Complete genome sequence of the myxobacterium Sorangium cellulosum

The genus Sorangium synthesizes approximately half of the secondary metabolites isolated from myxobacteria, including the anti-cancer metabolite epothilone. We report the complete genome sequence of the model Sorangium strain S. cellulosum So ce56, which produces several natural products and has morphological and physiological properties typical of the genus. The circular genome, comprising 13,033,779 base pairs, is the largest bacterial genome sequenced to date. No global synteny with the genome of Myxococcus xanthus is apparent, revealing an unanticipated level of divergence between these myxobacteria. A large percentage of the genome is devoted to regulation, particularly post-translational phosphorylation, which probably supports the strain's complex, social lifestyle. This regulatory network includes the highest number of eukaryotic protein kinase-like kinases discovered in any organism. Seventeen secondary metabolite loci are encoded in the genome, as well as many enzymes with potential utility in industry.     

Isolation and Characterization of a Hybrid Respiratory Supercomplex Consisting of Mycobacterium tuberculosis Cytochrome bcc and Mycobacterium smegmatis Cytochrome aa3

Recently, energy production pathways have been shown to be viable antitubercular drug targets to combat multidrug-resistant tuberculosis and eliminate pathogen in the dormant state. One family of drugs currently under development, the imidazo[1,2-a]pyridine derivatives, is believed to target the pathogen''s homolog of the mitochondrial bc₁ complex. This complex, denoted cytochrome bcc, is highly divergent from mitochondrial Complex III both in subunit structure and inhibitor sensitivity, making it a good target for drug development. There is no soluble cytochrome c in mycobacteria to transport electrons from the bcc complex to cytochrome oxidase. Instead, the bcc complex exists in a “supercomplex” with a cytochrome aa₃-type cytochrome oxidase, presumably allowing direct electron transfer. We describe here purification and initial characterization of the mycobacterial cytochrome bcc-aa₃ supercomplex using a strain of M. smegmatis that has been engineered to express the M. tuberculosis cytochrome bcc. The resulting hybrid supercomplex is stable during extraction and purification in the presence of dodecyl maltoside detergent. It is hoped that this purification procedure will potentiate functional studies of the complex as well as crystallographic studies of drug binding and provide structural insight into a third class of the bc complex superfamily.     

Mechanical ventilation with high tidal volumes attenuates myocardial dysfunction by decreasing cardiac edema in a rat model of LPS-induced peritonitis

Background

Injurious mechanical ventilation (MV) may augment organ injury remote from the lungs. During sepsis, myocardial dysfunction is common and increased endothelial activation and permeability can cause myocardial edema, which may, among other factors, hamper myocardial function. We investigated the effects of MV with injuriously high tidal volumes on the myocardium in an animal model of sepsis.

Methods

Normal rats and intraperitoneal (i.p.) lipopolysaccharide (LPS)-treated rats were ventilated with low (6 ml/kg) and high (19 ml/kg) tidal volumes (Vt) under general anesthesia. Non-ventilated animals served as controls. Mean arterial pressure (MAP), central venous pressure (CVP), cardiac output (CO) and pulmonary plateau pressure (P_plat) were measured. Ex vivo myocardial function was measured in isolated Langendorff-perfused hearts. Cardiac expression of endothelial vascular cell adhesion molecule (VCAM)-1 and edema were measured to evaluate endothelial inflammation and leakage.

Results

MAP decreased after LPS-treatment and Vt-dependently, both independent of each other and with interaction. MV Vt-dependently increased CVP and Pplat and decreased CO. LPS-induced peritonitis decreased myocardial function ex vivo but MV attenuated systolic dysfunction Vt-dependently. Cardiac endothelial VCAM-1 expression was increased by LPS treatment independent of MV. Cardiac edema was lowered Vt-dependently by MV, particularly after LPS, and correlated inversely with systolic myocardial function parameters ex vivo.

Conclusion

MV attenuated LPS-induced systolic myocardial dysfunction in a Vt-dependent manner. This was associated with a reduction in cardiac edema following a lower transmural coronary venous outflow pressure during LPS-induced coronary inflammation.     

Sarcocystis neurona: molecular characterization of enolase domain I region and a comparison to other protozoa

Sarcocystis neurona causes protozoal myeloencephalitis and has the ability to infect a wide host range in contrast to other Sarcocystis species. In the current study, five S. neurona isolates from a variety of sources, three Sarcocystis falcatula, one Sarcocystis dasypi/S. neurona-like isolate, and one Besnoitia darlingi isolate were used to compare the enolase 2 gene segment containing the domain I region to previously sequenced enolase genes from Neospora caninum, Neospora hughesi, Toxoplasma gondii, Plasmodium falciparum, and Trypanosoma cruzi; enolase 2 segment containing domain I region is highly conserved amongst these parasites of veterinary and medical importance. Immunohistochemistry results indicates reactivity of T. gondii enolase 1 and 2 antibodies to S. neurona merozoites and metrocytes, but no reactivity of anti-enolase 1 to the S. neurona bradyzoite stage despite reactivity to T. gondii bradyzoites, suggesting expression differences between organisms.     

Annual variation in source contributions to a mixed stock: implications for quantifying connectivity

Connectivity among populations of highly migratory species is an area of active research and is often quantified with genetic markers. We determined mitochondrial DNA (mtDNA) sequences in 350 green turtles, Chelonia mydas, in 10 annual samples over a 12-year period from an aggregation of immature green turtles in the southern Bahamas. We found significant temporal structuring in haplotype frequencies among years for all turtles and for recruits. These significant differences were reflected in substantial variation in the relative contributions from different rookeries among years estimated by a Bayesian hierarchical model. Because this foraging aggregation has been the subject of a demographic study for over 30 years, we were able to determine that, among the three potential causes of temporal structuring-differential recruitment, mortality and emigration-recruitment accounts for most of this variation. We found that estimates of connectivity and genetic diversity in sea turtle populations are affected by the level of temporal variation reported here. More studies on the extent of temporal variation in composition of mixed stocks of other migratory species are needed to determine how this affects measures of connectivity.     

Structure of the DNA-bound BRCA1 C-terminal Region from Human Replication Factor C p140 and Model of the Protein-DNA Complex

BRCA1 C-terminal domain (BRCT)-containing proteins are found widely throughout the animal and bacteria kingdoms where they are exclusively involved in cell cycle regulation and DNA metabolism. Whereas most BRCT domains are involved in protein-protein interactions, a small subset has bona fide DNA binding activity. Here, we present the solution structure of the BRCT region of the large subunit of replication factor C bound to DNA and a model of the structure-specific complex with 5′-phosphorylated double-stranded DNA. The replication factor C BRCT domain possesses a large basic patch on one face, which includes residues that are structurally conserved and ligate the phosphate in phosphopeptide binding BRCT domains. An extra α-helix at the N terminus, which is required for DNA binding, inserts into the major groove and makes extensive contacts to the DNA backbone. The model of the protein-DNA complex suggests 5′-phosphate recognition by the BRCT domains of bacterial NAD⁺-dependent ligases and a nonclamp loading role for the replication factor C complex in DNA transactions.     

Association Between Na<Superscript>+</Superscript>,K<Superscript>+</Superscript>-ATPase Activity and the Vulnerability/Resilience to Mood Disorders induced by Early Life Experience

There is increasing evidence that early life events can influence neurodevelopment and later susceptibility to disease. Chronic variable stress (CVS) has been used as a model of depression. The objective of this study was to evaluate the interaction between early experience and vulnerability to chronic variable stress in adulthood, analyzing emotional, metabolic and neurochemical aspects related to depression. Pups were (1) handled (10 min/day) or (2) left undisturbed from day 1 to 10 after birth. When the animals reached adulthood, the groups were subdivided and the rats were submitted or not to CVS, which consisted of daily exposure to different stressors for 40 days, followed by a period of behavioral tasks, biochemical (plasma corticosterone and insulin sensitivity) and neurochemical (Na⁺,K⁺-ATPase activity in hippocampus, amygdala and parietal cortex) measurements. Neonatally-handled rats demonstrated shorter immobility times in the forced swimming test, independently of the stress condition. There was no difference concerning basal corticosterone or insulin sensitivity between the groups. Na⁺,K⁺-ATPase activity was decreased in hippocampus and increased in the amygdala of neonatally-handled rats. CVS decreased the enzyme activity in the three structures, mainly in the non-handled group. These findings suggest that early handling increases the ability to cope with chronic variable stress in adulthood, with animals showing less susceptibility to neurochemical features associated with depression, confirming the relevance of the precocious environment to vulnerability to psychiatric conditions in adulthood.     

Living with myotonic dystrophy; what can be learned from couples? a qualitative study

Background  

Myotonic dystrophy type 1 (MD1) is one of the most prevalent neuromuscular diseases, yet very little is known about how MD1 affects the lives of couples and how they themselves manage individually and together. To better match health care to their problems, concerns and needs, it is important to understand their perspective of living with this hereditary, systemic disease.