RESUMO
Glutathione transferases (GSTs; EC 2.5.1.18) play important roles in stress tolerance and metabolic detoxification in plants.In higher plants, studies on GSTs have focussed largely on agricultural plants. There is restricted information about molecular characterization of GSTs in gymnosperms. To date, only tau class GST enzymes have been characterized from some pinus species. For the first time, the present study reports cloning and molecular characterization of two zeta class GST genes, namely PbGSTZ1 and PbGSTZ2 from Pinus brutia Ten., which is an economically important pine native to the eastern Mediterranean region and have to cope with several environmental stress conditions. The PbGSTZ1 gene was isolated from cDNA, whereas PbGSTZ2 was isolated from genomic DNA. Sequence analysis of PbGSTZ1 and PbGSTZ2 revealed the presence of an open reading frame of 226 amino acids with typical consensus sequences of the zeta class plant GSTs. Protein and secondary structure prediction analysis of two zeta class PbGSTZs have shared common features of other plant zeta class GSTs. Genomic clone, PbGSTZ2 gene, is unexpectedly intronless. Extensive sequence analysis of PbGSTZ2, with cDNA clone, PbGSTZ1, revealed 87% identity at nucleotide and 81% identity at amino acid levels with 41 amino acids differences suggesting that genomic PbGSTZ2 gene might be an allelic or a paralogue version of PbGSTZ1.
Assuntos
Glutationa Transferase/genética , Pinus/enzimologia , Proteínas de Plantas/genética , Sequência de Aminoácidos , Sequência de Bases , Northern Blotting , Densitometria , Glutationa Transferase/química , Glutationa Transferase/metabolismo , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Dados de Sequência Molecular , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Alinhamento de Sequência , Análise de Sequência de DNARESUMO
The effect of Thapsigargicin (TGC), a non-phorbol ester type tumor promoter, on Ca2+ movements has been investigated using L1210 mouse lymphoma cells. Ca2+ release from intact and digitonin permeabilized cells was evaluated using Fura-2 and Fura-3. TGC like Thapsigargin (TG) has the ability to discharge the intracellular Ca2+ stores and to increase intracellular free Ca2+ concentrations. TGC in a concentration dependent manner (0.16-16 nM) also inhibited cell growth and this effect was at least partially reversed by arachidonate.