Anthrax toxin: the long and winding road that leads to the kill

The past five years have led to a tremendous increase in our molecular understanding of the mode of action of the anthrax toxin, one of the two main virulence factors produced by Bacillus anthracis. The structures of each of the three components of the toxin--lethal factor (LF), edema factor (EF) and protective antigen (PA)--have been solved not only in their monomeric forms but, depending on the subunit, in a heptameric form, bound to their substrate, co-factor or receptor. The endocytic route followed by the toxin has also been unraveled and the enzymatic mechanisms of EF and LF elucidated.     

About lipids and toxins

Many mono or multicellular organisms secrete soluble proteins, referred to as protein toxins, which alter the behavior of foreign, or target cells, possibly leading to their death. These toxins affect either the cell membrane by forming pores or modifying lipids, or some intracellular target. To reach this target, they must cross one of the cellular membranes, generally that of an intracellular organelle. As described in this minireview, lipids play crucial roles in the intoxication process of most if not all toxins, by allowing/promoting binding, endocytosis, trafficking and/or translocation into the cytoplasm.     

Biochemical genetics of chromosome forms of Venezuelan spiny rats of the proechimys guairae and proechimys trinitatis superspecies

Spiny rats from Venezuela show an extensive karyotypic diversification (2n=24 to 2n=62) and little morphological differentiation. This study reports genetic distance, heterozygosity and polymorphism based upon 22 loci in semispecies and allospecies of the Proechimys guairae superspecies from N Central Venezuela, as compared with Proechimys urichi, a member of the Proechimys trinitatis superspecies from eastern Venezuela. Four chromosome forms of the P. guairae complex are included, each characterized by karyotypes of 2n=46 (Fundamental Number=72), 2n=48 (FN=72), 2n=50 (FN=72) and 2n=62 (FN=74). Proechimys urichi has a distinetive karyotype of 2n=62 (FN=88). The overall mean value of Nei's genetic identity index for all pair-wise comparisons is I=0.942±0.011. Mean identity within the P. guairae complex is =0.969±0.033. Mean identity between P. urichi and members of that complex is =0.889±0.011. Within the P. guairae complex, increased genetic divergence is correlated with higher karyotypic divergence. Heterozygosity varies from H=0.059 to H=0.153, with a mean value of H0.059. The mean percent of polymorphic loci is P=18.2±3.9 after the 0,95% polymorphism criterion, and P=20,5±5.2 after the 0.99% criterion. These results are compared with similar data from fossorial and non-fossorial rodents. Spiny rats are non-fossorial, forest-dwelling rodents which have undergone a speciation process with little genetic divergence and extensive chromosome rearrangements.     

Hormonal and Nutritional Changes in the Flavedo Regulating Rind Color Development in Sweet Orange [Citrus sinensis (L.) Osb.]

The objective of this research was to determine the changes in the levels of endogenous gibberellins GA₁ and GA₄, abscisic acid (ABA), and ethylene during fruit coloring of on-tree fruits of sweet orange. The time course of carbohydrates and nitrogen content in the flavedo prior to fruit color break and during peel ripening were also studied. To identify nutritional and hormonal changes in the fruit, 45?days before fruit color break the peduncles of 15?C30 fruits per tree of ??Washington?? navel, ??Navelate,?? and ??Valencia Delta Seedless?? sweet orange, located in single-fruited shoots, were girdled to intercept phloem transport. A set of 15?C30 fruits per tree remained intact on the peduncle for control. Girdling significantly delayed fruit coloration for more than 2?months; the delay paralleled higher GA₁ and GA₄ concentrations in the flavedo and retarded the rise of ABA concentration prior to color break. Girdling also reduced carbohydrate concentrations and increased N concentrations in the flavedo compared to control fruits; no ethylene production was detected. Therefore, in sweet orange, fruit changes color by reducing active gibberellin concentrations in the flavedo, which are involved in regulating sugars and ABA accumulation and in reducing N fraction concentration as rind color develops. This was demonstrated in vivo without removing the fruit from the tree. Comparable results were obtained with experiments carried out over four consecutive years in two countries (Spain and Uruguay).     

Single coronary artery with transeptal anterior interventricular artery: a rare anatomical feature

A case of a single coronary artery is described in a 50-year-old male, who died of asphyxia. The artery originated in the right aortic sinus and from it another artery emerged which crossed the crista supraventricularis and the interventricular septum and returned to occupy a subepicardial position in the lower half of the anterior interventricular sulcus. This partially intramyocardial artery was considered as the anterior interventricular artery. A literature survey showed only five cases with similar characteristics. The importance of this anomaly derives from the risk of damage occurring to the intramyocardial artery during a manipulation of the infundibulum of the right ventricle in a cardiac surgery or from problems of perfusion during coronary bypass procedures.     

A Two-Dimensional Electrophoresis Study of Phosphorylation and Dephosphorylation of Chromaffin Cell Proteins in Response to a Secretory Stimulus

Phosphorylated proteins of bovine chromaffin cells, radioactively labeled with [32P]orthophosphate, have been analyzed by two-dimensional polyacrylamide gel electrophoresis and autoradiography. Complex two-dimensional electrophoretograms were studied with the aid of computer-assisted image analysis (CAIA). A database map of 32P-labeled proteins was constructed; approximately 500 polypeptides have been detected, numbered, and characterized according to the intensity of labeling, molecular weight, and isoelectric point. The database was constructed from cells kept in resting conditions or stimulated with 59 mM K+ in 2.5 mM Ca2+ or in 0 Ca2+ solution. These manipulations caused statistically significant changes in the degree of phosphorylation of 20 proteins; they were classified as Ca2+-dependent substrates for the phosphorylation or dephosphorylation processes. These changes were also shown in cells stimulated in the presence of the Ca2+ channel activator Bay K 8644. New proteins that show as much as a fivefold increase in their phosphorylation state during cell stimulation have been located with this methodology, as well as many others that had not previously been detected with conventional methods. These experiments provide the first CAIA database of chromaffin cell phosphoproteins; the map constructed with these data will allow the location of specific phosphoproteins and serve as a reference for future ongoing studies. The database will continue to grow to identify more proteins and to facilitate the comparison of complex patterns obtained in different laboratories for normal and transformed pheochromocytoma PC12 cells.     

Reconstitution of a hormone-sensitive adenylate cyclase with membrane extracts from Neurospora and avian erythrocytes

Adenylate cyclase activity associated to wild type Neurospora membranes is highly dependent on Mn2+ and insensitive to fluoride, guanyl nucleotides, and cholera toxin. These membranes are able to interact with components of detergent extracts from turkey erythrocyte ghosts. The reconstituted cyclase system is catalytically active in the presence of Mg2+ and it is activated by guanyl-5'-yl imidodiphosphate plus isoproterenol and fluoride. When detergent extracts were prepared from avian erythrocyte membranes treated with cholera toxin, the reconstituted system was stimulated by guanyl-5'-yl imidodiphosphate in the absence of isoproterenol and cyclase activities were higher than those observed with extracts from membranes not treated with the toxin. Dose-response curves for isoproterenol and fluoride in the reconstituted system were similar to those reported for avian erythrocyte and liver membranes, respectively.