Identification of 3-hydroxypalmitic acid methyl ester as a novel autoregulator controlling virulence in Ralstonia solanacearum

Expression of virulence genes in Ralstonia solanacearum , a phytopathogenic bacterium, is controlled by a complex regulatory network that integrates multiple signal inputs. Production of several virulence determinants is co-ordinately reduced by inactivation of phcB , but is restored by growth in the presence of a volatile extracellular factor (VEF) produced by wild-type strains of R. solanacearum . The VEF was purified from spent culture broth by distillation, solvent extraction, and liquid chromatography. Gas chromatography and mass spectroscopy identified 3-hydroxypalmitic acid methyl ester (3-OH PAME) as the major component in the single peak of VEF activity. Authentic 3-OH PAME and the purified VEF were active at ≤1 nM, and had nearly equivalent specific activities for stimulating the expression of eps (the biosynthetic locus for extracellular polysaccharide) in a phcB mutant. Authentic 3-OH PAME also increased the production of three virulence factors by a phcB mutant over 20-fold to wild-type levels, restored normal cell density-associated expression of eps and increased expression of eps when delivered via the vapour phase. Reanalysis of the PhcB amino acid sequence suggested that it is a small-molecule S -adenosylmethionine-dependent methyltransferase, which might catalyse synthesis of 3-OH PAME from a naturally occurring fatty acid. Biologically active concentrations of extracellular 3-OH PAME were detected before the onset of eps expression, suggesting that it is an intercellular signal that autoregulates virulence gene expression in wild-type R. solanacearum . Other than acyl-homoserine lactones, 3-OH PAME is the only endogenous fatty acid derivative shown to be an autoregulator and may be the first example of a new family of compounds that can mediate long-distance intercellular communication.     

Expression of the IL-7 Receptor Alpha-Chain Is Down Regulated on the Surface of CD4 T-Cells by the HIV-1 Tat Protein

HIV infection elicits defects in CD4 T-cell homeostasis in both a quantitative and qualitative manner. Interleukin-7 (IL-7) is essential to T-cell homeostasis and several groups have shown reduced levels of the IL-7 receptor alpha-chain (CD127) on both CD4 and CD8 T-cells in viremic HIV+ patients. We have shown previously that soluble HIV Tat protein specifically down regulates cell surface expression of CD127 on human CD8 T-cells in a paracrine fashion. The effects of Tat on CD127 expression in CD4 T-cells has yet to be described. To explore this effect, CD4 T-cells were isolated from healthy individuals and expression levels of CD127 were examined on cells incubated in media alone or treated with Tat protein. We show here that, similar to CD8 T-cells, the HIV-1 Tat protein specifically down regulates CD127 on primary human CD4 T-cells and directs the receptor to the proteasome for degradation. Down regulation of CD127 in response to Tat was seen on both memory and naive CD4 T-cell subsets and was blocked using either heparin or anti-Tat antibodies. Tat did not induce apoptosis in cultured primary CD4 T-cells over 72 hours as determined by Annexin V and PI staining. Pre-incubation of CD4 T-cells with HIV-1 Tat protein did however reduce the ability of IL-7 to up regulate Bcl-2 expression. Similar to exogenous Tat, endogenously expressed HIV Tat protein also suppressed CD127 expression on primary CD4 T-cells. In view of the important role IL-7 plays in lymphocyte proliferation, homeostasis and survival, down regulation of CD127 by Tat likely plays a central role in immune dysregulation and CD4 T-cell decline. Understanding this effect could lead to new approaches to mitigate the CD4 T-cell loss evident in HIV infection.     

Substituent effects on the binding of ethidium and its derivatives to natural DNA

The binding of eight ethidium derivatives to short (approximately 35 base-pair), random sequence DNA has been investigated using 1H-NMR. At 35 degrees C, all drugs cause upfield shifts of the DNA imino proton resonances characteristic of intercalative binding to DNA, but the line shapes vary significantly with the nature of the drug. The results confirm our previous proposal that removal of the amino group at position-3, but not at position-8, on the parent ethidium shortens the lifetime of the intercalative state (less than 1-2 ms at 35 degrees C). These results suggest that hydrogen-bonding interactions with the 3-NH2 group are involved in stabilization of the drug-DNA complex or that changes in charge distribution that accompany removal of the 3-NH2 group reduce the complex stability. The magnitude of the shift of the drug-DNA spectra indicates a slight preference for binding of the drugs adjacent to G X C base-pairs.     

The Network Structure of Symptoms of the Diagnostic and Statistical Manual of Mental Disorders

Although current classification systems have greatly contributed to the reliability of psychiatric diagnoses, they ignore the unique role of individual symptoms and, consequently, potentially important information is lost. The network approach, in contrast, assumes that psychopathology results from the causal interplay between psychiatric symptoms and focuses specifically on these symptoms and their complex associations. By using a sophisticated network analysis technique, this study constructed an empirically based network structure of 120 psychiatric symptoms of twelve major DSM-IV diagnoses using cross-sectional data of the National Epidemiologic Survey on Alcohol and Related Conditions (NESARC, second wave; N = 34,653). The resulting network demonstrated that symptoms within the same diagnosis showed differential associations and indicated that the strategy of summing symptoms, as in current classification systems, leads to loss of information. In addition, some symptoms showed strong connections with symptoms of other diagnoses, and these specific symptom pairs, which both concerned overlapping and non-overlapping symptoms, may help to explain the comorbidity across diagnoses. Taken together, our findings indicated that psychopathology is very complex and can be more adequately captured by sophisticated network models than current classification systems. The network approach is, therefore, promising in improving our understanding of psychopathology and moving our field forward.     

Gastrocnemius medialis muscle architecture and physiological cross sectional area in adult males with Duchenne muscular dystrophy

Objectives:

To describe muscle size and architecture of the gastrocnemius medialis (GM) muscle in eleven adult males with Duchenne Muscular Dystrophy (DMD, age 24.5±5.4 years), and a control group of eleven males without DMD (CTRL, age 22.1±0.9 years).

Methods:

GM anatomical cross sectional area (ACSA), volume (VOL), physiological cross sectional area (PCSA), fascicle length (Lf) and pennation angle (θ) were assessed using B-Mode Ultrasonography. GM ACSA was measured at 25, 50 and 75% of muscle length (Lm), from which VOL was calculated. At 50% of Lm, sagittal plane images were analysed to determine GM Lf and θ. GM PCSA was calculated as VOL/Lf. The ratio of Lf and Lm was also calculated.

Results:

GM ACSA at 50% Lm, VOL and PCSA were smaller in DMD males compared to CTRL males by 36, 47 and 43%, respectively (P<0.01). There were no differences in Lf and θ. GM Lm was 29% shorter in DMD compared to CTRL. Lf/Lm was 29% longer in DMD (P<0.01).

Conclusions:

Unlike previous data in children with DMD, our results show significant atrophy in adult males with DMD, and no change in Lf or θ. The shorter Lm may have implications for joint flexibility.