Molecular Determinants and Consequences of Specificity in Histone 2A Ubiquitination

Specific ubiquitination of histones H2A is a crucial decision making point in the response to DNA damage. This thesis analysis the role of three distinct groups of lysines on H2A that are specifically ubiquitinated by three different E3 ligases, RING1b/BMI1, BRCA1/BARD1 and RNF168. The mechanistic basics underlying this specificity are discussed. The work describes how specific ubiquitination is employed to guide repair pathway choice between homologous recombination and non-homologous end joining. It shows that USP48, a deubiquitinating enzyme specific for the BRCA1 ubiquitination site, guides repair pathway choice by determining the extent of DNA end resection. Analysis of the E3 ligase RNF168 shows how specific interaction of the E3 with the nucleosomal acidic patch defines site-specificity. Furthermore, a general framework for structural analysis of E3-E2-Substrate complexes is presented

USP48 restrains resection by site-specific cleavage of the BRCA1 ubiquitin mark from H2A

BRCA1 ligase activity is tightly regulated to maintain genome stability and confer DNA double strand repair. Here the authors identify USP48 as a H2A deubiquitinating enzyme that acts as a BRCA1 E3 ligase antagonist and characterize its role during DNA repair

A complex proinflammatory cascade mediates the activation of HSCs upon LPS exposure in vivo

Infections are a key source of stress to the hematopoietic system. While infections consume short-lived innate immune cells, their recovery depends on quiescent hematopoietic stem cells (HSCs) with long-term self-renewal capacity. Both chronic inflammatory stress and bacterial infections compromise competitive HSC capacity and cause bone marrow failure. However, our understanding of how HSCs act during acute and contained infections remains incomplete. Here, we used advanced chimeric and genetic mouse models in combination with pharmacological interventions to dissect the complex nature of the acute systemic response of HSCs to lipopolysaccharide (LPS), a well-established model for inducing inflammatory stress. Acute LPS challenge transiently induced proliferation of quiescent HSCs in vivo. This response was not only mediated via direct LPS-TLR4 conjugation on HSCs, but also involved indirect TLR4 signaling in CD115+ monocytic cells, inducing a complex pro-inflammatory cytokine cascade in the bone marrow. Downstream of LPS-TLR4 signaling, the combined action of pro-inflammatory cytokines such as IFNα, IFNγ, TNFα, IL-1α, IL-1β and many others is required to mediate full HSC activation in vivo. Together, our study reveals detailed mechanistic insights into the interplay of proinflammatory cytokine-induced molecular pathways and cell types that jointly orchestrate the complex process of emergency hematopoiesis and HSC activation upon LPS exposure in vivo

Strategy for development of site-specific ubiquitin antibodies

Protein ubiquitination is a key post-translational modification regulating a wide range of biological processes. Ubiquitination involves the covalent attachment of the small protein ubiquitin to a lysine of a protein substrate. In addition to its well-established role in protein degradation, protein ubiquitination plays a role in protein-protein interactions, DNA repair, transcriptional regulation, and other cellular functions. Understanding the mechanisms and functional relevance of ubiquitin as a signaling system requires the generation of antibodies or alternative reagents that specifically detect ubiquitin in a site-specific manner. However, in contrast to other post-translational modifications such as acetylation, phosphorylation, and methylation, the instability and size of ubiquitin-76 amino acids-complicate the preparation of suitable antigens and the generation antibodies detecting such site-specific modifications. As a result, the field of ubiquitin research has limited access to specific antibodies. This severely hampers progress in understanding the regulation and function of site-specific ubiquitination in many areas of biology, specifically in epigenetics and cancer. Therefore, there is a high demand for antibodies recognizing site-specific ubiquitin modifications. Here we describe a strategy for the development of site-specific ubiquitin antibodies. Based on a recently developed antibody against site-specific ubiquitination of histone H2B, we provide detailed protocols for chemical synthesis methods for antigen preparation and discuss considerations for screening and quality control experiments.Chemical Immunolog

Antigen presentation safeguards the integrity of the hematopoietic stem cell pool

Hematopoietic stem and progenitor cells (HSPCs) are responsible for the production of blood and immune cells. Throughout life, HSPCs acquire oncogenic aberrations that can cause hematological cancers. Although molecular programs maintaining stem cell integrity have been identified, safety mechanisms eliminating malignant HSPCs from the stem cell pool remain poorly characterized. Here, we show that HSPCs constitutively present antigens via major histocompatibility complex class II. The presentation of immunogenic antigens, as occurring during malignant transformation, triggers bidirectional interactions between HSPCs and antigen-specific CD4(+4) T cells, causing stem cell proliferation, differentiation, and specific exhaustion of aberrant HSPCs. This immunosurveillance mechanism effectively eliminates transformed HSPCs from the hematopoietic system, thereby preventing leukemia onset. Together, our data reveal a bidirectional interaction between HSPCs and CD4(+4) T cells, demonstrating that HSPCs are not only passive receivers of immunological signals but also actively engage in adaptive immune responses to safeguard the integrity of the stem cell pool

A survey of literature on automated storage and retrieval systems from 2009 to 2019

This paper provides a literature review on automated storage and retrieval systems based on the Scopus database. The last comprehensive literature review that analyses the subject in its entirety dates back to 2009 by Roodbergen and Vis. Since then, the scientific community has provided many new efficient solutions, most of which are collected in Scopus. All these new published solutions need to be classified and summarised; thus, for this paper, more than one thousand scientific publications were analysed in order to classify the proposed solutions, deliver new academic insights to the field, and provide clear guidelines for researchers and practitioner

Integrated billing mechanisms in the Internet of Things to support information sharing and enable new business opportunities

The Internet of Things (IOT) concept and enabling technologies such as RFID offer the prospect of linking the real world of physical objects with the virtual world of information technology to improve visibility and traceability information within supply chains and across the entire lifecycles of products, as well as enabling more intuitive interactions and greater automation possibilities. There is a huge potential for savings through process optimization and profit generation within the IOT, but the sharing of financial benefits across companies remains an unsolved issue. Existing approaches towards sharing of costs and benefits have failed to scale so far. The integration of payment solutions into the IOT architecture could solve this problem. We have reviewed different possible levels of integration. Multiple payment solutions have been researched. Finally we have developed a model that meets the requirements of the IOT in relation to openness and scalability. It supports both hardware-centric and software-centric approaches to integration of payment solutions with the IOT. Different requirements concerning payment solutions within the IOT have been defined and considered in the proposed model. Possible solution providers include telcos, e-payment service providers and new players such as banks and standardization bodies. The proposed model of integrating the Internet of Things with payment solutions will lower the barrier to invoicing for the more granular visibility information generated using the IOT. Thus, it has the potential to enable recovery of the necessary investments in IOT infrastructure and accelerate adoption of the IOT, especially for projects that are only viable when multiple benefits throughout the supply chain need to be accumulated in order to achieve a Return on Investment (ROI). In a long-term perspective, it may enable IT-departments to become profit centres instead of cost centres. © 2010 - IOS Press and the authors. All rights reserved