The multiplicity of troponin T isoforms. Distribution in normal rabbit muscles and effects of chronic stimulation.
Polyclonal antibodies were raised in guinea pigs against troponin-T (TnT) isoforms purified from fast- and slow-twitch rabbit muscles. With the use of these antibodies and immunoblots of one- and two-dimensional electrophoreses, the distribution of fast and slow TnT isoforms was investigated in normal and chronically stimulated hindlimb muscles of the rabbit. According to differences in their apparent molecular masses, six fast TnT isoforms (TnTcf, TnT1f, TnT2f, TnT3f, TnT4f, TnT5f) were distinguished in normal tibialis anterior and extensor digitorum longus muscles. These muscles also contained low amounts of TnT1s and TnT2s which were the predominant TnT isoforms in slow-twitch soleus muscle. Fast and slow TnT isoforms were found to exist in several charge variants, i.e. one for TnTcf, three different charge forms for TnT1f, seven for TnT2f, four for TnT3f, three for TnT4f, one for TnT5f, four for TnT1s, and three for TnT2s. Some charge variants were phosphorylated isoforms because treatment with alkaline phosphatase reduced the number of the 19 fast and 7 slow variants to 12 and 3, respectively. The stimulation-induced fast-to-slow transition caused progressive decreases in fast and increases in slow isoforms. The decrease and the disappearance of the major fast isoforms followed a sequence of TnT2f, TnTcf, TnT4f, TnT1f, and TnT3f. This decrease in fast isoforms fits well with the reduction of fast TnT mRNAs assessed by Northern blot analysis. Prolonged stimulation ultimately created a TnT isoform pattern similar to that found in normal slow-twitch muscle. Stimulation also induced changes in the tropomyosin subunit pattern with a decrease in the fast and an increase in the slow alpha-tropomyosin subunit without altering the alpha/beta-tropomyosin subunit ratio. Similar to slow-twitch soleus muscle, long-term stimulated muscles contained appreciable amounts of the fast alpha-tropomyosin subunit, but only traces of fast TnT isoforms. This combination indicated that the predominant slow TnT isoforms may be capable of interacting with fast tropomyosin in these muscles.
Härtner
K T
KT
Fakultät für Biologie, Universität Konstanz, Federal Republic of Germany.
Kirschbaum
B J
BJ
Pette
D
D
D000818:Animals; D000917:Antibody Formation; D004558:Electric Stimulation; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D007536:Isomerism; D008297:Male; D009119:Muscle Contraction; D009132:Muscles / Q000294:innervation / Q000378:metabolism*; D010766:Phosphorylation; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000302:isolation & purification; D011817:Rabbits; D014335:Tropomyosin / Q000302:isolation & purification; D014336:Troponin / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification*; D020107:Troponin T
D012333:RNA, Messenger; D014335:Tropomyosin; D014336:Troponin; D020107:Troponin T