Leucine-rich diet supplementation modulates foetal muscle protein metabolism impaired by Walker-256 tumour

Background

Cancer-cachexia induces a variety of metabolic disorders of protein turnover and is more pronounced when associated with pregnancy. Tumour-bearing pregnant rats have impaired protein balance, which decreases protein synthesis and increases muscle breakdown. Because branched-chain amino acids, especially leucine, stimulate protein synthesis, we investigated the effect of a leucine-rich diet on protein metabolism in the foetal gastrocnemius muscles of tumour-bearing pregnant rats.

Methods

Foetuses of pregnant rats with or without Walker 256 tumours were divided into six groups. During the 20 days of the experiment, the pregnant groups were fed with either a control diet (C, control rats; W, tumour-bearing rats; Cp, rats pair-fed the same normoprotein-diet as the W group) or with a leucine-rich diet (L, leucine rats; LW, leucine tumour-bearing rats; and Lp, rats pair-fed the same leucine-rich diet as the LW group). After the mothers were sacrificed, the foetal gastrocnemius muscle samples were resected, and the protein synthesis and degradation and tissue chymotrypsin-like, cathepsin and calpain enzyme activities were assayed. The muscle oxidative enzymes (catalase, glutathione-S-transferase and superoxide dismutase), alkaline phosphatase enzyme activities and lipid peroxidation (malondialdehyde) were also measured.

Results

Tumour growth led to a reduction in foetal weight associated with decreased serum protein, albumin and glucose levels and low haematocrit in the foetuses of the W group, whereas in the LW foetuses, these changes were less pronounced. Muscle protein synthesis (measured by L-[3H]-phenylalanine incorporation) was reduced in the W foetuses but was restored in the LW group. Protein breakdown (as assessed by tyrosine release) was enhanced in the L and W groups, but chymotrypsin-like activity increased only in group W and tended toward an increase in the LW foetuses. The activity of cathepsin H was significantly higher in the W group foetuses, but the proteolytic calcium-dependent pathway showed similar enzyme activity. In parallel, an intense oxidative stress process was observed only in the group W foetuses.

Conclusions

These data suggested that the proteasomal and lysosomal proteolytic pathways and oxidative stress are likely to participate in the process of foetal muscle catabolism of Walker’s tumour-bearing pregnant rats. The present work shows that foetal muscle can be protected by supplementation with a leucine-rich diet.     

A substrate ambiguous enzyme facilitates genome reduction in an intracellular symbiont

Background

Genome evolution in intracellular microbial symbionts is characterized by gene loss, generating some of the smallest and most gene-poor genomes known. As a result of gene loss these genomes commonly contain metabolic pathways that are fragmented relative to their free-living relatives. The evolutionary retention of fragmented metabolic pathways in the gene-poor genomes of endosymbionts suggests that they are functional. However, it is not always clear how they maintain functionality. To date, the fragmented metabolic pathways of endosymbionts have been shown to maintain functionality through complementation by host genes, complementation by genes of another endosymbiont and complementation by genes in host genomes that have been horizontally acquired from a microbial source that is not the endosymbiont. Here, we demonstrate a fourth mechanism.

Results

We investigate the evolutionary retention of a fragmented pathway for the essential nutrient pantothenate (vitamin B5) in the pea aphid, Acyrthosiphon pisum endosymbiosis with Buchnera aphidicola. Using quantitative analysis of gene expression we present evidence for complementation of the Buchnera pantothenate biosynthesis pathway by host genes. Further, using complementation assays in an Escherichia coli mutant we demonstrate functional replacement of a pantothenate biosynthesis enzyme, 2-dehydropantoate 2-reductase (E.C. 1.1.1.169), by an endosymbiont gene, ilvC, encoding a substrate ambiguous enzyme.

Conclusions

Earlier studies have speculated that missing enzyme steps in fragmented endosymbiont metabolic pathways are completed by adaptable endosymbiont enzymes from other pathways. Here, we experimentally demonstrate completion of a fragmented endosymbiont vitamin biosynthesis pathway by recruitment of a substrate ambiguous enzyme from another pathway. In addition, this work extends host/symbiont metabolic collaboration in the aphid/Buchnera symbiosis from amino acid metabolism to include vitamin biosynthesis.

     

The Deuterator: software for the determination of backbone amide deuterium levels from H/D exchange MS data

Background  

The combination of mass spectrometry and solution phase amide hydrogen/deuterium exchange (H/D exchange) experiments is an effective method for characterizing protein dynamics, and protein-protein or protein-ligand interactions. Despite methodological advancements and improvements in instrumentation and automation, data analysis and display remains a tedious process. The factors that contribute to this bottleneck are the large number of data points produced in a typical experiment, each requiring manual curation and validation, and then calculation of the level of backbone amide exchange. Tools have become available that address some of these issues, but lack sufficient integration, functionality, and accessibility required to address the needs of the H/D exchange community. To date there is no software for the analysis of H/D exchange data that comprehensively addresses these issues.     

Hes1 Increases the Invasion Ability of Colorectal Cancer Cells via the STAT3-MMP14 Pathway

The Notch pathway contributes to self-renewal of tumor-initiating cell and inhibition of normal colonic epithelial cell differentiation. Deregulated expression of Notch1 and Jagged1 is observed in colorectal cancer. Hairy/enhancer of split (HES) family, the most characterized targets of Notch, involved in the development of many cancers. In this study, we explored the role of Hes1 in the tumorigenesis of colorectal cancer. Knocking down Hes1 induced CRC cell senescence and decreased the invasion ability, whereas over-expression of Hes1 increased STAT3 phosphorylation activity and up-regulated MMP14 protein level. We further explored the expression of Hes1 in human colorectal cancer and found high Hes1 mRNA expression is associated with poor prognosis in CRC patients. These findings suggest that Hes1 regulates the invasion ability through the STAT3-MMP14 pathway in CRC cells and high Hes1 expression is a predictor of poor prognosis of CRC.     

Generation and characterization of human insulin-releasing cell lines

Background  

The in vitro culture of insulinomas provides an attractive tool to study cell proliferation and insulin synthesis and secretion. However, only a few human beta cell lines have been described, with long-term passage resulting in loss of insulin secretion. Therefore, we set out to establish and characterize human insulin-releasing cell lines.     

A molecular basis for the physiological variation in shade avoidance responses: A tale of two ecotypes

Using two ecotypes of Stellaria longipes with contrasting responses to shade, we found that plants can differ in their responses to similar light cues, reflecting adaptations to their natural habitat. It was also observed that the plants could distinguish between distinct shade signals. Furthermore, the activity of wall modifying proteins, expansins and xyloglucan endotransglucosylase/hydrolase(s) (XTHs) was regulated during these responses. However, only expansin activity and gene expression profiles correlated with observed growth trends. The differential expression of expansins was light signal specific and ecotype specific and could account for both the trends in growth and their magnitude. We have thus established a potential molecular basis for the observed plasticity in responses to shade.Key words: shade avoidance, cell wall modification, expansins, XTHs, Stellaria longipes, phenotypic plasticity, light quality     

Zn(2+) modulation of neuronal transient K(+) current: fast and selective binding to the deactivated channels

Modulation of voltage-dependent transient K(+) currents (A type K(+) or K(A) current) by Zn(2+) was studied in rat hippocampal neurons by the whole-cell patch-clamp technique. It is found that Zn(2+) selectively binds to the resting (deactivated or closed) K(A) channels with a dissociation constant (K(d)) of approximately 3 &mgr;M, whereas the affinity between Zn(2+) and the inactivated K(A) channels is 1000-fold lower. Zn(2+) therefore produces a concentration-dependent shift of the K(A) channel inactivation curve and enhances the K(A) current elicited from relatively positive holding potentials. It is also found that the kinetics of Zn(2+) action are fast enough to compete with the transition rates between different gating states of the channel. The rapid and selective binding of Zn(2+) to the closed K(A) channels keeps the channel in the closed state and explains the ion's concentration-dependent slowing effect on the activation of K(A) current. This in turn accounts for the inhibitory effect of Zn(2+) on the K(A) current elicited from hyperpolarized holding potentials. Because the molecular mechanisms underlying these gating changes are kinetic interactions between the binding-unbinding of Zn(2+) and the intrinsic gating processes of the channel, the shift of the inactivation curve and slowing of K(A) channel activation are quantitatively correlated with ambient Zn(2+) over a wide concentration range without "saturation"; i.e., The effects are already manifest in micromolar Zn(2+), yet are not saturated even in millimolar Zn(2+). Because the physiological concentration of Zn(2+) could vary over a similarly wide range according to neural activities, Zn(2+) may be a faithful physiological "fine tuner," controlling and controlled by neural activities through its effect on the K(A) current.     

Longitudinal evaluation of repellent activity of Ocimum gratissimum (Labiatae) volatile oil against Simulium damnosum

To determine the repellent activity of Ocimum gratissimum volatile oil against Simulium damnosum (blackflies), a 12 month (January-December 2003) field study was conducted in three onchocerciasis endemic communities (Idomido, Obio camp, and Ikot Adaha) in Ini Local Government Area of Akwa Ibom State, Nigeria. The result revealed that topical application of 20% (v/v) concentration of the oil with liquid paraffin as a base, reduced the biting rate of S. damnosum by 90.2, 81.6, and 79.7%, in Idomido, Obiocamp, and Ikot Adaha respectively. The oil gave protection against the bite of S. damnosum for at least 3 h. A total of 710 adults S. damnosum were caught by individuals treated with Ocimum oil, as against 4296 caught by the control group. When the flies caught by the treated individuals were dissected none of them was infected with microfilariae of Onchocerca volvulus. Human-vector contact and onchocerciasis transmission could be reduced by the topical application of the volatile oil during the peak biting periods of the vector