Characterisation and Expression of Calpain Family Members in Relation to Nutritional Status, Diet Composition and Flesh Texture in Gilthead Sea Bream (Sparus aurata)

Abstract

Calpains are non-lysosomal calcium-activated neutral proteases involved in a wide range of cellular processes including muscle proteolysis linked to post-mortem flesh softening. The aims of this study were (a) to characterise several members of the calpain system in gilthead sea bream and (b) to examine their expression in relation to nutritional status and muscle tenderisation. We identified the complete open reading frame of gilthead sea bream calpains1-3, sacapn1, sacapn2, sacapn3, and two paralogs of the calpain small subunit1, sacapns1a and sacapns1b. Proteins showed 63–90% sequence identity compared with sequences from mammals and other teleost fishes, and the characteristic domain structure of vertebrate calpains. Transcripts of sacapn1, sacapn2, sacapns1a and sacapns1b had a wide tissue distribution, whereas sacapn3 was almost exclusively detected in skeletal muscle. Next, we assessed transcript expression in skeletal muscle following alteration of nutritional status by (a) fasting and re-feeding or (b) feeding four experimental diets with different carbohydrate-to-protein ratios. Fasting significantly reduced plasma glucose and increased free fatty acids and triglycerides, together with a significant increase in sacapns1b expression. Following 7 days of re-feeding, plasma parameters returned to fed values and sacapn1, sacapn2, sacapns1a and sacapns1b expression was significantly reduced. Furthermore, an increase in dietary carbohydrate content (11 to 39%) diminished growth but increased muscle texture, which showed a significant correlation with decreased sacapn1 and sacapns1a expression, whilst the other calpains remained unaffected. This study has demonstrated that calpain expression is modulated by nutritional status and diet composition in gilthead sea bream, and that the expression of several calpain members is correlated with muscle texture, indicating their potential use as molecular markers for flesh quality in aquaculture production.

Regulatory mechanisms involved in muscle and bone remodeling during refeeding in gilthead sea bream

“…Several members of the main proteolytic systems have been characterized and their responses upon different experimental challenges investigated in different fish species 18,[55][56][57][58][59][60] , including gilthead sea bream [20][21][22] ; however, due to the different experimental conditions (i.e. duration of periods, fish age, etc.)…”

“…This is more important in fish that have a specific and enhanced protein metabolism. The main endogenous proteolytic systems, each one performing specific degradative or regulatory functions according to the nutritional conditions are, calpains, cathepsins and ubiquitin-proteasome, all of which are well-known in fish, especially salmonids [17][18][19] , but also in gilthead sea bream [20][21][22] .In addition to muscle, bone is also an important tissue for skeletal and locomotion functions, but also, as a reservoir of minerals that is clearly affected when nutrition is restricted. Essential during development, synchronicity between bone and muscle is required for proper musculoskeletal growth 23,24 .…”

“…This is more important in fish that have a specific and enhanced protein metabolism. The main endogenous proteolytic systems, each one performing specific degradative or regulatory functions according to the nutritional conditions are, calpains, cathepsins and ubiquitin-proteasome, all of which are well-known in fish, especially salmonids [17][18][19] , but also in gilthead sea bream [20][21][22] .…”

Regulation of lipid metabolism and peroxisome proliferator-activated receptors in rainbow trout adipose tissue by lipolytic and antilipolytic endocrine factors

“…There is limited knowledge on the hormonal control of lipid turnover in fish adipose tissue [7][8][9]. It has been reported that growth hormone (GH), in addition to being a growth promoter, exerts a lipolytic effect in gilthead sea bream adipocytes [7,10]. GH transgenic coho salmon [11] are shown to increase their utilization of lipids for synthetic roles to maintain accelerated growth, but the specific effects in adipose tissue of GH overexpression remain unknown.…”

“…Insulin also stimulates lipoprotein lipase (LPL) activity in rainbow trout adipose tissue [17] and reduces the basal lipolysis level in rainbow trout and gilthead sea bream adipocytes [7,8]. Insulin-like growth factor (IGF)-I is structurally and functionally similar to insulin but is more potent as a growth factor and a metabolic controller in rainbow trout and gilthead sea bream myocytes and adipocytes [10,14,15,18]. In vertebrates, many of the growth-promoting actions of GH are known to be mediated indirectly through the stimulation of IGF transcription, mainly by the liver, or locally by extra hepatic tissues [19], but the mechanisms of action involved in GH proliferative and metabolic effects in fish are not well known [20].…”