Myofibrillar gene expression in differentiating lobster claw muscles

Medler, Scott; Lilley, Travis R.; Riehl, Jocelyn H.; Mulder, Eva P.; Chang, Ernest S.; Mykles, Donald L.

doi:10.1002/jez.375

Cited by 11 publications

(13 citation statements)

References 38 publications

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“…Blots were probed for TnT, TnI or P75 using polyclonal antibodies raised in rabbits against these proteins isolated from lobster muscles (Medler and Mykles, 2003;Medler et al, 2004;Medler et al, 2007). After transfer of proteins, membranes were washed in deionized water and then allowed to air dry and stored for up to several days before probing with antibodies.…”

Section: Analysis Of Myofibrillar Proteins By Sds Page and Western Blmentioning

confidence: 99%

“…We propose that subtle shifts in contractile properties may stem from the continuum of myofibrillar assemblage known to exist in crustacean muscle fibers Mykles et al, 2002). Developing lobster claw muscles exhibit a gradual shift in myofibrillar isoform expression, as the muscles differentiate into specialized fast and slow fibers over a period of months (Medler et al, 2007). Further studies are needed to understand precisely how these differences influence the running capability of ghost crabs.…”

Section: Scale Effects and Muscle Organizationmentioning

confidence: 99%

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Skeletal muscle fiber types in the ghost crab,Ocypode quadrata:implications for running performance

Perry

Tait

et al. 2009

Journal of Experimental Biology

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SUMMARYGhost crabs possess rapid running capabilities, which make them good candidates for comparing invertebrate exercise physiology with that of more extensively studied vertebrates. While a number of studies have examined various aspects of running physiology and biomechanics in terrestrial crabs, none to date have defined the basic skeletal muscle fiber types that power locomotion. In the current study, we investigated skeletal muscle fiber types comprising the extensor and flexor carpopodite muscles in relation to running performance in the ghost crab. We used kinematic analyses to determine stride frequency and muscle shortening velocity and found that both parameters are similar to those of comparably sized mammals but slower than those observed in running lizards. Using several complementary methods, we found that the muscles are divided into two primary fiber types: those of the proximal and distal regions possess long sarcomeres (6.2±2.3 μm) observed in crustacean slow fibers and have characteristics of aerobic fibers whereas those of the muscle mid-region have short sarcomeres (3.5±0.4 μm) characteristic of fast fibers and appear to be glycolytic. Each fiber type is characterized by several different myofibrillar protein isoforms including multiple isoforms of myosin heavy chain (MHC), troponin I (TnI), troponin T (TnT) and a crustacean fast muscle protein, P75. Three different isoforms of MHC are differentially expressed in the muscles, with fibers of the mid-region always coexpressing two isoforms at a 1:1 ratio within single fibers. Based on our analyses, we propose that these muscles are functionally divided into a two-geared system, with the aerobic fibers used for slow sustained activities and the glycolytic mid-region fibers being reserved for explosive sprints. Finally, we identified subtle differences in myofibrillar isoform expression correlated with crab body size, which changes by several orders of magnitude during an animalʼs lifetime.

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Section: Analysis Of Myofibrillar Proteins By Sds Page and Western Blmentioning

confidence: 99%

Section: Scale Effects and Muscle Organizationmentioning

confidence: 99%

Skeletal muscle fiber types in the ghost crab,Ocypode quadrata:implications for running performance

Perry

Tait

et al. 2009

Journal of Experimental Biology

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“…PCR conditions were the same as reported by Yu and Mykles (2003). Whole cutter and crusher claws from 9 th and 10 th stage juvenile lobsters were homogenized and used for RNA isolation (Medler et al, 2007). Log-transformed values were used to compare expression levels in cutter and crusher claws using a paired T-test.…”

Section: Real-time Pcr Of Ha-calpmmentioning

confidence: 99%

Muscle-specific calpain is localized in regions near motor endplates in differentiating lobster claw muscles

Medler¹,

Chang²,

Mykles³

2007

Comparative Biochemistry and Physiology Part A: Molecular &

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Calpains are Ca 2+ -dependent proteinases that mediate protein turnover in crustacean skeletal muscles. We used an antibody directed against lobster muscle-specific calpain (Ha-CalpM) to examine its distribution in differentiating juvenile lobster claw muscles. These muscles are comprised of both fast and slow fibers early in development, but become specialized into predominantly fast or exclusively slow muscles in adults. The transition into adult muscle types requires that myofibrillar proteins specific for fast or slow muscles to be selectively removed and replaced by the appropriate proteins. Using immunohistochemistry, we observed a distinct staining pattern where staining was preferentially localized in the fiber periphery along one side of the fiber. Immunolabeling with an antibody directed against synaptotagmin revealed that the calpain staining was greatest in the cytoplasm adjacent to synaptic terminals. In complementary analyses, we used sequence-specific primers with real-time PCR to quantify the levels of Ha-CalpM in whole juvenile claw muscles. These expression levels were not significantly different between cutter and crusher claws, but were positively correlated with the expression of fast myosin heavy chain. The anatomical localization of Ha-CalpM near motor endplates, coupled with the correlation with fast myofibrillar gene expression, suggests a role for this intracellular proteinase in fiber type switching.

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“…The homogenates were centrifuged at 16,000g for 20min at 4°C. The soluble protein in the supernatant fraction was dialyzed against two 1-l changes of 20mmoll -1 ammonium acetate, frozen, lyophilized, and dissolved in 1-2 ml SDS sample buffer (Medler et al, 2007) at 60°C for 10-15min. The myofibrillar protein in the pellet obtained from centrifugation was washed three times with 40ml buffer A and extracted with five volumes 0.6moll -1 NaCl and 5mmoll -1 sodium phosphate (pH7.4) at 4°C for 30min.…”

Section: Organ Culturementioning

confidence: 99%

“…Incorporation was expressed as nmol [ . For autoradiography, proteins were separated using 10% SDS-PAGE (Medler et al, 2007) and exposed to X-ray film at -80°C.…”

Section: Organ Culturementioning

confidence: 99%

Molt cycle regulation of protein synthesis in skeletal muscle of the blackback land crab, Gecarcinus lateralis, and the differential expression of a myostatin-like factor during atrophy induced by molting or unweighting

Covi

Bader

Chang

et al. 2010

Journal of Experimental Biology

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SUMMARYIn decapod crustaceans, claw muscle undergoes atrophy in response to elevated ecdysteroids while thoracic muscle undergoes atrophy in response to unweighting. The signaling pathways that regulate muscle atrophy in crustaceans are largely unknown. Myostatin is a negative regulator of muscle growth in mammals, and a myostatin-like cDNA is preferentially expressed in muscle of the land crab, Gecarcinus lateralis (Gl-Mstn). Contrary to prediction, levels of Gl-Mstn mRNA decreased dramatically in both the claw closer and weighted thoracic muscles when molting was induced by either eyestalk ablation (ESA) or multiple limb autotomy (MLA). However, the effect of molt induction was greater in the claw muscle. By late premolt, Gl-Mstn mRNA in the claw muscle decreased 81% and 94% in ESA and MLA animals, respectively, and was negatively correlated with ecdysteroids. Gl-Mstn mRNA in thoracic muscle decreased 68% and 82% in ESA and MLA animals, respectively, but was only weakly correlated with ecdysteroid. Claw and thoracic muscles also differed to varying extents in the expression of ecdysteroid receptor (Gl-EcR and Gl-RXR), elongation factor-2 (Gl-EF-2), and calpain T (Gl-CalpT) in response to molt induction, but levels of the four transcripts were not correlated with ecdysteroid. The downregulation of Gl-Mstn expression in premolt claw muscle coincided with 11-and 13-fold increases in protein synthesis in the myofibrillar and soluble protein fractions, respectively. Furthermore, the rate of the increase in the synthesis of soluble proteins was greater than that of myofibrillar proteins during early premolt (1.4:1, soluble:myofibrillar), but the two were equivalent during late premolt. By contrast, Gl-Mstn mRNA increased 3-fold and Gl-CalpT mRNA decreased 40% in unweighted thoracic muscle; there was little or no effect on Gl-EF-2, Gl-EcR, and Gl-RXR mRNA levels. These data indicate that Gl-Mstn expression is negatively regulated by both ecdysteroids and load-bearing contractile activity. The downregulation of GlMstn in claw muscle may induce the elevated protein turnover associated with remodeling of the contractile apparatus during molt-induced atrophy. The upregulation of Gl-Mstn in unweighted thoracic muscle suggests that this factor is also involved in disuse atrophy when hemolymph ecdysteroid levels are low.

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Myofibrillar gene expression in differentiating lobster claw muscles

Cited by 11 publications

References 38 publications

Skeletal muscle fiber types in the ghost crab,Ocypode quadrata:implications for running performance

Skeletal muscle fiber types in the ghost crab,Ocypode quadrata:implications for running performance

Muscle-specific calpain is localized in regions near motor endplates in differentiating lobster claw muscles

Molt cycle regulation of protein synthesis in skeletal muscle of the blackback land crab, Gecarcinus lateralis, and the differential expression of a myostatin-like factor during atrophy induced by molting or unweighting

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