A new piece of an old jigsaw: glucose kinase is activated posttranslationally in a glucose transport-dependent manner in streptomyces coelicolor A3(2)

Members of the soil-dwelling prokaryotic genus Streptomyces are indispensable for the recycling of complex polysaccharides, and produce a wide range of natural products. Nutrient limitation is likely to be a major signal for the onset of their development, resulting in spore formation by specialized aerial hyphae. Streptomycetes grow on numerous carbon sources, which they utilize in a preferential manner. The main signaling pathway underlying this phenomenon is carbon catabolite repression, which in streptomycetes is totally dependent on the glycolytic enzyme glucose kinase (Glk). How Glk exerts this fascinating dual role (metabolic and regulatory) is still largely a mystery. We show here that while Glk is made constitutively throughout the growth of Streptomyces coelicolor A3(2), its catalytic activity is modulated in a carbon source-dependent manner: while cultures growing exponentially on glucose exhibit high Glk activity, mannitol- grown cultures show negligible activity. Glk activity was directly proportional to the amount of two Glk isoforms observed by Western blot analysis. The activity profile of GlcP, the major glucose permease, correlated very well with that of Glk. Our data are consistent with a direct interaction between Glk and GlcP, suggesting that a Glk-GlcP permease complex is required for efficient glucose transport by metabolic trapping. This is supported by the strongly reduced accumulation of glucose in glucose kinase mutants. A model to explain our data is presented.     

Pseudoxanthoma Elasticum: Cardiac Findings in Patients and Abcc6-Deficient Mouse Model

Background

Pseudoxanthoma elasticum (PXE), caused by mutations in the ABCC6 gene, is a rare multiorgan disease characterized by the mineralization and fragmentation of elastic fibers in connective tissue. Cardiac complications reportedly associated with PXE are mainly based on case reports.

Methods

A cohort of 67 PXE patients was prospectively assessed. Patients underwent physical examination, electrocardiogram, transthoracic echocardiography, cardiac magnetic resonance imaging (CMR), treadmill testing, and perfusion myocardial scintigraphy (SPECT). Additionally, the hearts of a PXE mouse models (Abcc6^−/−) and wild-type controls (WT) were analyzed.

Results

Three patients had a history of proven coronary artery disease. In total, 40 patients underwent exercise treadmill tests, and 28 SPECT. The treadmill tests were all negative. SPECT showed mild perfusion abnormalities in two patients. Mean left ventricular (LV) dimension and function values were within the normal range. LV hypertrophy was found in 7 (10.4%) patients, though the hypertrophy etiology was unknown for 3 of those patients. Echocardiography revealed frequent but insignificant mitral and tricuspid valvulopathies. Mitral valve prolapse was present in 3 patients (4.5%). Two patients exhibited significant aortic stenosis (3.0%). While none of the functional and histological parameters diverged significantly between the Abcc6^−/− and WT mice groups at age of 6 and 12 months, the 24-month-old Abcc6^−/− mice developed cardiac hypertrophy without contractile dysfunction.

Conclusions

Despite sporadic cases, PXE does not appear to be associated with frequent cardiac complications. However, the development of cardiac hypertrophy in the 24-month-old Abcc6^−/− mice suggests that old PXE patients might be prone to developing late cardiopathy.     

A Major Gene for Resting Metabolic Rate Unassociated with Body Composition: Results from the Québec Family Study

A major gene hypothesis for resting metabolic rate (RMR) was investigated using segregation analysis (POINTER) of data on families participating in Phase 2 of the Québec Family Study. Complete analyses were conducted on RMR adjusted for age, and also on RMR adjusted for age and other covariates, primarily fat mass (FM) and fat-free mass (FFM). Prior to adjustment for covariates, support for a major gene hypothesis was equivocal — i.e., there was evidence for either a major gene or a multifactorial component (i.e., polygenic and/or familial environment). The multifactorial model was preferred over the major gene model, although the latter did segregate according to Mendelian expectations. However, after the effects of FM and FFM were accounted for, a major gene effect was unambiguous and compelling. The putative locus accounted for 57% of the variance, affected 7% of the sample, and led to high values of RMR. The lack of a significant multifactorial effect suggested that the familial etiology of RMR adjusted for FM and FFM was likely to be entirely a function of the major locus. Comparing the RMR results from pre- and post-adjustment for FM and FFM suggests a plausible hypothesis. We know from earlier studies in this sample that there is a putative major gene for FM and a major non-Mendelian effect for FFM. The current study leads us to speculate that: (1) the gene(s) affecting body size and body composition also may have an effect on RMR, and further (2) removal of the effect of the major gene(s) for body size and composition allowed for detection of an additional major gene affecting only the RMR. Thus, RMR appears to be an oligogenic trait.     

Detection of refractive photokeratectomy traces during eye banking: impossible with organ culture but possible with an active storage machine: case report

Cell and Tissue Banking - The detection of corneas operated on for refractive surgery [LASIK or photorefractive keratectomy (PRK)] will become a major concern for eye banks in the coming years...     

In vivo simulation of human pharmacokinetics in the rabbit

The evaluation of drugs in vivo is often based on experimental models using small animals such as mice, rats and rabbits. However, these models could be improved to correspond more closely to the human situation if the pharmacokinetics of the drugs tested in animals were similar to that observed in humans. The use of a computer-controlled pump allowing an adequate flow of tobramycin and amikacin to be infused into rabbits enabled us to simulate the human pharmacokinetics of these antibiotics in vivo in this study. The function defining the rate of infusion required to perform the simulation of an intravenous bolus was first determined generally and symbolically for linear pharmacokinetic models independently from the number of compartments involved. The practical simulation of a decreasing monoexponential serum profile with a half-life of 2 h (one-compartment model for the human pharmacokinetics of aminoglycosides) was then studied for tobramycin and amikacin on the basis of a two-compartment model in the animal. The kinetics obtained had an apparent elimination half-life of 1.97 and 1.86 h, respectively. Linearity of the semilogarithmic regressions of the profiles obtained was quite sound. Finally, an a posteriori analysis of the pharmacokinetic model and its parameters is proposed on the basis of the results obtained after simulation.     

Mek2 is dispensable for mouse growth and development

MEK is a dual-specificity kinase that activates the extracellular signal-regulated kinase (ERK) mitogen-activated protein (MAP) kinase upon agonist binding to receptors. The ERK/MAP kinase cascade is involved in cell fate determination in many organisms. In mammals, this pathway is proposed to regulate cell growth and differentiation. Genetic studies have shown that although a single Mek gene is present in Caenorhabditis elegans, Drosophila melanogaster, and Xenopus laevis, two Mek homologs, Mek1 and Mek2, are present in the mammalian cascade. The inactivation of the Mek1 gene leads to embryonic lethality and has revealed the unique role played by Mek1 during embryogenesis. To investigate the biological function of the second homolog, we have generated mice deficient in Mek2 function. Mek2 mutant mice are viable and fertile, and they do not present flagrant morphological alteration. Although several components of the ERK/MAP kinase cascade have been implicated in thymocyte development, no such involvement was observed for MEK2, which appears to be nonessential for thymocyte differentiation and T-cell-receptor-induced proliferation and apoptosis. Altogether, our findings demonstrate that MEK2 is not necessary for the normal development of the embryo and T-cell lineages, suggesting that the loss of MEK2 can be compensated for by MEK1.     

Structure of a highly phosphorylated lipopolysaccharide core in the Delta algC mutants derived from Pseudomonas aeruginosa wild-type strains PAO1 (serogroup O5) and PAC1R (serogroup O3)

Lipopolysaccharides (LPS) were isolated from rough-type mutant strains of Pseudomonas aeruginosa (Delta algC) derived from wild-type strains PAO1 (serogroup O5) and PAC1R (serogroup O3). Structural studies of the LPS core region with a special focus on the phosphorylation pattern were performed by 2D NMR spectroscopy, including a 1H,(31)P HMQC-TOCSY experiment, MALDI-TOF MS, and Fourier-transform ion cyclotron resonance ESIMS using the capillary skimmer dissociation technique. Both LPS were found to contain two residues each of 3-deoxy-D-manno-oct-2-ulosonic acid (Kdo) and L-glycero-D-manno-heptose (Hep), one residue of N-(L-alanyl)-D-galactosamine and one O-carbamoyl group (Cm) on the distal Hep residue. The following structures of a tetrasaccharide trisphosphate from strain PAC1R Delta algC and that carrying an additional ethanolamine phosphate group (PEtN) from strain PAO1 Delta algC were elucidated: [carbohydrate structre: see text] where R=P in PAC1R Delta algC and PPEtN in PAO1 Delta algC. To our knowledge, in this work the presence of ethanolamine diphosphate is unambiguously confirmed and its position established for the first time in the LPS core of a rough-type strain of P. aeruginosa. In addition, the structure of the complete LPS core of wild-type strain P. aeruginosa PAO1 was reinvestigated and the position of the phosphorylation sites was revised.