Whole genome sequencing of a natural recombinant Toxoplasma gondii strain reveals chromosome sorting and local allelic variants

Background  

Toxoplasma gondii is a zoonotic parasite of global importance. In common with many protozoan parasites it has the capacity for sexual recombination, but current evidence suggests this is rarely employed. The global population structure is dominated by a small number of clonal genotypes, which exhibit biallelic variation and limited intralineage divergence. Little is known of the genotypes present in Africa despite the importance of AIDS-associated toxoplasmosis.     

Regulation of glucose transport in Ca2+-tolerant myocytes from adult rat heart

Calcium-tolerant cardiac myocytes were isolated from adult rat ventricles and sarcolemmal glucose transport was assessed by measuring linear initial uptake rates of the nonmetabolized glucose analog 3-O-methyl-D-glucose in the presence and absence of Ca2+ in the incubation medium. (1) Agents which are known to increase internal Na+ and thus stimulate Ca2+ influx via Na+-Ca2+ exchange stimulated 3-methylglucose transport in the presence of external Ca2+. These include low-Na+ medium, 10(-6) M ouabain and K+-free medium, cyanide and the sodium ionophore, monensin. Hyperosmolarity stimulated transport also in the absence of Ca2+, consistent with release of Ca2+ from internal stores. Transport was decreased in a hypo-osmolar medium and with 10(-9) M ouabain, a concentration which stimulates the Na+ pump. (2) The calcium ionophore A23187 increased basal 3-methylglucose transport but opposed stimulation of transport by insulin. (3) Insulin-stimulated transport was antagonized by palmitate and this effect was reversed by 2-bromostearate, an inhibitor of fatty acid oxidation. These results are identical in all respects to those obtained in intact cardiac and skeletal muscle preparations, confirming that hexose transport in muscle shows Ca2+ dependence and indicating that isolated cardiac myocytes are suitable for the study of this phenomenon.     

Bi-transgenic Mice Reveal that K-rasVal12 Augments a p53-independent Apoptosis When Small Intestinal Villus Enterocytes Reenter the Cell Cycle

Studies in cell culture systems have indicated that oncogenic forms of Ras can affect apoptosis. Activating mutations of Ras occur in ~30% of all human tumors and 50% of colorectal carcinomas. Since these mutations appear at early or intermediate stages in multistep journeys to neoplasia, an effect on apoptosis may help determine whether initiated cells progress towards a more neoplastic state. We have tested the effects of K-ras^Val12 on apoptosis in transgenic mice. A lineage-specific promoter was used to direct expression of human K-ras^Val12, with or without wild-type (wt) or mutant SV-40 T antigens (TAg), in postmitotic villus enterocytes, the principal cell type of the small intestinal epithelium. Enterocytes can be induced to reenter the cell cycle by TAg^Wt. Reentry is dependent upon the ability of TAg to bind pRB and is associated with a p53-independent apoptosis. Analyses of K-ras^Val12 × TAg^Wt bi-transgenic animals indicated that K-ras^Val12 can enhance this apoptosis threefold but only in cycling cells; increased apoptosis does not occur when K-ras^Val12 is expressed alone or with a TAg containing Glu^107,108→ Lys^107,108 substitutions that block its ability to bind pRB. Analysis of bi-transgenic K-ras^Val12 × TAg^Wt mice homozygous for wild-type or null p53 alleles established that the enhancement of apoptosis occurs through a p53-independent mechanism, is not attributable to augmented proliferation or to an increase in abortive cell cycle reentry (compared to TAg^Wt mice), and is not associated with detectable changes in the crypt–villus patterns of expression of apoptotic regulators (Bcl-2, Bcl-x_L, Bak, and Bax) or mediators of epithelial cell–matrix interactions and survival (e.g., α₅β₁ integrin and its ligand, fibronectin). Coexpression of K-ras^Val12 and TAg^Wt produces dysplasia. The K-ras^Val12-augmented apoptosis is unrelated to this dysplasia; enhanced apoptosis is also observed in cycling nondysplastic enterocytes that produce K-ras^Val12 and a TAg with a COOH-terminal truncation. The dysplastic epithelium of K-ras^Val12 × TAg^Wt mice does not develop neoplasms. Our results are consistent with this finding: (a) When expressed in initiated enterocytes with a proliferative abnormality, K-ras^Val12 facilitates progression to a dysplastic phenotype; (b) by diminishing cell survival on the villus, the oncoprotein may impede further progression; and (c) additional mutations may be needed to suppress this proapoptotic response to K-ras^Val12.     

Endothelial cell junctions

In the course of a freeze-cleave study on intercellular junctions in the regenerating rat liver, we observed an unusual array of intramembranous particles located in regions of contact between endothelial cells lining the hepatic sinusoids. These arrays were characterized by an accumulation of particles which resembled a zonula occludens in their linear deployment but differed in that the contact regions were composed of individual particles which remained separated from each other by regular particle-free intervals.