Incorporation of amino acids by rat liver nucleoli in vitro.

Isolated rat liver nucleoli are shown to incorporate radioactive amino acids into proteins. The characteristics of this nucleolar protein-synthesizine system indicated that the incorporation observed was not due to cytoplasmic or bacterial contamination. The products synthesized by isolated nucleoli have been analyzed by polyacrylamide gel electrophoresis and subsequent autoradiography; this revealed a few radioactive bands suggesting that in vitro only a few proteins are labelled. The results presented are discussed in view of a possible nucleolar protein synthesis.

Protein synthesis inhibition in rat liver by the mycotoxin patulin.

Patulin, a carcinogenic mycotoxin, inhibits in vivo and in vitro protein synthesis in rat liver. The in vivo inhibition culminates 5 h after toxin administration and reaches a maximum of 65% with regard to control values; a breakdown of polysomes is associated with the translational blockage. However in in vitro systems, the cellular fractions obtained from patulin-treated rats appear equally as active in protein synthesizing ability and as sensitive to the toxin action as those prepared from control animals. The in vitro inhibition by patulin is dose related. The postmitochondrial system is less sensitive than that functioning with isolated polysomes and pH 5 enzyme. This difference might be due to the presence of soluble factor(s) that counteract patulin action. We propose that the inhibition of protein synthesis by patulin may result from an interaction of the drug with active SH groups at the membrane level (amino acid transport, ion equilibrium) and/or at the cytoplasmic level (enzymes and factors involved in the translational process).

Mechanism of the inhibition of transcription by PR toxin, a mycotoxin from Penicillium roqueforti.

PR toxin, a mycotoxin synthesized by Penicillium roqueforti, impairs the transcriptional process in liver cells; the two main RNA polymerase systems (enzymes A and B) are affected by PR toxin. The toxin does not require an enzymic conversion before interfering with in vitro RNA synthesis. Addition of ammonium sulphate completely prevents the inhibition of transcription by PR toxin. In vitro results, using RNA polymerase purified from E. coli, suggest that PR toxin impairs the activity of the RNA polymerase itself. Regarding the step of the transcription process affected, it is shown that PR toxin inhibits both initiation and elongation of the polynucleotide chain.

Relationships between the chemical structure and the biological properties of some eremophilane compounds related to PR toxin.

The problem of the chemical structure--biological effects relationships has been studied for various eremophilane compounds related to PR toxin (PRT), a mycotoxin synthesized by Penicillium roqueforti. The biological tests were based on in vivo toxicity for male Swiss mice and inhibition of in vitro transcription and tranlation. The results showed a good correlation between the responses obtained for the three tests by the different compounds; thus, the toxic potency and the capacity of inhibiting transcription and translation should be directed by a common chemical structure. The data also indicated that the biological properties are related to the existence of an aldehyde group in position 12. In addition, evidence has been obtained demonstrating the hydrolysis of PRT imine with formation of a PRT-like compound.

Effects of the mycotoxin botryodiplodin on mammalian cells in culture.

The effects of botryodiplodia, a mycotoxin synthesized by Botryodiplodia theobromae and by some strains of Penicillium roqueforti. The toxin inhibited cell multiplication in growing cultures at concentrations which were without toxic effect on cultures nearing or at confluence. In growing cultures the toxin affected DNA, RNA and protein synthesis, the effect on DNA synthesis being the greatest. The removal of mycotoxin from the culture medium brought about partial recovery of the synthetic activities of the cells. The implications of the results with respect of the recovery of DNA synthesis are discussed.

Action of monovalent cations on the biological properties of PR toxin, a mycotoxin from Penicillium roqueforti.

PR toxin impairs liver cell metabolism by inhibiting RNA and protein synthesis. In vitro, the drug inhibits the transcription carried out by isolated rat liver nuclei and the translation promoted by polysomes. The action of monovalent cations on the biological activities of PR toxin has been studied. The increased ionic strength due to the presence of salt in the incubation medium, lowers the inhibitory action of PR toxin on in vitro transcription and translation activities; this action is reversible. Besides the overall effect of the ionic strength, ammonium salts possess a specific ability to suppress irreversibly the biological properties of PR toxin (in vivo toxicity and capacity of inhibiting RNA and protein synthesis). The mechanism of this action is discussed.

Induction of cross-links between DNA and protein by PR toxin, a mycotoxin from Penicillium roqueforti.

PR toxin, a mycotoxin from Penicillium roqueforti, induces DNA--protein cross-links in chromatin of both cultured cells and isolated rat-liver nuclei. The presence of the aldehyde group in the PRT molecule is required for the induction of cross-linking; methylene bridges between nucleic acid and protein are presumably involved in the complex formation. The role of other functional groups of PR toxin is discussed.

Induction and mutagenesis of prophage lambda in Escherichia coli K12 by metabolites of aflatoxin B1.

Like most carcinogens, aflatoxin B1 must be activated by mammalian microsomal enzymes to give rise to coupounds active on bacteria. These compounds act as inducers of E. coli K12 (lambda) at a high efficiency, whereas unmodified aflatoxin B1 has no effect. Moreover, metabolites of aflatoxin B1 have a mutagenic action on phage lambda, as shown by the appearance of clear plaque mutants. We propose the hypothesis that the same derivative is responsible for carcinogenesis of liver cells by aflatoxin B1. Therefore, our system provides a simple way of measuring in vitro, in the same assay, the mutagenic and inducing activities of compounds to which the cells are permeable, thereby detecting potentially carcinogenic agents.

Induced reactivity of UV-damaged phage gamma in E. coli K12 host cells treated with aflatoxin B1 metabolites.

The metabolites of aflatoxin B1, the most potent hepatocarcinogen so far known, promote in E. coli K12 cells the reactivation of phage lambda damaged by ultraviolet (UV) radiation. This reactivation process is error prone; 25% of the phage DNA lesions are repaired, but mutagenesis, scored as clear plaque formation, is increased as much as 10-fold. Such reactivation of UV-damaged phage lambda, which occurs in wild-type and in uvrA but not in recA bacteria, is inducible: phage reactivation is obtained even after a long delay following treatment of the host by the short-lived metabolites. This induced reactivation of UV-damaged phage in hosts treated with metabolites of aflatoxin B1 is similar to direct of indirect UV reactivation. Metabolites of aflatoxin B1 produce induced phage reactivation as well as prophage lambda induction in lysogens and cell filamentation in non-lysogens. These cellular events are also triggered by DNA lesions caused by UV radiation and result from the induction of a metabolic pathway (SOS functions). We postulate that, in eucaryotes, carcinogens may induce cellular SOS functions similar to those in E. coli. Induction of such functions might be responsible for the transformation of mammalian cells.