How does the kinase Lck phosphorylate the T cell receptor? Spatial organization as a regulatory mechanism

T cell signaling begins with the ligation of the T cell antigen receptor (TCR) by a cognate peptide and the phosphorylation of the receptor’s immunoreceptor tyrosine-based activation motif domains by the kinase Lck. However, the canonical receptor model is insufficient to explain how the constitutively active kinase Lck can discriminate between non-ligated and ligated TCRs. Here, we discuss the factors that are thought to regulate the spatial distribution of the TCR and Lck, and therefore critically influence TCR signaling initiation

Valproic acid use in fertile women with genetic generalized epilepsies

Objectives: In genetic generalized epilepsies (GGE), valproic acid (VPA) is the most efficacious compound. However, due to teratogenicity and increased risk for impaired cognitive development after intrauterine exposure, its use in women of fertile age is strictly regulated but sometimes unavoidable. Methods: All patients with GGE treated at the outpatient clinic of a tertiary epilepsy center with at least one visit between January 2015 and April 2020 were included in this retrospective study. The rate of women aged 18 to 49 years taking VPA was compared to that of men of the same age group and to women > 49 years. Furthermore, in each group, clinical variables associated with VPA use were sought. Results: Twenty-eight out of 125 women of fertile age (22%) were treated with VPA, compared to 28 out of 56 men ≤ 49 years (50%; p = .002) and to 22 out of 40 female patients > 49 years (55%; p < .001). VPA dose was lower in fertile women compared to men, with no difference in seizure freedom rates. In women ≤ 49 years, multivariate analysis demonstrated age as the only variable independently associated with VPA use (OR 1.095; 95% CI 1.036-1.159). In the other two groups, no associated variables were identified. Conclusions: Despite warnings with respect to teratogenicity and impaired cognitive development with VPA, from 2015 to 2020, almost every fourth women of fertile age with GGE received this compound. Inevitably lower VPA doses in these women seem sufficient for favorable seizure freedom rates

Continuing Professional Development at Two Rural Hospitals in Ecuador

Background: E-learning Continuing Professional Development (CPD) is an activity demonstrated to improve the quality of healthcare delivery. The CPD of medical and nursing staff in high income countries (HICs) is commonplace. CPD of administrative staff is less common, but increasingly frequent. In low- and middle-income countries (LMICs), CPD of any kind is infrequent, particularly in rural and remote areas. Objective: The aim of this study was to describe a hospital-based e-learning CPD program for clinical and non-clinical personnel as a unique example of a successful, ongoing educational pilot, quality improvement program involving a broad cohort of employees, in a country that does not require such activities. Methods: Using the online educational platform Chamilo, e-learning modules were created for eight groups including clinical and non-clinical employees. Upon completion of each module, one to two paragraph discussions were provided for each incorrect answer submitted. Two additional chances were offered for the employee to achieve a passing score of 70%. This study reports on the first 10-month period of the program. Findings: All participants achieved the 70% passing threshold after the first or second attempt. There was 100% participation by the employees required to complete the e-learning modules. Employee feedback suggested the modules were good for continuing education, but some felt the CPD was imposed on them. Conclusion: E-learning CPD is an important and emerging element for CPD and may provide opportunities for healthcare service quality improvement as part of broader pedagogical modalities, such as conferences and directed readings, in rural and remote areas of LMICs. These pilot programs could provide important information to develop Spanish-language e-learning CPD programs across a broader region, promote collaboration with regional professional societies, and possibly contribute to the establishment of national health program CPD standards

Angular-Differential Cross Sections for H(2p) Formation in Intermediate-Energy Proton-Helium Collisions

Angular-differential cross sections for charge transfer with simultaneous emission of a photon in collisions of protons with helium atoms have been measured. The incident proton energies were 25, 50, and 100 keV and the center-of-mass scattering angles were between 0 and 2.0 mrad. In the experiment, hydrogen atoms that scattered through an angle θ were detected in coincidence with photons emitted perpendicular to the scattering plane with a wavelength between 1140 and 1400 Å. Differential cross sections for capture into the 2p state of the hydrogen atom were determined from the variation in the coincidence signal with θ. The experimental results are compared with the results of a classical trajectory Monte Carlo (CTMC) simulation and with the results of a calculation for H(2p) capture using the Coulomb-Brinkman-Kramers (CBK) approximation. The agreement between the experimental results and the CTMC calculation is good at all three energies while the agreement between the shape of the data and the CBK calculation is good at 50 and 100 keV

The Reorientation of T-Cell Polarity and Inhibition of Immunological Synapse Formation by CD46 Involves Its Recruitment to Lipid Rafts

Many infectious agents utilize CD46 for infection of human cells, and therapeutic applications of CD46-binding viruses are now being explored. Besides mediating internalization to enable infection, binding to CD46 can directly alter immune function. In particular, ligation of CD46 by antibodies or by measles virus can prevent activation of T cells by altering T-cell polarity and consequently preventing the formation of an immunological synapse. Here, we define a mechanism by which CD46 reorients T-cell polarity to prevent T-cell receptor signaling in response to antigen presentation. We show that CD46 associates with lipid rafts upon ligation, and that this reduces recruitment of both lipid rafts and the microtubule organizing centre to the site of receptor cross-linking. These data combined indicate that polarization of T cells towards the site of CD46 ligation prevents formation of an immunological synapse, and this is associated with the ability of CD46 to recruit lipid rafts away from the site of TCR ligation

Functional role of T-cell receptor nanoclusters in signal initiation and antigen discrimination

Antigen recognition by the T-cell receptor (TCR) is a hallmark of the adaptive immune system. When the TCR engages a peptide bound to the restricting major histocompatibility complex molecule (pMHC), it transmits a signal via the associated CD3 complex. How the extracellular antigen recognition event leads to intracellular phosphorylation remains unclear. Here, we used single-molecule localization microscopy to quantify the organization of TCR–CD3 complexes into nanoscale clusters and to distinguish between triggered and nontriggered TCR–CD3 complexes. We found that only TCR–CD3 complexes in dense clusters were phosphorylated and associated with downstream signaling proteins, demonstrating that the molecular density within clusters dictates signal initiation. Moreover, both pMHC dose and TCR–pMHC affinity determined the density of TCR–CD3 clusters, which scaled with overall phosphorylation levels. Thus, TCR–CD3 clustering translates antigen recognition by the TCR into signal initiation by the CD3 complex, and the formation of dense signaling-competent clusters is a process of antigen discrimination

Dephosphorylation accelerates the dissociation of ZAP70 from the T cell receptor

Protein–protein binding domains are critical in signaling networks. Src homology 2 (SH2) domains are binding domains that interact with sequences containing phosphorylated tyrosines. A subset of SH2 domain–containing proteins has tandem domains, which are thought to enhance binding affinity and specificity. However, a trade-off exists between long-lived binding and the ability to rapidly reverse signaling, which is a critical requirement of noise-filtering mechanisms such as kinetic proofreading. Here, we use modeling to show that the unbinding rate of tandem, but not single, SH2 domains can be accelerated by phosphatases. Using surface plasmon resonance, we show that the phosphatase CD45 can accelerate the unbinding rate of zeta chain–associated protein kinase 70 (ZAP70), a tandem SH2 domain–containing kinase, from biphosphorylated peptides from the T cell receptor (TCR). An important functional prediction of accelerated unbinding is that the intracellular ZAP70–TCR half-life in T cells will not be fixed but rather, dependent on the extracellular TCR–antigen half-life, and we show that this is the case in both cell lines and primary T cells. The work highlights that tandem SH2 domains can break the trade-off between signal fidelity (requiring long half-life) and signal reversibility (requiring short half-life), which is a key requirement for T cell antigen discrimination mediated by kinetic proofreading.https://www.biorxiv.org/content/10.1101/2020.02.12.945170v