Liver glutathione S-transferase polymorphism in Japanese and its pharmacogenetic importance

Summary A total of 168 autopsy liver extracts from Japanese individuals were examined for the glutathione S-transferase (GST) isozymes by means of starch gel electrophoresis. The gene frequencies of GST1^*1, GST1^*2, and GST1^*0 in Japanese were 0.252, 0.057, and 0.691, respectively. GST1^*3 was detected as a rare variant allele. The incidence of GST1 0 in 41 liver biopsy samples from patients suffering from various liver diseases was investigated using polyacrylamide gel isoelectric focusing. The GST1 0 phenotype was found more frequently in livers with hepatitis and carcinoma than in control livers. The isozymes coded by different GST loci were partially purified and characterized to study their biochemical properties. The apparent Km values with 1-chloro-2,4-dinitrobenzene (CDNB) as substrate for the isozymes at the GST1, GST2, GST3, and GST4 loci were 604, 1345, 776 and 591 M, respectively.     

A Comparison of the Effect of Salt on Polypeptides and Translatable mRNAs in Roots of a Salt-Tolerant and a Salt-Sensitive Cultivar of Barley

The effect of salt stress on polypeptide and mRNA levels in roots of two barley (Hordeum vulgare L.) cultivars differing in salt tolerance (cv CM 72, tolerant; cv Prato, sensitive) was analyzed using two-dimensional polyacrylamide gel electrophoresis. Preliminary experiments indicated that germination of Prato was inhibited significantly in the presence of NaCl, but growth of the surviving Prato seedlings was not substantially different from that of CM 72. Fluorographs of two-dimensional gels containing in vivo labeled polypeptides or in vitro translation products were computer analyzed to identify and quantitate changes that resulted when plants were grown in the presence of 200 millimolar NaCl for 6 days. The patterns of in vivo labeled polypeptides and in vitro products of CM 72 and Prato were qualitatively the same. Salt caused quantitative changes in numerous polypeptides and translatable mRNAs, but, overall, the changes were relatively small. Salt did not induce the synthesis of unique polypeptides or translatable mRNAs and did not cause any to disappear. Because of the similarities of the two cultivars with respect to growth and polypeptide patterns and the slight changes in polypeptide and translation product levels caused by salt, specific polypeptides or translatable mRNAs that are related to salt tolerance in barley could not be identified.     

Estrogen can regulate the cell cycle in the early G1 phase of yeast by increasing the amount of adenylate cyclase mRNA

The effects of beta-estradiol (estrogen; a minor component of yeast cells) on S. cerevisiae cells in the G0 and G1 phases were examined. Results showed that estrogen stimulated the recovery of growth from G0 arrest induced by nutrient limitation or ts mutation of cdc35 (adenylate cyclase) in the early G1 phase, and inhibited entry into the resting G0 phase by increasing the intracellular cAMP level. However, estrogen had no effect on late G1 arrest induced by the alpha factor or ts mutation of cdc36. Estrogen was found to lead to higher steady-state levels of adenylate cyclase mRNA but not to affect the expression of the RAS1 and RAS2 genes, although these can also alter the intracellular cAMP level. These results suggest that estrogen influences the cell cycle of yeast in the early G1 phase by controlling the level of cAMP through the increase of adenylate cyclase mRNA.     

Effect of histamine H1-receptor antagonist on the regulation of carbohydrate and lipid metabolism in rat liver

In order to elucidate the role of histamine in the liver, we studied the effect of a histamine H1-receptor antagonist on the carbohydrate and lipid metabolism in the rat liver. The administration of the H1-receptor antagonist decreased significantly the contents of glycogen and malonyl-CoA in the liver. However, it did not affect the levels of serum glucose and free fatty acid. These results suggest that histamine may play a part in the regulation of metabolism of carbohydrates and lipids in the liver.     

Quantitative determination of sulfated glycopeptide by two-dimensional electrophoresis on cellulose acetate membrane

Quantitative determination of the sulfated glycoproteins present in tissue and secretion fluid was performed. After digestion of the specimen with pronase in order to convert glycoproteins to glycopeptides, the sulfated glycopeptides were separated from a mixture of acidic glycans (glycosaminoglycans, sialoglycopeptides and sulfated glycopeptides) by two-dimensional electrophoresis on cellulose acetate membrane [(1986) J. Biochem. Biophys. Methods 12, 239-246]. After staining with alcian blue, the spot of sulfated glycopeptide on the cellulose acetate membrane was cut out, and then only the dye bound to the sulfated glycopeptide was extracted with a 5% cetylpyridinium chloride solution at 100 degrees C for 15 min. The extract was then measured by absorbance at 615 nm using an authentic sulfated glycopeptide as a standard. This method facilitated the determination of sulfated glycopeptides, which were separated from other acidic glycans, within the range 0-25 micrograms.     

cDNA cloning of an mRNA encoding a sulfur-rich 10 kDa prolamin polypeptide in rice seeds

Using a rice maturing seed pUC9 expression library, we isolated a cDNA clone corresponding to 10 kDa sulfurrich prolamin by immunoscreening. A longer cDNA clone was obtained from a gtll library by plaque hybridization using this ³²P-labeled cDNA as a probe. A polypeptide sequence composed of 134 amino acids was deduced from the nucleotide sequence. A 24 amino acid signal peptide was assigned by computer calculation for the membrane spanning region and Edman sequencing of the purified mature polypeptide. Remarkably, 20% of methionine and 10% of cysteine were found in the mature polypeptide as well as high contents of glutamine, and hydrophobic amino acids. Part of the amino acid sequence was homologous with a conserved cysteine-rich region found in other plant prolamins. Two repeats of amino acid sequence were found in the polypeptide.     

Decrease in ovarian platelet-activating factor during ovulation in the gonadotropin-primed immature rat

Platelet-activating factor (PAF) is a biologically active phospholipid that is released locally during acute inflammatory reactions and tissue injury. Since there is evidence that the biochemical events of mammalian ovulation resemble an inflammatory reaction, the objective of this study was to determine whether ovarian levels of PAF change during ovulation. At 2-h intervals during the ovulatory process in gonadotropin-primed 25-day-old Wistar rats, the ovaries were extirpated, homogenized, and extracted for lipids. The extracts were subjected to thin-layer chromatography (TLC), and the portion of the silica gel that comigrated with PAF was re-extracted and assayed for PAF activity. The PAF was measured (in fmole equivalents of synthetic PAF) by a bioassay based on the capacity of aliquots of the extracts to release [3H]-serotonin from platelets isolated from whole blood of rabbits and prelabeled with [3H]-serotonin. The ovarian level of PAF decreased (p less than 0.01) by 36% from 6.67 +/- 0.77 to 4.27 +/- 0.45 fmoles/mg ovary by 2 h after treatment with human chorionic gonadotropin (hCG), and it declined another 14% by 4 h after hCG. The ovarian PAF remained at this reduced level for up to 24 h after hCG. The administration of indomethacin (5 mg/rat, s.c.) or epostane (5 mg/rat, s.c.) at 1 h after hCG prevented ovulation, but neither drug affected the decline in ovarian PAF. Preliminary tests showed that the lipid extracts from the ovaries also contained PAF inhibitor(s) that comigrated with PAF on the TLC plates. Similar to PAF, the lipid-soluble inhibitor(s) decreased (p less than 0.05) in the ovaries within 4 h after hCG treatment.(ABSTRACT TRUNCATED AT 250 WORDS)