Sirtuins belong to a class of NAD-dependent deacetylases, and include seven distinct isoforms, of which SIRT7 is the least studied member. In the present study, the subcellular expression of SIRT7 in primary fibroblasts undergoing senescence was evaluated by immunocytochemistry and immunoblot assay. Expression of nucleolar SIRT7 in young fibroblast was very prominent, decreased in pre-senescent cells, and became undetectable in the senescent cells. Interestingly, we observed previously unreported staining for cytoplasmic SIRT7 in fibroblasts, and report the existence of a steady-state level of SIRT7 in cytoplasm. Selective localization of the high-molecular-mass (47.5 kDa) SIRT7 in the cytoplasmic fraction and the low-molecular-mass (45 kDa) SIRT7 in the nuclear fraction was observed in the immunoblot analysis of various cell types. The specificity of the N-terminal antibodies for detection of cytoplasmic SIRT7 was confirmed by RNAi and peptide competition assays. The two forms of SIRT7 showed reciprocal expression following serum starvation, nocodazole and okadaic acid treatments, and also during senescence. Using a combination of deletion constructs and site-directed mutagenesis, we defined the role of two distinct SIRT7 sequences in the N-terminal region (amino acids 61-76, LQGRSRRREGLKRRQE) and the C-terminal region (amino acids 392-400, KRTKRKKVT) for nuclear and nucleolar import, respectively. In conclusion, we report for the first time the existence of a cytoplasmic pool of SIRT7 in addition to its well-known nucleolar form, identify distinct localization signals for its nuclear/nucleolar targeting, and describe an association between loss of nucleolar SIRT7 and replicative senescence.
Glycogen content and metabolic enzyme activities viz. lactate dehydrogenase (LDH), malate dehydrogenase (MDH), aspartate amino transferase (AST) and alanine amino transferase (ALT) in Indian major carps, Labeo rohita, Catla catla and Cirrhinus mrigala, were investigated after a 6 h transportation trial to compare the species‐specific variation and the effect of increased packing density on the metabolism. Fish (45±5 mm, 0.5±0.1 g) were packed in three densities (100, 150 and 200 L−1) for the experiment, and 12 specimens of each species were randomly sampled from all the treatments at the end of transportation. The glycogen content of L. rohita ingerlings decreased significantly (P<0.05) with increasing packing density. The activities of enzymes LDH, MDH, AST and ALT showed a rising trend with increasing packing density in all the three species. Species‐specific differences were observed in various tested parameters at the lowest packing density (100 fry L−1). Alanine amino transferase and LDH activities were significantly (P<0.05) lower in C. mrigala as compared with the other two species. However, glycogen reserves and MDH activity were not significantly different (P>0.05) among the species. The present study reveals that the optimum packing density for Indian major carp fry (100 fry L−1) for transportation up to 6 h and metabolic regimes are species specific during transportation.
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