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1.
2.
A pertussis toxin-sensitive G-protein mediates some aspects of insulin action in BC3H-1 murine myocytes. 总被引:9,自引:0,他引:9
L Luttrell E Kilgour J Larner G Romero 《The Journal of biological chemistry》1990,265(28):16873-16879
The involvement of G-proteins in the insulin signal transduction system has been studied in detail using the murine BC3H-1 myocyte system. Pertussis toxin (PT) treatment, previously shown to attenuate some of the metabolic effects of insulin in this cell line (Luttrell, L.M., Hewlett, E.L., Romero, G., and Rogol, A.D. (1988) J. Biol. Chem. 263, 6134-6141), abolished insulin-induced generation of diacylglycerol and inositolglycan mediators with no effects on either the autophosphorylation of the insulin receptor or the phosphorylation of the major endogenous substrates for insulin-stimulated tyrosine kinase activity (pp185 and pp42-45). In vitro ADP-ribosylation and immunoblotting studies suggest that the major PT substrate is a 40-kDa protein of the G alpha family. This protein band did not exhibit detectable tyrosine phosphorylation upon stimulation of either intact cells or cell membranes with insulin. In the presence of low concentrations of GTP, insulin treatment of isolated myocyte plasma membranes resulted in a small (30-40%) but significant stimulation of GTP hydrolysis. This effect was best observed in the presence of small concentrations of sodium dodecyl sulfate. The rate of guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) binding to BC3H-1 membranes was also significantly increased in the presence of insulin. The effects of insulin on GTP hydrolysis and GTP gamma S binding were found to be dependent on the concentration of insulin. These effects were not detected in plasma membranes prepared from PT-pretreated BC3H-1 myocytes. In contrast, pretreatment with the B (inactive) subunit of PT did not alter the response of myocyte membranes to insulin. High affinity binding of [125I]iodoinsulin to myocyte plasma membranes was reduced by 60-70% in the presence of guanine nucleotides. Similar effects on insulin binding were produced by PT pretreatment of the cells. In contrast, adenine nucleotides had no effect on insulin binding. Scatchard analysis of the binding data showed that the observed effects of guanine nucleotides and PT on insulin binding resulted either from a reduction in the number of high affinity insulin binding sites or from a significant reduction of the affinity of insulin for its receptor. Low affinity binding sites did not appear to be affected by either guanine nucleotides nor PT pretreatment. These results provide substantial evidence suggestive of a noncovalent interaction between the insulin receptor and a regulatory G-protein system during the process of insulin signaling. 相似文献
3.
Intracellular accumulation of nitrate, indicative of the operation of an active nitrate transport system, has been measured in intact cells of the cyanobacterium Anacystis nidulans. The ability of the cells to accumulate nitrate was effectively hindered by either ammonium addition or selective inhibition of CO2 fixation by DL-glyceraldehyde, with the effect of either compound being prevented by previously blocking ammonium assimilation. The results support the contention that nitrate utilization in cyanobacteria is regulated at the level of nitrate transport through the concerted action of ammonium assimilation and CO2 fixation. 相似文献
4.
M M Almar J Díaz-Mayans F J Romero 《Comp. Biochem. Physiol. C, Comp. Pharmacol. Toxicol.》1987,87(2):433-435
1. Glutathione content and GSH S-transferase activity in the midgut gland of Procambarus clarkii (P. c.) of different sex and body weight are presented. 2. Procambarus clarkii females' GSH concentration in the midgut gland decreases to a higher extent upon fasting, compared with males. 3. Procambarus clarkii females, both in control and fasting conditions, have a slightly higher GSH S-transferase activity than males. 4. Cadmium present in water only affects GSH content and GSH S-transferase activity (after 96 hr) in midgut gland, with cadmium chloride concentrations higher than 100 micrograms/l. 相似文献
5.
6.
The purpose of these studies was to determine if Interleukin-1 (IL-1) alters the rate of prostaglandin biosynthesis by human amnion. Primary monolayer cultures of amnion cells were established from women undergoing elective cesarean section before the onset of labor. Natural purified and recombinant human IL-1 alpha and IL-1 beta were incubated with amnion cells in culture, and prostaglandin E2 (PGE2) biosynthesis was measured by radioimmunoassay in cell-free media. A concentration-dependent increase in PGE2 production by amnion cells occurred in response to natural purified and recombinant IL-1 preparations. No differences in the parameters of the dose-response curves between the two IL-1 gene products could be determined (p greater than 0.05). Indomethacin blocked the effect of IL-1 in prostaglandin biosynthesis by human amnion. Interleukin-1, a fever mediator, could serve as a signal for the initiation of labor in cases of intrauterine or systemic infection. 相似文献
7.
A sensitive procedure has been developed for the determination of intracellular nitrate. The method includes: (i) preparation of cell lysates in 2 M-H3PO4 after separation of cells from the outer medium by rapid centrifugation through a layer of silicone oil, and (ii) subsequent nitrate analysis by ion-exchange h.p.l.c. with, as mobile phase, a solution containing 50 mM-H3PO4 and 2% (v/v) tetrahydrofuran, adjusted to pH 1.9 with NaOH. The determination of nitrate is subjected to interference by chloride and sulphate when present in the samples at high concentrations. Nitrite also interferes, but it is easily eliminated by treatment of the samples with sulphamic acid. The method has been successfully applied to the study of nitrate transport in the unicellular cyanobacterium Anacystis nidulans. 相似文献
8.
One of the most widely used mechanisms by which the role of glutathione (GSH) in cellular functions has been withdrawn, is to deplete GSH intracellularly. The importance of the procedure and xenobiotic chosen to get it is discussed. Mitochondrial GSH plays certainly an important role in maintaining cellular homeostasis. This contribution varies depending on the tissue and the conclusions obtained about the functions of this GSH pool in one organ may not be applied to others. Original data on the subcellular distribution of GSH in myocardial tissue of the rat are presented, and the effect of phorone on both cardiac GSH pools is compared with the effect in liver. The mechanical failure of myocardium after ischemic or reperfusion damage might involve mitochondrial GSH, in view of the literature data referring to the role of thiol groups in energy transfer from mitochondria to cytosol. 相似文献
9.
Relationship between a 47-kDa cytoplasmic membrane polypeptide and nitrate transport in Anacystis nidulans 总被引:2,自引:0,他引:2
M N Sivak C Lara J M Romero R Rodríguez M G Guerrero 《Biochemical and biophysical research communications》1989,158(1):257-262
The polypeptide composition of cytoplasmic membranes of the cyanobacterium Anacystis nidulans changes in response to variations in the nitrogen source available to the cells, differing specifically in the amount of a polypeptide of 47-kDa molecular mass. Synthesis of the polypeptide and expression of nitrate transport activity are repressed by ammonium. Transfer of ammonium-grown cells to a medium containing nitrate as the sole nitrogen source results in parallel development of the 47-kDa polypeptide and nitrate transport activity of the cells. These results suggest the involvement of the 47-kDa cytoplasmic membrane polypeptide in nitrate transport by A. nidulans. 相似文献
10.
Cecilia Gotor Eloísa Pajuelo Luís C. Romero Antonio J. Márquez José M. Vega 《Archives of microbiology》1990,153(3):230-234
Polyclonal antiserum specific for ferredoxin-nitrite reductase (EC 1.7.7.1) from the green alga Chlamydomonas reinhardii recognized the nitrite reductase from other green algae, but did not cross-react with the corresponding enzyme from different cyanobacteria or higher plant leaves. An analogous situation was also found for ferredoxin-glutamate synthase (EC 1.4.7.1), using its specific antiserum. Besides, the antibodies raised against C. reinhardii ferredoxin-glutamate synthase were able to inactivate the ferredoxin-dependent activity of nitrite reductase from green algae.These results suggest that there exist similar domains in ferredoxin-nitrite reductases and ferredoxin-glutamate synthases from green algae. In addition, both types of enzymes share common antigenic determinants, probably located at the ferredoxin-binding domain. In spite of their physicochemical resemblances, no apparent antigenic correlation exists between the corresponding enzymes from green algae and those from higher plant leaves or cyanobacteria.Abbreviations Fd
ferredoxin
- GOGAT
glutamate synthase
- MV+
reduced methyl viologen (radical cation)
- NiR
nitrite reductase
- PMSF
phenylmethylsulphonyl fluoride
- SDS
sodium dodecyl sulfate 相似文献