Insights into autosomal dominant polycystic kidney disease by quantitative mass spectrometry-based proteomics

Autosomal dominant polycystic kidney disease (ADPKD) is a common monogenetic disorder that is caused by mutations in the genes PKD1 and PKD2 encoding polycystin-1 and polycystin-2, respectively. Polycystin-1 and -2 form a complex, interact with several proteins involved in signal transduction and localize to discrete subcellular positions, most importantly the primary cilium. Whereas the causative mutations leading to ADPKD are known, the underlying deregulated cellular pathways are not well understood. In the current review, we introduce state-of-the-art mass spectrometry (MS)-based proteomic techniques and summarize their use in kidney and ADPKD research. Proteomic profiling approaches, the elucidation of ADPKD- relevant protein-protein interactions and the regulation of posttranslational modifications are included. We also discuss the use of MS-based methods for ADPKD prognosis, diagnosis and disease monitoring by using protein- and peptide-based biomarkers

Early loading of plalatal implants (ortho-type II) a prospective multicenter randomized controlled clinical trial

<p>Abstract</p> <p>Background</p> <p>In orthodontic treatment, anchorage control is a fundamental aspect. Usually conventional mechanism for orthodontic anchorage control can be either extraoral or intraoral that is headgear or intermaxillary elastics. Their use are combined with various side effects such as tipping of occlusal plane or undesirable movements of teeth. Especially in cases, where key-teeth are missing, conventional anchorage defined as tooth-borne anchorage will meet limitations. Therefore, the use of endosseous implants for anchorage purposes are increasingly used to achieve positional stability and maximum anchorage.</p> <p>Methods/Design</p> <p>The intended study is designed as a prospective, multicenter randomized controlled trial (RCT), comparing and contrasting the effect of early loading of palatal implant therapy versus implant loading after 12 weeks post implantation using the new ortho-implant type II anchor system device (Orthosystem Straumann, Basel, Switzerland).</p> <p>124 participants, mainly adult males or females, whose diagnoses require temporary stationary implant-based anchorage treatment will be randomized 1:1 to one of two treatment groups: group 1 will receive a loading of implant standard therapy after a healing period of 12 week (gold standard), whereas group 2 will receive an early loading of orthodontic implants within 1 week after implant insertion. Participants will be at least followed for 12 months after implant placement.</p> <p>The primary endpoint is to investigate the behavior of early loaded palatal implants in order to find out if shorter healing periods might be justified to accelerate active orthodontic treatment. Secondary outcomes will focus e.g. on achievement of orthodontic treatment goals and quantity of direct implant-bone interface of removed bone specimens. As tertiary objective, a histologic and microtomography evaluation of all retrieved implants will be performed to obtain data on the performance of the SLA surface in human bone evaluation of all retrieved implants. Additionally, resonance frequency analysis (RFA, Osstell™ mentor) will be used at different times for clinically monitoring the implant stability and for histological comparison in order to measure the reliability of the resonance frequency measuring device.</p> <p>Trial registration</p> <p>Current Controlled Trials ISRCTN97142521.</p

Retromer- and WASH-dependent sorting of nutrient transporters requires a multivalent interaction network with ANKRD50

Retromer and the associated actin-polymerizing WASH complex are essential for the endocytic recycling of a wide range of integral membrane proteins. A hereditary Parkinson's-disease-causing point mutation (D620N) in the retromer subunit VPS35 perturbs retromer's association with the WASH complex and also with the uncharacterized protein ankyrin-repeat-domain-containing protein 50 (ANKRD50). Here, we firmly establish ANKRD50 as a new and essential component of the SNX27– retromer–WASH super complex. Depletion of ANKRD50 in HeLa or U2OS cells phenocopied the loss of endosome-to-cell-surface recycling of multiple transmembrane proteins seen upon suppression of SNX27, retromer or WASH- complex components. Mass-spectrometry-based quantification of the cell surface proteome of ANKRD50-depleted cells identified amino acid transporters of the SLC1A family, among them SLC1A4, as additional cargo molecules that depend on ANKRD50 and retromer for their endocytic recycling. Mechanistically, we show that ANKRD50 simultaneously engages multiple parts of the SNX27–retromer–WASH complex machinery in a direct and co-operative interaction network that is needed to efficiently recycle the nutrient transporters GLUT1 (also known as SLC2A1) and SLC1A4, and potentially many other surface proteins

Discrete cytosolic macromolecular BRAF complexes exhibit distinct activities and composition

As a central element within the RAS/ERK pathway, the serine/threonine kinase BRAF plays a key role in development and homeostasis and represents the most frequently mutated kinase in tumors. Consequently, it has emerged as an important therapeutic target in various malignancies. Nevertheless, the BRAF activation cycle still raises many mechanistic questions as illustrated by the paradoxical action and side effects of RAF inhibitors. By applying SEC‐PCP‐SILAC, we analyzed protein–protein interactions of hyperactive BRAFV600E and wild‐type BRAF (BRAFWT). We identified two macromolecular, cytosolic BRAF complexes of distinct molecular composition and phosphorylation status. Hyperactive BRAFV600E resides in large complexes of higher molecular mass and activity, while BRAFWT is confined to smaller, slightly less active complexes. However, expression of oncogenic K‐RasG12V, either by itself or in combination with RAF dimer promoting inhibitors, induces the incorporation of BRAFWT into large, active complexes, whereas pharmacological inhibition of BRAFV600E has the opposite effect. Thus, the quaternary structure of BRAF complexes is shaped by its activation status, the conformation of its kinase domain, and clinically relevant inhibitors

Retromer/WASH dependent sorting of nutrient transporters requires a multivalent interaction network with ANKRD50

Retromer and the associated actin-polymerizing WASH complex are essential for the endocytic recycling of a wide range of integral membrane proteins. A hereditary Parkinson's-disease-causing point mutation (D620N) in the retromer subunit VPS35 perturbs retromer's association with the WASH complex and also with the uncharacterized protein ankyrin-repeat-domain-containing protein 50 (ANKRD50). Here, we firmly establish ANKRD50 as a new and essential component of the SNX27–retromer–WASH super complex. Depletion of ANKRD50 in HeLa or U2OS cells phenocopied the loss of endosome-to-cell-surface recycling of multiple transmembrane proteins seen upon suppression of SNX27, retromer or WASH-complex components. Mass-spectrometry-based quantification of the cell surface proteome of ANKRD50-depleted cells identified amino acid transporters of the SLC1A family, among them SLC1A4, as additional cargo molecules that depend on ANKRD50 and retromer for their endocytic recycling. Mechanistically, we show that ANKRD50 simultaneously engages multiple parts of the SNX27–retromer–WASH complex machinery in a direct and co-operative interaction network that is needed to efficiently recycle the nutrient transporters GLUT1 (also known as SLC2A1) and SLC1A4, and potentially many other surface proteins

АНАЛИЗ МЕТОДОВ И СРЕДСТВ ОЦЕНКИ ТЕМПЕРАТУРЫ И ВЛАЖНОСТИ ЗЕРНА ПРИ ЕГО ДЛИТЕЛЬНОМ ХРАНЕНИИ

В работе описывается основные факторы, влияющие на качество зерна при его хранении. А также методы контроля их с помощью приборов

Phospho-proteomic analyses of B-Raf protein complexes reveal new regulatory principles

B-Raf represents a critical physiological regulator of the Ras/RAF/MEK/ERK-pathway and a pharmacological target of growing clinical relevance, in particular in oncology. To understand how B-Raf itself is regulated, we combined mass spectrometry with genetic approaches to map its interactome in MCF-10A cells as well as in B-Raf deficient murine embryonic fibroblasts (MEFs) and B-Raf/Raf-1 double deficient DT40 lymphoma cells complemented with wildtype or mutant B-Raf expression vectors. Using a multi-protease digestion approach, we identified a novel ubiquitination site and provide a detailed B-Raf phospho-map. Importantly, we identify two evolutionary conserved phosphorylation clusters around T401 and S419 in the B-Raf hinge region. SILAC labelling and genetic/biochemical follow-up revealed that these clusters are phosphorylated in the contexts of oncogenic Ras, sorafenib induced Raf dimerization and in the background of the V600E mutation. We further show that the vemurafenib sensitive phosphorylation of the T401 cluster occurs in trans within a Raf dimer. Substitution of the Ser/Thr-residues of this cluster by alanine residues enhances the transforming potential of B-Raf, indicating that these phosphorylation sites suppress its signaling output. Moreover, several B-Raf phosphorylation sites, including T401 and S419, are somatically mutated in tumors, further illustrating the importance of phosphorylation for the regulation of this kinase

Multi-objective Problems in Terms of Relational Algebra

Abstract. Relational algebra has been shown to be a powerful tool for solving a wide range of combinatorial optimization problems with small computational and programming effort. The problems considered in recent years are single- objective ones where one single objective function has to be optimized. With this paper we start considerations on the use of relational algebra for multi-objective problems. In contrast to singleobjective optimization multiple objective functions have to be optimized at the same time usually resulting in a set of different trade-offs with respect to the different functions. On the one hand, we examine how to solve the mentioned problem exactly by using relational algebraic programs. On the other hand, we address the problem of objective reduction that has recently been shown to be NP-hard. We propose an exact algorithm for this problem based on relational algebra. Our experimental results show that this algorithm drastically outperforms the currently best one.

Space for Opinions

Wrede B, Wolter D, Kirsch A, Bhatt M, Kersting K, Meyer U. Space for Opinions. KUENSTLICHE INTELLIGENZ. 2017;31(2):117-119