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