Nucleotide and protein sequences for dog masticatory tropomyosin identify a novel Tpm4 gene product

Abstract

Jaw-closing muscles of several vertebrate species, including members of Carnivora, express a unique, "masticatory", isoform of myosin heavy chain, along with isoforms of other myofibrillar proteins that are not expressed in most other muscles. It is generally believed that the complement of myofibrillar isoforms in these muscles serves high force generation for capturing live prey, breaking down tough plant material and defensive biting. A unique isoform of tropomyosin (Tpm) was reported to be expressed in cat jaw-closing muscle, based upon two-dimensional gel mobility, peptide mapping, and immunohistochemistry. The objective of this study was to obtain protein and gene sequence information for this unique Tpm isoform. Samples of masseter (also a jawclosing muscle), tibialis (with predominantly fast-twitch fibers), and the deep lateral gastrocnemius (predominantly slow-twitch fibers) were obtained from adult dogs. Expressed Tpm isoforms were cloned and sequencing yielded cDNAs that were identical to genomic predicted striated muscle Tpm1.1 St(a,b,b,a) (historically referred to as αTpm), Tpm2.2 St(a,b,b,a) (βTpm) and Tpm3.12 St(a,b,b,a) (cTpm) isoforms (nomenclature reflects predominant tissue expression ("St"-striated muscle) and exon splicing pattern), as well as a novel 284 amino acid isoform observed in jaw-closing muscle that is identical to a genomic predicted product of the Tpm4 gene (δTpm) family. The novel isoform is designated as Tpm4.3 St(a,b,b,a) . The myofibrillar Tpm isoform expressed in dog masseter exhibits a unique electrophoretic mobility on gels containing 6 M urea, compared to other skeletal Tpm isoforms. To validate that the cloned Tpm4.3 isoform is the Tpm expressed in dog masseter, E. coli-expressed Tpm4.3 was electrophoresed in the presence of urea. Results demonstrate that Tpm4.3 has identical electrophoretic mobility to the unique dog masseter Tpm isoform and is of different mobility from that of muscle Tpm1.1, Tpm2.2 and Tpm3.12 isoforms. We conclude that the unique Tpm isoform in dog masseter is a product of the Tpm4 gene and that the 284 amino acid protein product of this gene represents a novel myofibrillar Tpm isoform never before observed to be expressed in striated muscle.

Desensitizing mouse cardiac troponin C to calcium converts slow muscle towards a fast muscle phenotype

“…The same 22 samples were also loaded on gels (12% acrylamide, 200:1 acrylamide/bis‐acrylamide crosslinking, without glycerol) that were silver‐stained or were transferred for troponin T (TnT) western blots, as previously described (Brundage et al . ). The blots were probed with anti‐slow/cardiac TnT CT3 antibody from the Developmental Studies Hybridoma Bank (DSHB) (DSHB Cat.…”