Effect of Ceramide on Nonraft Proteins

The currently accepted model of biological membranes involves a heterogeneous, highly dynamic organization, where certain lipids and proteins associate to form cooperative platforms (“rafts”) for cellular signaling or transport processes. Ceramides, a lipid species occurring under conditions of cellular stress and apoptosis, are considered to stabilize these platforms, thus modulating cellular function. The present study focuses on a previously unrecognized effect of ceramide generation. In agreement with previous studies, we find that ceramide leads to a depletion of sphingomyelin from mixtures with palmitoyl oleoyl phosphatidylcholine bilayers, forming a ceramide–sphingomyelin-rich gel phase that coexists with a fluid phase rich in palmitoyl oleoyl phosphatidylcholine. Interestingly, however, this latter phase has an almost fourfold smaller bending rigidity compared to a sphingomyelin–palmitoyl oleoyl phosphatidylcholine mixture lacking ceramide. The significant change of membrane bulk properties can have severe consequences for conformational equilibria of membrane proteins. We discuss these effects in terms of the lateral pressure profile concept for a simple geometric model of an ion channel and find a significant inhibition of its activity.     

Mutations in the SPTLC2 subunit of serine palmitoyltransferase cause hereditary sensory and autonomic neuropathy type I

Hereditary sensory and autonomic neuropathy type I (HSAN-I) is an axonal peripheral neuropathy associated with progressive distal sensory loss and severe ulcerations. Mutations in the first subunit of the enzyme serine palmitoyltransferase (SPT) have been associated with HSAN-I. The SPT enzyme catalyzes the first and rate-limiting step in the de novo sphingolipid synthesis pathway. However, different studies suggest the implication of other genes in the pathology of HSAN-I. Therefore, we screened the two other known subunits of SPT, SPTLC2 and SPTLC3, in a cohort of 78 HSAN patients. No mutations were found in SPTLC3, but we identified three heterozygous missense mutations in the SPTLC2 subunit of SPT in four families presenting with a typical HSAN-I phenotype. We demonstrate that these mutations result in a partial to complete loss of SPT activity in vitro and in vivo. Moreover, they cause the accumulation of the atypical and neurotoxic sphingoid metabolite 1-deoxy-sphinganine. Our findings extend the genetic heterogeneity in HSAN-I and enlarge the group of HSAN neuropathies associated with SPT defects. We further show that HSAN-I is consistently associated with an increased formation of the neurotoxic 1-deoxysphinganine, suggesting a common pathomechanism for HSAN-I.     

Functional Gene Group Analysis Reveals a Role of Synaptic Heterotrimeric G Proteins in Cognitive Ability

Although cognitive ability is a highly heritable complex trait, only a few genes have been identified, explaining relatively low proportions of the observed trait variation. This implies that hundreds of genes of small effect may be of importance for cognitive ability. We applied an innovative method in which we tested for the effect of groups of genes defined according to cellular function (functional gene group analysis). Using an initial sample of 627 subjects, this functional gene group analysis detected that synaptic heterotrimeric guanine nucleotide binding proteins (G proteins) play an important role in cognitive ability (P_EMP = 1.9 × 10⁻⁴). The association with heterotrimeric G proteins was validated in an independent population sample of 1507 subjects. Heterotrimeric G proteins are central relay factors between the activation of plasma membrane receptors by extracellular ligands and the cellular responses that these induce, and they can be considered a point of convergence, or a “signaling bottleneck.” Although alterations in synaptic signaling processes may not be the exclusive explanation for the association of heterotrimeric G proteins with cognitive ability, such alterations may prominently affect the properties of neuronal networks in the brain in such a manner that impaired cognitive ability and lower intelligence are observed. The reported association of synaptic heterotrimeric G proteins with cognitive ability clearly points to a new direction in the study of the genetic basis of cognitive ability.     

Expression and function of the empty spiracles gene in olfactory sense organ development of Drosophila melanogaster

In Drosophila, the cephalic gap gene empty spiracles plays key roles in embryonic patterning of the peripheral and central nervous system. During postembryonic development, it is involved in the development of central olfactory circuitry in the antennal lobe of the adult. However, its possible role in the postembryonic development of peripheral olfactory sense organs has not been investigated. Here, we show that empty spiracles acts in a subset of precursors that generate the olfactory sense organs of the adult antenna. All empty spiracles-expressing precursor cells co-express the proneural gene amos and the early patterning gene lozenge. Moreover, the expression of empty spiracles in these precursor cells is dependent on both amos and lozenge. Functional analysis reveals two distinct roles of empty spiracles in the development of olfactory sense organs. Genetic interaction studies in a lozenge-sensitized background uncover a requirement of empty spiracles in the formation of trichoid and basiconic olfactory sensilla. MARCM-based clonal mutant analysis reveals an additional role during axonal targeting of olfactory sensory neurons to glomeruli within the antennal lobe. Our findings on empty spiracles action in olfactory sense organ development complement previous studies that demonstrate its requirement in olfactory interneurons and, taken together with studies on the murine homologs of empty spiracles, suggest that conserved molecular genetic programs might be responsible for the formation of both peripheral and central olfactory circuitry in insects and mammals.     

A nationwide population-based skin cancer screening in Germany: Proceedings of the first meeting of the International Task Force on Skin Cancer Screening and Prevention (September 24 and 25, 2009)

Skin cancer incidence is increasing worldwide in white populations and mortality rates have not declined throughout most of the world. An extraordinarily high proportion of at-risk individuals have yet to be screened for melanoma but guidelines from esteemed bodies do not currently endorse population-based screening. Evidence for the effectiveness of skin cancer screening is imperative. To this end, scientists in Germany have launched a nationwide skin cancer screening campaign. Herein, we review pilot screening data from Schleswig-Holstein, discuss the launch of the major new national initiative, review issues related to evaluation of that program, and propose seven recommendations from the International Task Force on Skin Cancer Screening and Prevention that was held in Hamburg, Germany, on September 24 and 25, 2009.     

Multicentric validation of proteomic biomarkers in urine specific for diabetic nephropathy

Background

Urine proteome analysis is rapidly emerging as a tool for diagnosis and prognosis in disease states. For diagnosis of diabetic nephropathy (DN), urinary proteome analysis was successfully applied in a pilot study. The validity of the previously established proteomic biomarkers with respect to the diagnostic and prognostic potential was assessed on a separate set of patients recruited at three different European centers. In this case-control study of 148 Caucasian patients with diabetes mellitus type 2 and duration ≥5 years, cases of DN were defined as albuminuria >300 mg/d and diabetic retinopathy (n = 66). Controls were matched for gender and diabetes duration (n = 82).

Methodology/Principal Findings

Proteome analysis was performed blinded using high-resolution capillary electrophoresis coupled with mass spectrometry (CE-MS). Data were evaluated employing the previously developed model for DN. Upon unblinding, the model for DN showed 93.8% sensitivity and 91.4% specificity, with an AUC of 0.948 (95% CI 0.898-0.978). Of 65 previously identified peptides, 60 were significantly different between cases and controls of this study. In <10% of cases and controls classification by proteome analysis not entirely resulted in the expected clinical outcome. Analysis of patient''s subsequent clinical course revealed later progression to DN in some of the false positive classified DN control patients.

Conclusions

These data provide the first independent confirmation that profiling of the urinary proteome by CE-MS can adequately identify subjects with DN, supporting the generalizability of this approach. The data further establish urinary collagen fragments as biomarkers for diabetes-induced renal damage that may serve as earlier and more specific biomarkers than the currently used urinary albumin.     

Contribution of marrow-derived progenitors to vascular and cardiac regeneration

Adult bone marrow is a rich reservoir of tissue-specific pluripotent stem and progenitor cells. Accumulating evidence suggest that these cells have the potential of contributing to tissue revascularization and cardiac regeneration. Physiological stress results in the release of specific chemokines and cytokines that promote mobilization of stem cells to the peripheral circulation. Incorporation of these mobilized cells contributes to formation of functional vasculature and sets up stage for tissue regeneration. Vascular Endothelial Growth Factor (VEGF) through interaction with its receptors VEGFR2 and VEGFR1 expressed on endothelial and hematopoietic stem cells promote recruitment of these cells into the sites of tissue injury accelerating vascular healing. Similarly, subset of CD34 + marrow derived cells are mobilized and recruited to the ischemic myocardium, differentiating into cardiac and vascular cells, restoring cardiac function. Identification of cellular mediators and tissue specific chemocytokines that facilitate selective recruitment of marrow-derived stem and progenitor cells to specific organs, will open up new avenues to accelerate cardiovascular regeneration and tissue revascularization.     

Phosphoinositide 3-kinase is required for process outgrowth and cell polarization of gastrulating mesendodermal cells

BACKGROUND: During vertebrate gastrulation, cell polarization and migration are core components in the cellular rearrangements that lead to the formation of the three germ layers, ectoderm, mesoderm, and endoderm. Previous studies have implicated the Wnt/planar cell polarity (PCP) signaling pathway in controlling cell morphology and movement during gastrulation. However, cell polarization and directed cell migration are reduced but not completely abolished in the absence of Wnt/PCP signals; this observation indicates that other signaling pathways must be involved. RESULTS: We show that Phosphoinositide 3-Kinases (PI3Ks) are required at the onset of zebrafish gastrulation in mesendodermal cells for process formation and cell polarization. Platelet Derived Growth Factor (PDGF) functions upstream of PI3K, while Protein Kinase B (PKB), a downstream effector of PI3K activity, localizes to the leading edge of migrating mesendodermal cells. In the absence of PI3K activity, PKB localization and cell polarization are strongly reduced in mesendodermal cells and are followed by slower but still highly coordinated and directed movements of these cells. CONCLUSIONS: We have identified a novel role of a signaling pathway comprised of PDGF, PI3K, and PKB in the control of morphogenetic cell movements during gastrulation. Furthermore, our findings provide insight into the relationship between cell polarization and directed cell migration at the onset of zebrafish gastrulation.     

Metamorphosis of Hydractinia echinata (Cnidaria) is caspase-dependent

Apoptotic cell death plays an important role in many developmental pathways in multicellular animals. Here, we show that metamorphosis in the basal invertebrate Hydractinia echinata (Cnidaria) depends on the activity of caspases, the central enzymes in apoptosis. Caspases are activated during metamorphosis and this activity can be measured with caspase-3 specific fluorogenic substrates. In affinity labelling experiments 23/25 kDa bands were obtained, which represented active caspase. Specific inhibition of caspase activity with caspase-3 inhibitors abolished metamorphosis completely, reversibly and in a dose-dependent manner. This suggests that caspase activity is indispensable for metamorphosis in Hydractinia echinata.