Evidence for inserted sequences in the head region of nonmuscle myosin specific to the nervous system. Cloning of the cDNA encoding the myosin heavy chain-B isoform of vertebrate nonmuscle myosin.
The complete amino acid sequence of a vertebrate nonmuscle myosin heavy chain-B isoform (MHC-B, 1976 amino acids, 229 kDa) has been deduced by using cDNA clones from chicken brain libraries. The chicken nonmuscle MHC-B shows overall similarity in primary structure to other MHCs in the areas contributing to the ATP-binding site and actin-binding site. Similar to other nonsarcomeric MHC IIs, there is a short uncoiled tail sequence at the carboxyl terminus of the molecule. It is in the uncoiled tail sequence that the greatest number of differences in amino acids sequence between MHC-A and B were found, which allowed generation of isoform-specific antibodies. These antibodies were used to determine the relative content of MHC-A and MHC-B in various tissues. During the cloning of the cDNA encoding chicken brain MHC-B, we found a 63-nucleotide insertion encoding 21 amino acids located in the head region of the MHC near to the actin-binding site and a 30 nucleotide insertion encoding 10 amino acids near to the ATP-binding site. Analysis using S-1 nuclease showed that both inserts are expressed in a tissue-dependent manner; mRNA containing the inserts is present in tissues of the nervous system, but is absent from other non-muscle cells, which contain the noninserted isoform of MHC-B. Similar inserts were found in corresponding positions in human cerebellar mRNA. Antibodies raised against a peptide synthesized based on the 21 amino acid insert found in chickens recognize a MHC isoform in the same tissues that are enriched for the mRNA. These insertions appear to be a mechanism for generating additional MHC-B isoforms specific to the nervous system.
Laboratory of Molecular Cardiology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
M
M
Takahashi
S
S
Kawamoto
R S
RS
Adelstein
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D001483:Base Sequence; D002531:Cerebellum / Q000502:physiology*; D002645:Chickens; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D008969:Molecular Sequence Data; D009132:Muscles / Q000502:physiology; D009218:Myosins / Q000235:genetics*; D009928:Organ Specificity; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D011061:Poly A / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid
D000906:Antibodies; D010455:Peptides; D012333:RNA, Messenger; D011061:Poly A; D004247:DNA; D009218:Myosins