Quantification of myosin heavy chain mRNA in somatic and branchial arch muscles using competitive PCR.
The purpose of this study was to quantify the type and amount of myosin heavy chain (MHC) mRNA within muscles of different developmental origins to determine whether the regulation of gene expression is comparable. Seven MHC isoforms were analyzed in rat adult limb (extensor digitorum longus, tibialis anterior, and soleus) and nonlimb (extraocular, thyroarytenoid, diaphragm, and masseter) muscles using a competitive PCR assay. An exogenous template that included oligonucleotide sequences specific for seven rat sarcomeric MHC isoforms (beta-cardiac, 2A, 2X, 2B, extraocular, embryonic, and neonatal) as well as beta-actin was constructed and used as the competitor. Only the extraocular muscle contained all seven isoforms. All seven muscles contained type 2A and type 2X MHC transcripts in varying percentages. As expected, the soleus muscle contained primarily beta-cardiac MHC (87.8 +/- 2.6%). Extraocular MHC was found only in the extraocular and thyroarytenoid muscles and in relatively small proportions (7.4 +/- 1.5% and 4.0 +/- 0.7%, respectively). Neonatal MHC was identified in extraocular (7.9 +/- 0. 3%), thyroarytenoid (4.4 +/- 0.4%), and masseter (1.0 +/- 0.2%) muscles, and embryonic MHC was identified both in extraocular (1.2 +/- 0.5%) and, unexpectedly, in soleus (0.6 +/- 0.1%) muscles. Absolute MHC mRNA mass was greatest in the masseter (106 pg/0.5 microg RNA) and least for the tibialis anterior (64 pg/0.5 microg RNA). These values suggest that MHC mRNA represents from 4 to 17% of the total mRNA pool in various skeletal muscles. Differences in MHC profile between somatic and branchial arch muscles suggest that the developmental origin of a muscle may, at least in part, be responsible for the MHC expression program that is implemented in the adult. An inverse relationship between the expression of beta-cardiac and type 2B MHC transcripts across muscles was noted, suggesting that the expression of these two isoforms may be reciprocally regulated.
Department of Surgery/Otolaryngology, University of California at San Diego School of Medicine and Veterans Affairs Medical Center, La Jolla, California 92093, USA.
H H
HH
Jung
R L
RL
Lieber
A F
AF
Ryan
D000375:Aging / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D001483:Base Sequence; D001934:Branchial Region / Q000378:metabolism*; D017931:DNA Primers; D004622:Embryo, Mammalian; D018507:Gene Expression Regulation, Developmental; D024510:Muscle Development; D018482:Muscle, Skeletal / Q000254:growth & development / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000096:biosynthesis*; D016376:Oligonucleotides, Antisense; D009928:Organ Specificity; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000032:analysis* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014158:Transcription, Genetic
D017931:DNA Primers; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D018995:Myosin Heavy Chains