Expression of a novel tropomyosin isoform in axolotl heart and skeletal muscle

Thomas, Anish; Rajan, Sudarsan; Thurston, Harold; Masineni, Sreeharsha N.; Dube, Preeti; Bose, Abhishek; Muthu, Vasundhara; Dube, Syamalima; Wieczorek, David F.; Poiesz, Bernard J.; Dube, Dipak K.

doi:10.1002/jcb.22599

Cited by 26 publications

(54 citation statements)

References 16 publications

(23 reference statements)

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“…On the other hand the relative expression of TPM1κ is higher in the heart. The results are consistent with our reported findings in mice (Dube et al, 2014), and axolotl (Thomas et al, 2010). However, when we compared the absolute copy number expression in heart and skeletal muscle, the expression of TPM1α compared to TPM1κ is much higher (~300 fold) in heart.…”

Section: Figsupporting

confidence: 92%

“…However, the functional role of TPM1κ is yet to be established. It is known that TPM1κ is ~5% of the total sarcomeric tropomyosin protein in human hearts, which is also true for axolotl (Thomas et al, 2010). We already reported that an anti-sense mediated down regulation of TPM1κ inhibited the cardiac contractility in axolotl in situ suggesting a critical role of TPM1κ protein in cardiac contractility.…”

Section: Figmentioning

confidence: 79%

“…For each sample in a total reaction mixture of 30 μl was made, 5 μl protein sample, 7.5 μl NuPage LDS sample buffer (4x), 15.5 μl distilled water and 2 μl NuPage sample reducing agent (10x). The samples were run on NuPage 4-12% Bis Tris gel and then transferred to nitrocellulose membrane according to the manufacturer's instruction as described earlier (Thomas et al, 2010). The membrane was stained with Ponceau S solution in order to verify the transfer of the protein onto the membrane itself.…”

Section: Western Blotmentioning

confidence: 99%

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Expression of various sarcomeric tropomyosin isoforms in equine striated muscles

et al. 2017

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In order to better understand the training and athletic activity of horses, we must have complete understanding of the isoform diversity of various myofibrillar protein genes like tropomyosin. Tropomyosin (TPM), a coiled-coil dimeric protein, is a component of thin filament in striated muscles. In mammals, four TPM genes (TPM1, TPM2, TPM3, and TPM4) generate a multitude of TPM isoforms via alternate splicing and/or using different promoters. Unfortunately, our knowledge of TPM isoform diversity in the horse is very limited. Hence, we undertook a comprehensive exploratory study of various TPM isoforms from horse heart and skeletal muscle. We have cloned and sequenced two sarcomeric isoforms of the TPM1 gene called TPM1α and TPM1κ, one sarcomeric isoform of the TPM2 and one of the TPM3 gene, TPM2α and TPM3α respectively. By qRT-PCR using both relative expression and copy number, we have shown that TPM1α expression compared to TPM1κ is very high in heart. On the other hand, the expression of TPM1α is higher in skeletal muscle compared to heart. Further, the expression of TPM2α and TPM3α are higher in skeletal muscle compared to heart. Using western blot analyses with CH1 monoclonal antibody we have shown the high expression levels of sarcomeric TPM proteins in cardiac and skeletal muscle. Due to the paucity of isoform specific antibodies we cannot specifically detect the expression of TPM1κ in horse striated muscle. To the best of our knowledge this is the very first report on the characterization of sarcmeric TPMs in horse striated muscle.

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Section: Figsupporting

confidence: 92%

Section: Figmentioning

confidence: 79%

Section: Western Blotmentioning

confidence: 99%

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Expression of various sarcomeric tropomyosin isoforms in equine striated muscles

et al. 2017

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“…The copy number per volume of clone in solution was determined using the equation number of copies = (ng of plasmid DNA × 6.02 × 10 23 )/(bp length of plasmid × 1 × 10 9 × 650), which was simplified by Andrew Staroscik at the URI Genomics and Sequencing Center. A dilution series of each clone was done for 1 × 10 1 −1 × 10 4 copies of template, which was used to create a standard curve after amplification (Figure 6) (Dube et al, , ; Thomas et al, ; Thurston et al, ). For better accuracy, each sample in the dilution series was run in triplicate.…”

Section: Methodsmentioning

confidence: 99%

Sarcomeric TPM3α in developing chicken

et al. 2017

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Cloning and sequencing of various tropomyosin isoforms expressed in chickens have been described since the early 1980s. However, to the best of our knowledge, this is the first report on the molecular characterization and the expression of the sarcomeric isoform of the TPM3 gene in cardiac and skeletal muscles from developing as well as adult chickens. Expression of TPM3α was performed by conventional RT-PCR as well as qRT-PCR using relative expression (by ΔC as well as ΔΔC methods) and by determining absolute copy number. The results employing all these methods show that the expression level of TPM3α is maximum in embryonic (10-day/15-day old) skeletal muscle and can barely be detected in both cardiac and skeletal muscles from the adult chicken. Our various RT-PCR analyses suggest that the expression of high molecular weight TPM3 isoforms are regulated at the transcription level from the proximal promoter at the 5'-end of the TPM3 gene.

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“…Axolotls are commonly employed in biomedical research due to their ability to quickly repair damaged tissues (Voss and Shaffer, 1997;Wakahara, 1996). Indeed, most studies of the axolotl heart focus on its ability to regenerate after partial ventricular amputation (Cano-Martínez et al, 2010) or on sarcomeric proteins mutations (Thomas et al, 2010;Ward et al, 1995). Given their relevance for biomedical research, it is surprising that very little is known about the intrinsic contractile properties of the axolotl heart.…”

Section: Introductionmentioning

confidence: 97%