Molecular basis for the loss-of-function effects of the Alzheimer's disease-associated R47H variant of the immune receptor TREM2.

Triggering receptor expressed on myeloid cells 2 (TREM2) is an immune receptor expressed on the surface of microglia, macrophages, dendritic cells, and osteoclasts. The R47H TREM2 variant is a significant risk factor for late-onset Alzheimer's disease (AD), and the molecular basis of R47H TREM2 loss of function is an emerging area of TREM2 biology. Here, we report three high-resolution structures of the extracellular ligand-binding domains (ECDs) of R47H TREM2, apo-WT, and phosphatidylserine (PS)-bound WT TREM2 at 1.8, 2.2, and 2.2 Å, respectively. The structures reveal that Arg47 plays a critical role in maintaining the structural features of the complementarity-determining region 2 (CDR2) loop and the putative positive ligand-interacting surface (PLIS), stabilizing conformations capable of ligand interaction. This is exemplified in the PS-bound structure, in which the CDR2 loop and PLIS drive critical interactions with PS via surfaces that are disrupted in the variant. Together with in vitro and in vivo characterization, our structural findings elucidate the molecular mechanism underlying loss of ligand binding, putative oligomerization, and functional activity of R47H TREM2. They also help unravel how decreased in vitro and in vivo stability of TREM2 contribute to loss of function in disease.

Early childhood caries epidemiology, aetiology, risk assessment, societal burden, management, education, and policy: Global perspective.

BACKGROUND: This paper is a summary of the proceedings of the International Association of Paediatric Dentistry Bangkok Conference on early childhood caries (ECC) held in 3-4 November 2018. AIM: The paper aims to convey a global perspective of ECC definitions, aetiology, risk factors, societal costs, management, educational curriculum, and policy. DESIGN: This global perspective on ECC is the compilation of the state of science, current concepts, and literature regarding ECC from worldwide experts on ECC. RESULTS: Early childhood caries is related to frequent sugar consumption in an environment of enamel adherent, acid-producing bacteria in a complex biofilm, as well as developmental defects of enamel. The seriousness, societal costs, and impact on quality of life of dental caries in pre-school children are enormous. Worldwide data show that ECC continues to be highly prevalent, yet infrequently treated. Approaches to reduce the prevalence include interventions that start in the first year of a child's life, evidence-based and risk-based management, and reimbursement systems that foster preventive care. CONCLUSIONS: This global perspective on ECC epidemiology, aetiology, risk assessment, global impact, and management is aimed to foster improved worldwide understanding and management of ECC.

Dental Students' and Dental School Graduates' Practical Skills: An International Survey of Perceptions of National Dental Associations in Europe.

PURPOSE: Dental students learn knowledge and practical skills to provide oral health care to the population. Practical skills must be maintained or continuously developed throughout a professional career. This cross-sectional survey aimed to evaluate the perception of practical skills of dental students and dental-school graduates by national dental associations (NDAs) in international comparison in the European Regional Organization of the FDI World Dental Federation (ERO-FDI) zone. MATERIALS AND METHODS: A questionnaire of 14 items collected information on pre-/postgraduate areas. RESULTS: A total of 25 countries participated (response rate: 69.4%), with 80.0% having minimum requirements for practical skills acquisition and 64.0% starting practical training in the 3rd year of study. In countries where clinical practical work on patients begins in the 2nd year of study, practical skills of graduates are perceived as average, starting in the 3rd year of study as mainly good, starting in the 4th as varying widely from poor to very good. In total, 76.0% of respondents feel that improvements are needed before entering dental practice. Improvements could be reached by treating more patients in dental school (32.0%), increasing the quantity of clinical training (20.0%), or having more clinical instructors (12.0%). In 56.0% of the countries, it is possible to open one's own dental practice immediately after graduation, and in 16.0%, prior vocational training is mandatory. CONCLUSIONS: All participating countries in the ERO-FDI zone reported practical training in dental school, most starting in the 3rd year of study. The perception of practical skills of dental students and dental-school graduates among NDAs is very heterogeneous. Reasons for the perceived deficiencies should be further explored.

The Fibronectin-ILT3 Interaction Functions as a Stromal Checkpoint that Suppresses Myeloid Cells.

Suppressive myeloid cells inhibit antitumor immunity by preventing T-cell responses. Immunoglobulin-like transcript 3 (ILT3; also known as LILRB4) is highly expressed on tumor-associated myeloid cells and promotes their suppressive phenotype. However, the ligand that engages ILT3 within the tumor microenvironment and renders tumor-associated myeloid cells suppressive is unknown. Using a screening approach, we identified fibronectin as a functional ligand for ILT3. The interaction of fibronectin with ILT3 polarized myeloid cells toward a suppressive state, and these effects were reversed with an ILT3-specific antibody that blocked the interaction of ILT3 with fibronectin. Furthermore, ex vivo treatment of human tumor explants with anti-ILT3 reprogrammed tumor-associated myeloid cells toward a stimulatory phenotype. Thus, the ILT3-fibronectin interaction represents a "stromal checkpoint" through which the extracellular matrix actively suppresses myeloid cells. By blocking this interaction, tumor-associated myeloid cells may acquire a stimulatory phenotype, potentially resulting in increased antitumor T-cell responses.

The DNA replication-related element-binding factor (DREF) is a transcriptional regulator of the Drosophila myb gene.

Drosophila melanogaster possesses a single gene, Dm myb, that is closely related to the vertebrate proto-oncogene c-Myb, and its other family members (A-Myb and B-Myb), all of which encode transcription factors. Dm myb is expressed in all proliferating cells throughout development, and previous studies demonstrate that Dm myb promotes both S-phase and M-phase in proliferating cells, while preserving diploidy by suppressing endoreduplication. We have initiated a characterization of the mechanisms that regulate Dm myb expression, and we report here that the transcriptional activator DREF (the DNA replication-related element binding factor) activates Dm myb transcription via two binding sites located in the 5' flanking region; that the Dm myb promoter lacks a prototypical TATA box sequence and instead appears to use an initiator/downstream promoter element (Inr/DPE) type promoter; and that Dm myb expression is regulated at the translational as well as transcriptional level.

Discovery of substrate for type I signal peptidase SpsB from Staphylococcus aureus.

Based on the kinetic model of substrate phage proteolysis, we have formulated a strategy for best manipulating the conditions in screening phage display libraries for protease substrates (Sharkov, N. A., Davis, R. M., Reidhaar-Olson, J. F., Navre, M., and Cai, D. (2001) J. Biol. Chem. 276, 10788-10793). This strategy is exploited in the present study with signal peptidase SpsB from Staphylococcus aureus. We demonstrate that highly active substrate phage clones can be isolated from a phage display library by systematically tuning the selection stringency in screening. Several of the selected clones exhibit superior reactivity over a control, the best clone, SIIIRIII-8, showing >100-fold improvement. Because no conserved sequence features were readily revealed that could allow delineation of the active and unreactive clones, the sequences identified in five of the active clones were tested as synthetic dodecamers, Ac-AGX(8)GA-NH(2). Using electrospray ionization mass spectrometry, we show that four of these peptides can be cleaved by SpsB and that Ala is the P1 residue exclusively and Ala or Leu the P3 residue, in keeping with the (-3, -1) rule for substrate recognition by signal peptidase. Our successful screening with SpsB demonstrated the general applicability of the screening strategy and allowed us to isolate the first peptide substrates for the enzyme.

Drosophila myb exerts opposing effects on S phase, promoting proliferation and suppressing endoreduplication.

Drosophila melanogaster possesses a single gene, Dm myb, that is closely related to the vertebrate family of Myb genes, which encode transcription factors that are involved in regulatory decisions affecting cell proliferation, differentiation and apoptosis. The vertebrate Myb genes have been specifically implicated in regulating the G(1)/S transition of the cell cycle. Dm myb is expressed in all proliferating tissues, but not at detectable levels in endoreduplicating cells. Analysis of loss-of-function mutations in Dm myb revealed a block at the G(2)/M transition and mitotic defects, but did not directly implicate Dm myb function in the G(1/)S transition. We have used the Gal4-UAS binary system of ectopic expression to further investigate the function of Dm myb. Our results demonstrate that depending upon the type of cell cycle, ectopic Dm myb activity can exert opposing effects on S phase: driving DNA replication and promoting proliferation in diploid cells, even when developmental signals normally dictate cell cycle arrest; but suppressing endoreduplication in endocycling cells, an effect that can be overcome by induction of E2F. We also show that a C-terminally truncated DMyb protein, which is similar to an oncogenic form of vertebrate Myb, has more potent effects than the full-length protein, especially in endoreduplicating tissues. This finding indicates that the C terminus acts as a negative regulatory domain, which can be differentially regulated in a tissue-specific manner. Our studies help to resolve previous discrepancies regarding myb gene function in Drosophila and vertebrates. We conclude that in proliferating cells, Dm myb has the dual function of promoting S phase and M phase, while preserving diploidy by suppressing endoreduplication.