Nucleic acids and growth of larval and early juvenile spider crab, Hyas araneus

The spider crab Hyas araneus L. was reared in the laboratory from hatching of zoea I (ZI) through the first juvenile instar (CI). Within a given moult cycle, individuals of the same age were sampled in intervals of two (ZI, ZII, CI) or 3d (megalopa) for analysis of dry weight, carbon (C), nitrogen (N), hydrogen (H), protein, DNA and RNA. Lipid was calculated from C. Biomass, growth rate and nucleic acid contents showed high variability during each moult cycle and between instars. Instantaneous growth rates of carbon and nitrogen were high in postmoult and intermoult, and low in the premoult periods of each moulting cycle. A shift was observed from high rates of lipid accumulation in the postmoult and intermoult stages to proportionally increasing protein accumulation during late premoult (ZI), or throughout a major part of the remaining moult cycle (in all other instars). DNA was accumulated throughout the ZI and ZII instars, cut it decreased in the late premoult megalopa. It increased again from late intermoult through intermediate premoult in the juveniles. RNA increased continuously during the ZI and ZII, and it decreased in the megalopa, almost to level that had been found immediately after hatching. In juveniles, variation in RNA followed it followed closely those in DNA. Cell multiplication (expressed by DNA increase) dominated over increase in cell size (defined by the C:DNA ratio) during the zoeal instars and in postmoult through early intermoult in the megalopa and CI. When specific (C-related) RNA values and DNA/RNA ratios were compared with instantaneous growth rates in C and N, no general correspondence was detected. The only significant relationship between specific RNA values and instantaneous C or N growth rates was found in the megalopa. The same held for the relationship between the RNA/DNA ratio and growth. Here, in addition to the megalopa, a correspondence with C growth was also found in the CI instar. Our results suggest that variation in nucleic acids may provide useful insights into mechanisms of growth on the cellular level (cell multiplication vs. cell enlargement), however, lack of general correlation with variation in growth rates of Hyas araneus larvae shows that the use of nucleic acids as a measure of growth is probably based upon too simplistic assumptions and hence, does not yield reliable predictions

Hepatocyte nuclear factor 4alpha transactivates the mitochondrial alanine aminotransferase gene in the kidney of Sparus aurata

Alanine aminotransferase (ALT) plays an important role in amino acid metabolism and gluconeogenesis. The preference of carnivorous fish for protein amino acids instead of carbohydrates as a source of energy lead us to study the transcriptional regulation of the mitochondrial ALT (mALT) gene and to characterize the enzyme kinetics and modulation of mALT expression in the kidney of gilthead sea bream (Sparus aurata) under different nutritional and hormonal conditions. 5′-Deletion analysis of mALT promoter in transiently transfected HEK293 cells, site-directed mutagenesis and electrophoretic mobility shift assays allowed us to identify HNF4α as a new factor involved in the transcriptional regulation of mALT expression. Quantitative RT-PCR assays showed that starvation and the administration of streptozotocin (STZ) decreased HNF4α levels in the kidney of S. aurata, leading to the downregulation of mALT transcription. Analysis of the tissue distribution showed that kidney, liver, and intestine were the tissues with higher mALT and HNF4α expression. Kinetic analysis indicates that mALT enzyme is more efficient in catalyzing the conversion of L-alanine to pyruvate than the reverse reaction. From these results, we conclude that HNF4α transactivates the mALT promoter and that the low levels of mALT expression found in the kidney of starved and STZ-treated fish result from a decreased expression of HNF4α. Our findings suggest that the mALT isoenzyme plays a major role in oxidazing dietary amino acids, and points to ALT as a target for a biotechnological action to spare protein and optimize the use of dietary nutrients for fish culture