Identification and characterization of nonmuscle myosin II-C, a new member of the myosin II family.
A previously unrecognized nonmuscle myosin II heavy chain (NMHC II), which constitutes a distinct branch of the nonmuscle/smooth muscle myosin II family, has recently been revealed in genome data bases. We characterized the biochemical properties and expression patterns of this myosin. Using nucleotide probes and affinity-purified antibodies, we found that the distribution of NMHC II-C mRNA and protein (MYH14) is widespread in human and mouse organs but is quantitatively and qualitatively distinct from NMHC II-A and II-B. In contrast to NMHC II-A and II-B, the mRNA level in human fetal tissues is substantially lower than in adult tissues. Immunofluorescence microscopy showed distinct patterns of expression for all three NMHC isoforms. NMHC II-C contains an alternatively spliced exon of 24 nucleotides in loop I at a location analogous to where a spliced exon appears in NMHC II-B and in the smooth muscle myosin heavy chain. However, unlike neuron-specific expression of the NMHC II-B insert, the NMHC II-C inserted isoform has widespread tissue distribution. Baculovirus expression of noninserted and inserted NMHC II-C heavy meromyosin (HMM II-C/HMM II-C1) resulted in significant quantities of expressed protein (mg of protein) for HMM II-C1 but not for HMM II-C. Functional characterization of HMM II-C1 by actin-activated MgATPase activity demonstrated a V(max) of 0.55 + 0.18 s(-1), which was half-maximally activated at an actin concentration of 16.5 + 7.2 microm. HMM II-C1 translocated actin filaments at a rate of 0.05 + 0.011 microm/s in the absence of tropomyosin and at 0.072 + 0.019 microm/s in the presence of tropomyosin in an in vitro motility assay.
Golomb
Eliahu
E
Laboratory of Molecular Cardiology, NHLBI, National Institute of Health, Bethesda, Maryland 20892, USA. golomb@szmc.org.il
Ma
Xuefei
X
Jana
Siddhartha S
SS
Preston
Yvette A
YA
Kawamoto
Sachiyo
S
Shoham
Nitza G
NG
Goldin
Ehud
E
Conti
Mary Anne
MA
Sellers
James R
JR
Adelstein
Robert S
RS
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005091:Exons; D006801:Humans; D015151:Immunoblotting; D051379:Mice; D008969:Molecular Sequence Data; D018995:Myosin Heavy Chains / Q000032:analysis / Q000235:genetics*; D024462:Myosin Type II / Q000032:analysis / Q000235:genetics*; D009928:Organ Specificity; D010802:Phylogeny; D017421:Sequence Analysis
D018995:Myosin Heavy Chains; D024462:Myosin Type II; C477830:MYH7b protein, human; C483905:Myh14 protein, mouse