Comparison of the effects of different isolates of Verticillium on phenylalanine ammonia lyase activity and phytoalexin biosynthesis in tissues of Medicago sativa L

Abstract

It is concluded that the receptor occupancy-response relationship is very similar for insulin and antibodies, in terms of the efficiency of coupling of occupied receptor to the signalling pathway which determines metabolic responses. However, this signalling pathway does not require autophosphorylation of receptors in the case of activation by anti-receptor antibodies. Indeed antibody 25-49 had no detectable effect on receptor phosphorylation in situ, even at a concentration 100 x higher than that required for maximum metabolic stimulation. Insulin under the same conditions dramatically increased receptor phosphorylation so that a response amounting to 1/20 of the maximum insulin effect could have been easily detected.The autophosphorylation of solubilized, purified receptor from both NIH 3T3 H1R3.5 and CHO-T cells was stimulated by those antibodies tested (83-7 and 18-44) in a manner very similar to that previously documented with placental receptor [9]. Presumably, the receptor in intact cells is subject to steric constraints or interactions with other proteins, which are not present in solubilized preparations, and which prevent autophosphorylation of the antibody-activated receptor.A mutant insulin receptor lacking kinase activity did not mediate the insulin-like effect of antibodies (25-49, 83-14 and 18-44) on glucose uptake when transfected into CHO cells [13]. It is indeed surprising that metabolic effects of antibodies apparently require a receptor with kinase activity, but do not involve autophosphorylation of the receptor which generally follows activation of this kinase. The explanation for this paradox is not at present clear. It is possible that effects of antibodies involve the increased internalization of a basally active receptor kinase, or the stimulation of kinase activity towards exogenous substrates without accompanying autophosphorylation. The study of further receptor mutants should help to define more precisely the structural features of receptors required to mediate metabolic responses to insulin and antibodies.We are very grateful to Drs R. Roth, L. Ellis, W. J. Rutter and J. Whittaker for the gift of transfected cell lines. We also thank the Medical Research Council, the Wellcome Trust and Serono Diagnostics Ltd for financial support. I , Ilcnton, R. M. ( I 986) Adv.

Binding saturation analysis of inositol-1,4,5-trisphosphate in suspension cultures of lucerne cells

“…A t t h e same t i m e t h e r e is increasing evidence t h a t inositol-1,4,5-trisphosphate ( I n s ( l 4 5)P3) may be responsible f o r t h e release of Ca24 from plant microsomes and vacuoles [l]. Previous work employing [3H]-inositol, has provided provisional identification of 1ns(1,4,5)P in lucerne cell suspension cultures and t h e latelling data obtained is consistent with a role for Ins (1,4,5)P3 in phytoalexin accumulation induced b y fungal elicitor (23. In the studies reported here, binding saturation analysis has been employed t o confirm these earlier findings.…”

“…A f t e r 6 days growth t h e lucerne cell-suspension cultures [3] were either used directly o r were challenged by addition of VerticiUium albo-atrum elicitor solution [4] o r an equivalent volume of H20. LiCl (lOpmol/ml cells) w a s added to t h e suspension cultures a t t h e same t i m e as treatment was initiated.…”

Adenosine 3',5'-Cyclic Monophosphate Extracted from Local Isolate of Bacillus Species

“…After mixing in vortex, 5 µL was injected into HPLC and by the detector recording the chromatogram chart and measuring the retention time. Same technique was used for determination the retention time of the purified cAMP eluted peak (step 3) 25,26 . TLC analysis: A uniform thin layer silica gel G60 has been used as the stationary phase in the project to separated cAMP.…”