Shemin pathway and peroxidase deficiency in a fully habituated and fully heterotrophic non-organogenic sugarbeet callus: an adaptative strategy or the consequence of modified hormonal balances and sensitivities in these cancerous cells? A review and reassessment.

There are many arguments for considering a specific fully habituated (auxin and cytokinin-independent) and fully heterotrophic non-organogenic (HNO) sugarbeet callus cell line as terminating a neoplastic progression, and thus to be made of cancerous cells. The similarities with animal tumour and cancer cells are recalled. All types of habituated tissues examined in the literature share at least three common biochemical characteristics: low apparent peroxidase activity, high content of polyamines (PAs) and low production of ethylene. However, results concerning their auxin and cytokinin levels are not consistent. Peroxidase synthesis in the achlorophyllous HNO callus appears to arise from aminolevulinic acid (ALA) synthesis through the Shemin pathway, commonly used by animals and fungi. This pathway is limited by disturbed nitrogen metabolism that diverts glutamate (directly used for ALA synthesis in green higher plants) from the Kreb's cycle into PA synthesis. There is no argument to suggest that the low ethylene production is caused by a competition with PAs for their common precursor, S-adenosylmethionine. The results we report here indicate modified anabolic and catabolic pathways of auxins and cytokinins but also the possibilities of unusual compounds playing similar roles (dehydrodiconiferyl alcohol glucosides, for instance). A higher turnover of PAs is shown in the HNO callus, which could suggest a role for H2O2 and gamma-aminobutyric acid, products or intermediates in the PA catabolic pathway, as secondary messengers. The habituated cells retain some sensitivity towards exogenous auxins and cytokinins. Their increased sensitivity to PAs and ethylene suggests modified hormonal balances for the control of these actively dividing cells.

Molecular cloning of the 5-aminolevulinic acid dehydratase gene from Rhodobacter sphaeroides.

A hemB mutant of Escherichia coli was used to clone the gene encoding 5-aminolevulinic acid dehydratase from Rhodobacter sphaeroides after physiological complementation of the mutation. A 2.9-kb DNA fragment was obtained and cloned in both orientations into the unique PstI restriction site of pUC19. This recombinant plasmid encodes a protein (Mr 39,000) that is immunoreactive with antibodies raised against the enzyme from higher plants.

Inhibitory effect of gabaculine on 5-aminolevulinate dehydratase activity in radish seedlings.

We have compared the activity of 5-aminolevulinate dehydratase (5-ALAD) with the amount of protein detected by specific antibodies in rocket immunoelectrophoresis. Parallel kinetic evolutions of enzymic activity and amount of antigen were observed in radish (Raphanus sativus L.) cotyledons, both in complete darkness or under standard far red light involving phytochrome. However, the treatment of seedlings with gabaculine leads to an important decrease in enzymic activity, while the specific protein content is maintained. This inhibition is not overcome by the addition of glutamic acid, but by 5-aminolevulinic acid which points to a specific control of 5-ALAD activity by its substrate. As there is no discrepancy between the enzymic activity and the amount of antigen during the time course development of seedlings, this could confirm a coordinate cellular control between 5-aminolevulinic acid formation and 5-ALAD protein synthesis, both being amplified by the action of phytochrome.

Production of monospecific antisera by immunization with precipitin line leads to misinterpretation of immunoblot.

Immunoblotting displays an artifact in the preparation of monospecific antibodies from immunocomplexes cut out of line immunoelectrophoresis. Agarose induces the formation of antibodies towards a 68 KDa protein which is contained in commercial agarose.

Lag-phases in phytochrome-mediated enzyme synthesis (PAL).

In experiments with the mustard seedling (Sinapis alba L.) it was confirmed that in the case of "secondary irradiation" induction of PAL by phytochrome is a very rapid process. The lag-phase after the onset of light is too brief to be detected. However, the data of other investigators, who found extended "secondary" lag-phases in their experimental material, can be imitated with the mustard seedling. We explain why these investigators were not able to eliminate the "secondary" lag-phase.