Effect of cold acclimation on troponin I isoform expression in striated muscle of rainbow trout

Abstract

In vertebrates each of the three striated muscle types (fast skeletal, slow skeletal, and cardiac) contain distinct isoforms of a number of different contractile proteins including troponin I (TnI). The functional characteristics of these proteins have a significant influence on muscle function and contractility. The purpose of this study was to characterize which TnI gene and protein isoforms are expressed in the different muscle types of rainbow trout (Oncorhynchus mykiss) and to determine whether isoform expression changes in response to cold acclimation (4°C). Semiquantitative real-time PCR was used to characterize the expression of seven different TnI genes. The sequence of these genes, cloned from Atlantic salmon (Salmo salar) and rainbow trout, were obtained from the National Center for Biotechnology Information databases. One-dimensional gel electrophoresis and tandem mass spectrometry were used to identify the TnI protein isoforms expressed in each muscle type. Interestingly, the results indicate that each muscle type expresses the gene transcripts of up to seven TnI isoforms. There are significant differences, however, in the expression pattern of these genes between muscle types. In addition, cold acclimation was found to increase the expression of specific gene transcripts in each muscle type. The proteomics analysis demonstrates that fast skeletal and cardiac muscle contain three TnI isoforms, whereas slow skeletal muscle contains four. No other vertebrate muscle to date has been found to express as many TnI protein isoforms. Overall this study underscores the complex molecular composition of teleost striated muscle and suggests there is an adaptive value to the unique TnI profiles of each muscle type.

Dissecting the role of the myofilament in diaphragm dysfunction during the development of heart failure in mice

“…All MS/MS samples were analyzed using Mascot (Matrix Science, London, UK; version Mascot) as in Alderman et al (3). Scaffold (version Scaffold 4.1, Proteome Software, Portland, OR) was used to validate MS/MS based peptide and protein identifications.…”

Cold acclimation increases cardiac myofilament function and ventricular pressure generation in trout

“…The level of phosphorylation of each protein was compared between the thermally acclimated samples via the ratio of the density of the Pro-Q stained bands/actin on the SYPRO-stained gel using ImageJ. We have previously identified cTnT, ssTnT, RLC and TnI using western blotting or tandem mass spectroscopy Alderman et al, 2012). The TnI isoform is not specified as trout heart contains multiple isoforms and these run together in the identified band (Alderman et al, 2012).…”

“…We have previously identified cTnT, ssTnT, RLC and TnI using western blotting or tandem mass spectroscopy Alderman et al, 2012). The TnI isoform is not specified as trout heart contains multiple isoforms and these run together in the identified band (Alderman et al, 2012). Protein loading between lanes was assessed by measuring the density of actin bands visible after SYPRO staining.…”

“…Previous studies have demonstrated that cold acclimation of rainbow trout results in an increase in the maximal rate of cardiac actomyosin ATPase (AM-ATPase) (Yang et al, 2000;Klaiman et al, 2011) and the speed of twitch kinetics of intact fibers (Aho and Vornanen, 1999), indicating changes to the contractile machinery of the heart. Cold acclimation of trout also affects transcript levels of troponin I (Alderman et al, 2012), troponin C (Genge et al, 2013) and sarcoplasmic reticulum Ca 2+ -ATPase (SERCA) (Korajoki and Vornanen, 2012), all of which are key proteins involved in contraction. Korajoki and Vornanen (Korajoki and Vornanen, 2013) have also found that cold acclimation of burbot causes a fourfold increase in the levels of SERCA in the heart, a modification thought to help maintain Ca 2+ cycling at low temperatures.…”

Chronic Hypoxia Exposure of Trout Embryos Alters Swimming Performance and Cardiac Gene Expression in Larvae

“…Transeript abundanees were measured in duplieate reaetions on a StepOne Plus (Life Teehnologies) using default eycling conditions and a dissociation cycle. Each 15-/iL reaction contained 1 x Power SYBR Green Master Mix (Life Technologies), 200 nM each gene-specific primer (Alderman et al 2012), and 1 : 15 vol : vol cDNA. All reactions generated a single-peaked dissociation curve at the predieted amplieon melting temperature.…”