The root-knot nematode resistance gene (Mi) in tomato: construction of a molecular linkage map and identification of dominant cDNA markers in resistant genotypes

A dominant allele at the Mi locus on chromosome 6 of tomato (Lycopersicon esculentum Mill) confers resistance to three species of root-knot nematodes (Meloidogyne). The resistance, which is associated with a localized necrotic response, was originally introduced into tomato from the wild species Lycopersicon peruvianum. As a step towards the molecular cloning of Mi, we have identified closely linked DNA markers from both cDNA and genomic DNA libraries as restriction fragment length polymorphisms (RFLPs). DNA from tomato populations segregating for nematode resistance was analyzed to generate a high-resolution genetic map of this region. Additional information on gene order was obtained by comparing the size of the introgressed L. peruvianum chromosomal segment within a collection of nematode-resistant tomato lines. Among the four cDNA markers that are tightly linked to Mi, three are dominant, i.e. L. peruvianum-specific. One cDNA marker corresponds to a gene family comprising 20-30 members, one of which is diagnostic for all nematode-resistant genotypes tested. The presence of non-homologous sequences around the Mi gene may contribute to the suppression of recombination in this region of the genome in crosses heterozygous for Mi. The potential of 'walking' from closely linked markers to Mi is discussed.     

心房钠泵因子对颈动脉窦压力感受器反射的易化作用

Effects of atriopeptin II (APII) on the carotid sinus baroreflex in anesthetized rats and on the sinus nerve afferent activity in the anesthetized rabbits were investigated. The results were as follows: (1) By perfusing the isolated left carotid sinus with APII (1 microgram/ml) in anesthetized rats (n = 10), the threshold pressure (TP) of the carotid baroreflex did not show any change, while the equilibrium pressure (EP), the saturation pressure (SP) and the operating range (OR) were decreased from 101 +/- 2.8 to 95 +/- 2.0 mmHg (P less than 0.05), 202 +/- 5.2 to 168 +/- 6.1 mmHg (P less than 0.001) and 128 +/- 5.5 to 93 +/- 6.3 mmHg (P less than 0.001), respectively. The function curve of the baroreflex was shifted to the left and downward with a peak slope (PS) increased during perfusing with APII. In contrast, by perfusing the carotid sinus with sodium nitroprusside (NP, 0.5 micrograms/ml), TP and EP remained unchanged, whereas SP and OR were increased from 188 +/- 6.4 to 218 +/- 6.0 mmHg (n = 6, P less than 0.01) and from 107 +/- 6.9 to 132 +/- 7.6 mmHg (P less than 0.05), respectively. The function curve of the baroreflex and its PS were not affected by NP. The sinus nerve afferent activity was quite stable with the perfusion of carotid sinus at constant intrasinus pressure (ISP) in the rabbits (n = 6) and increased during the elevation of ISP.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence for age-related changes in pyridine nucleotide content of isolated rat islets

The purpose of this study was to document the effect of age on alpha-glycerophosphate activity and pyridine nucleotide concentration in pancreatic islets isolated from rats. In order to do this, islets were isolated from pancreases of 2 and 12 month-old rats, and measurements made of alpha-glycerophosphate activity and of NAD+ and NADH, determinations were made following incubation at both basal (5.6 mM) and elevated glucose concentrations (28 mM). The results indicated that islet alpha-glycerophosphate dehydrogenase activity was decreased (P less than 0.001) by approximately 50% in the older rats. This was associated with an increase in mean (+/- SEM) basal NADH content (pmol/microgram DNA) in 12 month-old (4.48 +/- 0.31) as compared to 2 month-old rats (2.73 +/- 0.49). Although mean (+/- SEM) basal NAD+ levels (pmol/microgram DNA) were the same in 2 and 12 month-old rats (29.4 +/- 2.5 and 30.8 +/- 2.8, respectively), NAD+ content following incubation at elevated levels of glucose declined (absolutely and relatively) to a significantly greater degree in the younger rats. The incremental rise in islet NADH concentration following incubation at the elevated glucose concentration was similar in the two groups, but the relative increase was only approximately half as great in islets from 12 month-old rats. These data indicate that the age-related decline in the activity of alpha-glycerophosphate dehydrogenase, the enzyme regulating the glycerophosphate shuttle system in 12 month-old rats, is associated with alterations in islet pyridine nucleotide composition.(ABSTRACT TRUNCATED AT 250 WORDS)     

Insulin induces activation and translocation of protein kinase FA (a multifunctional protein phosphatase activator) in human platelet

Protein kinase FA (an activator of the ATP.Mg-dependent multifunctional protein phosphatase) has been identified in both cytosol and plasma membrane isolated from human platelets. The FA activity in the cytosol is active whereas the FA activity in the membrane is inactive. Quantitative analysis further indicates that approximately 90% of total FA is present in the membrane whereas only 10% of FA is localized in the cytosol, suggesting that the inactive membrane-associated FA might be regulated. This notion has subsequently been demonstrated that exposure of platelets to physiological concentrations of insulin for only 1 min resulted in an increase in cytosolic FA activity to about 300% of control values in the absence of insulin and in a corresponding decrease in FA activity in the membrane. It is concluded that the molecular basis for insulin action on cellular metabolism may partly be mediated through the activation and translocation of protein kinase FA in the membrane. It is suggested that redistribution of protein kinase FA may represent a transmembrane signal of insulin.     

Effects of microorganisms on effective oxygen diffusion coefficients and solubilities in fermentation media

Effective oxygen diffusion coefficients and solubilities were measured for submerged cultures of Saccharomyces cerevisiae, Escherichia coli, and Penicillium chrysogenum. Both effective oxygen diffusion coefficients and solubilities were found to decrease with increasing cell concentrations in the fermentation media. Comparison of the experimental results of effective oxygen diffusion coefficients in fermentation media with values theoretically predicted on the assumption of unpenetrable microbial cells indicates that oxygen molecules diffuse through the cells during the diffusion process. Within the cell concentration range of typical submerged fermentations, the effective oxygen diffusion coefficient of the fermentation media can be described as D(e) = A(1)f + A(2)f(2). In this equation, fis the cell volume fraction and both A(1) and A(2) are functions of the shape of the cells and the ratio of effective oxygen diffusion coefficient in microbial cells to that in the medium.     

Protein kinase C: subcellular redistribution by increased Ca2+ influx. Evidence that Ca2+-dependent subcellular redistribution of protein kinase C is involved in potentiation of beta-adrenergic stimulation of pineal cAMP and cGMP by K+ and A23187

Phenylephrine is known to stimulate translocation of protein kinase C in rat pinealocytes (Sugden, D., Vanecek, J., Klein, D.C., Thomas, T.P., and Anderson, W. B. (1985) Nature 314, 359-361). In the present study, the receptor mediating this effect was found to belong to the alpha 1-adrenoceptor subclass. Activation of this receptor is also known to produce a sustained increase in [Ca2+]i by increasing net influx (Sugden, A. L., Sugden, D., and Klein, D. C. (1985) J. Biol. Chem. 261, 11608-11612), which points to the possible importance of Ca2+ influx in the subcellular redistribution (activation) of protein kinase C in intact cells. This possibility was investigated by reducing extracellular Ca2+ ((Ca2+]o) with EGTA or by inhibiting Ca2+ influx with inorganic Ca2+ blockers. These treatments reduced alpha 1-adrenoceptor-mediated translocation of protein kinase C. This suggested that elevation of Ca2+ influx alone triggers activation of protein kinase C. In support of this, it was found that treatments which elevate Ca2+ influx, including increased extracellular K+ and addition of the Ca2+ ionophore A23187, cause redistribution of protein kinase C. The effect of K+ was blocked by nifedipine and that of A23187 by EGTA, indicating that effects of these agents are Ca2+-dependent. The possible role of phospholipase C activation in these effects was examined by measuring the formation of [3H]diacylglycerol by cells labeled with [3H]arachidonic acid. Although [3H]diacylglycerol formation was easily detected in the presence or absence of an effective concentration of an inhibitor of diacylglycerol kinase, none of the agents which cause rapid translocation of protein kinase C were found to cause a rapid increase in the generation of [3H]diacylglycerol. These findings establish that an increase in Ca2+ influx is sufficient to trigger translocation of protein kinase C. In addition, we found that a very close correlation exists between translocation of protein kinase C by phenylephrine, K+, and A23187 and their ability to potentiate beta-adrenergic stimulation of cAMP and cGMP accumulation. This provides strong support to the proposal that translocation of protein kinase C is required for potentiation of beta-adrenergic stimulation of pinealocyte cAMP and cGMP accumulation.     

Four- and five-coordinate species in nickel-reconstituted hemoglobin and myoglobin: Raman identification of the nickel-histidine stretching mode

Nickel(II)-reconstituted hemoglobin (NiHb) and myoglobin (NiMb) and model Ni porphyrins have been investigated by Soret-resonance Raman difference spectroscopy. Two sets of frequencies for the oxidation-state and core-size marker lines in the region from 1300 to 1700 cm-1 indicate two distinct sites in NiHb. Only one of these sites is evident in the Raman spectra of NiMb. This result is consistent with the UV-visible absorption spectrum of NiHb, which shows two Soret bands at 397 and 420 nm and one Soret at 424 nm for NiMb. Excitation at the blue Soret component of NiHb with 406.7-nm laser radiation preferentially enhances the set of Raman marker lines typical of Ni-protoporphyrin IX [Ni(ProtoP )] in noncoordinating solvents. The wavelength of the blue Soret component and the Raman spectrum indicate four-coordination for this site in NiHb. Laser excitation in the red Soret band enhances a set of lines whose frequencies are compatible with neither four- nor six-coordinate frequencies but are intermediate between the two. The red Soret band of the proteins is also considerably less red shifted than six-coordinate Ni-porphyrin models. These results suggest that Ni in the second site possesses a single axial ligand. Raman spectra of 64Ni-reconstituted and natural abundance Ni-reconstituted hemoglobins, obtained simultaneously in a Raman difference spectrometer, have identified the Ni-ligand stretch at 236 cm-1. The line shifts to 229 cm-1 for the 64Ni-reconstituted Hb. For a pure Ni-ligand stretch a 10-cm-1 shift would be predicted.(ABSTRACT TRUNCATED AT 250 WORDS)