Moulting their shells and misplacing limbs, crabs continually depend on their versatile musculature to reshape their bodies. According to Donald Mykles and colleagues from Colorado State University and the University of Wisconsin – Stevens Point, USA, the massive claw muscle of blackback land crabs withers by as much as 78% before they can slip their shells off. However, Mykles and his colleagues made an interesting discovery in 2010. Instead of shutting down protein synthesis to allow the muscle to wither, the crabs ramp it up 11-fold by shutting down myostatin expression – which usually keeps protein synthesis in check. Knowing that myostatin regulates protein expression in mammals via the mTOR (metazoan target of rapamycin) signalling cascade, which controls mRNA translation, Mykles decided to investigate the mTOR pathway in blackback land crabs to find out whether it also regulates muscle rebuilding in crabs during moulting (p. 590).
Stimulating crabs to moult, the team collected muscles from the animals and searched for the mRNA of four key mTOR signalling proteins – Rheb, mTOR, Akt and s6k. The crabs expressed all four genes, confirming that the mTOR pathway is active in moulting crab’s atrophied muscle.
Next, the team measured the expression levels of each gene in crabs that had been induced to moult using methods that raise the animals’ moulting hormone (ecdysteroid, which regulates myostatin levels) over different time scales. They found that Rheb levels increased in the moulting crabs’ claws muscles and in the atrophied thoracic muscles of crabs that had lost a limb. Mykles says ‘Rheb is a key activator of mTOR. Its up-regulation by moulting indicates that Rheb plays a role in the stimulation of mTOR-mediated protein synthesis by ecdysteroids’.
However, when the team investigated the mechanism by which Rheb is upregulated in the two muscles, there were differences. They found a strong correlation between the levels of Rheb in the crabs’ claws and the ecdysteroid levels, suggesting that ecdysteroid does regulate protein synthesis via mTOR in the claw muscle. Yet, when they compared the ecdysteroid levels and Rheb levels in the thoracic muscles – that wither in response to limb loss – the team found no correlation. Ecdysteroid hormone does not control muscle atrophy in response to limb loss.
Analysing the expression patterns of Rheb and myostatin in atrophied claw and thoracic muscles, Mykles and colleagues also saw different expression relationships between the proteins in the two tissues. ‘If blackback land crab myostatin plays a role in both types of atrophy, any unifying mechanism must reconcile the differences between myostatin and Rheb expression in the two muscles’, the team says.
Having shown that Rheb is a key component of the signalling pathway that regulates muscle atrophy in crabs, the team says that it is keen to build, ‘a mechanistic understanding of the interactions between ecdysteroids and the signalling pathways that control protein metabolism in crustacean skeletal muscle’.