Molecular architecture of the αβ T cell receptor–CD3 complex

αβ T-cell receptor (TCR) activation plays a crucial role for T-cell function. However, the TCR itself does not possess signaling domains. Instead, the TCR is noncovalently coupled to a conserved multisubunit signaling apparatus, the CD3 complex, that comprises the CD3εγ, CD3εδ, and CD3ζζ dimers. How antigen ligation by the TCR triggers CD3 activation and what structural role the CD3 extracellular domains (ECDs) play in the assembled TCR–CD3 complex remain unclear. Here, we use two complementary structural approaches to gain insight into the overall organization of the TCR–CD3 complex. Small-angle X-ray scattering of the soluble TCR–CD3εδ complex reveals the CD3εδ ECDs to sit underneath the TCR α-chain. The observed arrangement is consistent with EM images of the entire TCR–CD3 integral membrane complex, in which the CD3εδ and CD3εγ subunits were situated underneath the TCR α-chain and TCR β-chain, respectively. Interestingly, the TCR–CD3 transmembrane complex bound to peptide–MHC is a dimer in which two TCRs project outward from a central core composed of the CD3 ECDs and the TCR and CD3 transmembrane domains. This arrangement suggests a potential ligand-dependent dimerization mechanism for TCR signaling. Collectively, our data advance our understanding of the molecular organization of the TCR–CD3 complex, and provides a conceptual framework for the TCR activation mechanism.T cells are key mediators of the adaptive immune response. Each αβ T cell contains a unique αβ T-cell receptor (TCR), which binds antigens (Ags) displayed by major histocompatibility complexes (MHCs) and MHC-like molecules (1). The TCR serves as a remarkably sensitive driver of cellular function: although TCR ligands typically bind quite weakly (1–200 μM), even a handful of TCR ligands are sufficient to fully activate a T cell (2, 3). The TCR does not possess intracellular signaling domains, uncoupling Ag recognition from T-cell signaling. The TCR is instead noncovalently associated with a multisubunit signaling apparatus, consisting of the CD3εγ and CD3εδ heterodimers and the CD3ζζ homodimer, which collectively form the TCR–CD3 complex (4, 5). The CD3γ/δ/ε subunits each consist of a single extracellular Ig domain and a single immunoreceptor tyrosine-based activation motif (ITAM), whereas CD3ζ has a short extracellular domain (ECD) and three ITAMs (6–11). The TCR–CD3 complex exists in 1:1:1:1 stoichiometry for the αβTCR:CD3εγ:CD3εδ:CD3ζζ dimers (12). Phosphorylation of the intracellular CD3 ITAMs and recruitment of the adaptor Nck lead to T-cell activation, proliferation, and survival (13, 14). Understanding the underlying principles of TCR–CD3 architecture and T-cell signaling is of therapeutic interest. For example, TCR–CD3 is the target of therapeutic antibodies such as the immunosuppressant OKT3 (15), and there is increasing interest in manipulating T cells in an Ag-dependent manner by using naturally occurring and engineered TCRs (16).Assembly of the TCR–CD3 complex is primarily driven by each protein’s transmembrane (TM) region, enforced through the interaction of evolutionarily conserved, charged, residues in each TM region (4, 5, 12). What, if any, role interactions between TCR and CD3 ECDs play in the assembly and function of the complex remains controversial (5): there are several plausible proposed models of activation, which are not necessarily mutually exclusive (5, 17–19). Although structures of TCR–peptide–MHC (pMHC) complexes (2), TCR–MHC-I–like complexes (1), and the CD3 dimers (6–10) have been separately determined, how the αβ TCR associates with the CD3 complex is largely unknown. Here, we use two independent structural approaches to gain an understanding of the TCR–CD3 complex organization and structure.     

Pharmacoepidemiology of Antipsychotic Use in Youth with ADHD: Trends and Clinical Implications

Although concern has been raised about antipsychotic prescribing to youth with attention-deficit/hyperactivity disorder (ADHD), the available database is limited to individual studies. Therefore, in order to provide a synthesis of prevalence and time trends, we conducted a systematic review and pooled analysis of pharmaco-epidemiologic data on antipsychotic use in ADHD youth. Of 1806 hits, 21 studies (N) were retained that reported analyzable data for three separate populations: 1) antipsychotic-treated youth (N?=?15, n?=?341,586); 2) ADHD youth (N?=?9, n?=?6,192,368), and 3) general population youth (N?=?5, n?=?14,284,916). Altogether, 30.5?±?18.5 % of antipsychotic-treated youth had ADHD. In longitudinal studies, this percentage increased over time (1998–2007) from 21.7?±?7.1 % to 27.7?±?7.7 %, ratio?=?1.3?±?0.4. Furthermore, 11.5?±?17.5 % of ADHD youth received antipsychotics. In longitudinal studies, this percentage also increased (1998–2006) from 5.5?±?2.6 % to 11.4?±?6.7 %, ratio?=?2.1?±?0.6. Finally, 0.12?±?0.07 % of youth in the general population were diagnosed with ADHD and received antipsychotics. Again, in longitudinal studies, this percentage increased over time (1993–2007): 0.13?±?0.09 % to 0.44?±?0.49 %, ratio?=?3.1?±?2.2. Taken together, these data indicate that antipsychotics are used by a clinically relevant and increasing number of youth with ADHD. Reasons for and risk/benefit ratios of this practice with little evidence base require further investigation.     

Tegillarca granosa extract Haishengsu (HSS) suppresses expression of mdr1, BCR/ABL and sorcin in drug-resistant K562/ADM tumors in mice

PurposeTo evaluate the effect of Haishengsu (HSS), a protein extract from Tegillarca granosa, on multidrug-resistance genes mdr1, BCR/ABL and sorcin in transplanted tumors.Material/MethodsMice were inoculated subcutaneously with a drug resistant leukemia cell line K562/ADM. Tumor-bearing animals were divided into control, adriamycin, HSS and combination therapy (adriamycin plus HSS) groups. Flow cytometry was used to detect apoptosis of tumor cells, and RT-PCR was used to evaluate the expression of mdr1, BCR/ABL and sorcin.ResultsThe apoptosis rate in the high (71.8%), medium (72.3%) and low doses HSS group (72.4%) was higher than in control (1.2%, p<0.01), adriamycin (34.4%, p<0.05) or combination therapy group (46.4%, p<0.05). The mean optical density of mdr1, BCR/ABL and sorcin in HSS groups was lower than in control, adriamycin and combination therapy group (p<0.01). The optical density of the three genes in high HSS group was lower than in medium and low HSS group (p<0.01).ConclusionsHaishengsu promotes apoptosis of drug-resistant K562/ADM tumors in mice in a dose-dependent manner. The pro-apoptotic effect of Haishengsu may be related to a reduced expression of multidrug-resistance genes mdr1, BCR/ABL and sorcin.     

Thermometry during coblation and radiofrequency ablation of vertebral metastases: a cadaver study

Purpose

To evaluate safety of coblation of simulated lytic metastases in human cadaveric vertebral bodies by measuring heat distribution during thermal tissue ablation and comparing it to radiofrequency ablation (RFA).

Materials and methods

Three devices were compared: a 10 mm single-needle RFA electrode, a 20 mm array RFA electrode and the coblation device. To simulate bone metastases, a spinal tumor model was used stuffing a created lytic cavity with muscle tissue. Measuring of heat distribution was performed during thermal therapy within the vertebral body, in the epidural space and at the ipsilateral neural foramen. Eight vertebral bodies were used for each device.

Results

Temperatures at heat-sensitive neural structures during coblation were significantly lower than using RFA. Maximum temperatures measured at the end of the procedure at the neural foramen: 46.4 °C (±2.51; RFA 10 mm), 52.2 °C (±5.62; RFA 20 mm) and 42.5 °C (±2.88; coblation). Maximum temperatures in the epidural space: 46.8 °C (±4.7; RFA 10 mm), 49.5 °C (±6.48; RFA 20 mm) and 42.1 °C (±2.5; coblation). Maximum temperatures measured within the vertebral body: 50.6 °C (±10.48; RFA 10 mm), 61.9 °C (±15.39; RFA 20 mm) and 54.4 °C (±15.77; coblation).

Conclusion

In addition to RFA, the application of coblation is a safe method to ablate vertebral lesions with regards to heat distribution at heat-sensitive neural spots. The measured temperatures did not harbor danger of thermal damage to the spinal cord or the spinal nerves.     

Associations of motor abilities with biological,sociodemographic, and behavioural factors in children: results from the ToyBox study

Sport Sciences for Health - In this cross-sectional study, we examined the association of selected basic motor abilities with biological (sex, age, and BMI), sociodemographic [socio-economic status...     

Survey on practice of venom immunotherapy in France

Introduction

Venom immunotherapy (VIT) is the only efficient prevention for sting-induced anaphylaxis, but its application is not without risks and needs precautions and standardization. European guidelines were proposed in 2005, but recent practice surveys and more recent knowledge raise the need for an update. The aim of this study was to analyze VIT practices in France, based on previous surveys in Europe but also extended to outcome event management.

Material and methods

A paper questionnaire was sent widely to persons involved in venom treatment.

Results

Eighty-six responses could be included from physicians actively involved in VIT induction evenly distributed in France. The survey shows that VIT was engaged from grade III down to grade I reactions, starting preferentially with the ultra-rush protocol. Premedication was used by 42% only and risks induced by co-treatment with β-blockers were well known but not with angiotensin-converting enzyme inhibitors. However, side effects were very variably managed from arrest to enhancement in doses, time-delay or duration. Similarly, we observed a large discrepancy in treatment evaluation (skin tests, biology, timing and interpretation), decision making for treatment termination (when and how long to be prolonged) and post-treatment follow-up (adrenaline kit, event record) as well as procedures in case of late relapse (new induction, different doses).

Conclusions

Our study shows that most recommendations were fully or partially followed and may need reminding, but many points need to be completed or updated with new tools and knowledge acquired during the last 10 years.     

Small genome of the fungus Escovopsis weberi,a specialized disease agent of ant agriculture

Many microorganisms with specialized lifestyles have reduced genomes. This is best understood in beneficial bacterial symbioses, where partner fidelity facilitates loss of genes necessary for living independently. Specialized microbial pathogens may also exhibit gene loss relative to generalists. Here, we demonstrate that Escovopsis weberi, a fungal parasite of the crops of fungus-growing ants, has a reduced genome in terms of both size and gene content relative to closely related but less specialized fungi. Although primary metabolism genes have been retained, the E. weberi genome is depleted in carbohydrate active enzymes, which is consistent with reliance on a host with these functions. E. weberi has also lost genes considered necessary for sexual reproduction. Contrasting these losses, the genome encodes unique secondary metabolite biosynthesis clusters, some of which include genes that exhibit up-regulated expression during host attack. Thus, the specialized nature of the interaction between Escovopsis and ant agriculture is reflected in the parasite’s genome.The highly evolved agricultural lifestyle of leaf-cutting ants has attracted particular attention because these ants cultivate a symbiotic fungus that serves as their major food source. These ants cut leaves, preprocess them into small pieces, and feed them to the cultivated fungus (1). The capacity of the cultivated fungus to break down plant material gives ant agriculturalists access to the vast nutrient stores locked within neotropical plants (Fig. 1A) (2–5). The symbiosis between fungus-growing ants and their cultivated fungi has persisted for at least 50 million years (6).Open in a separate windowFig. 1.Escovopsis weberi, a specialized mycoparasite of the fungus-growing ant symbiosis, has a small genome compared with other Pezizomycotina fungi. (A) Both fungus-growing ants and the mycoparasite E. weberi use the ants’ cultivated fungi as their primary food source. The ability of the cultivated fungi to efficiently break down plant material gives both consumers access to the biomass of neotropical plants. (B) Size and protein-coding gene content of genomes of diverse fungi in the Pezizomycotina. Bayesian phylogeny estimated using partial amino acid alignments of three genes (Rpb1, Rpb2, ef1-α). All posterior probabilities are greater than 0.95. Phylogeny is rooted with Sacchormyces cervesiae (not shown). (C) Relationship between genome size and gene content. A list of genomes included in this panel is in SI Appendix, Table S1.Like human agriculture, ant agriculture is hampered by disease. The ants’ fungal crops are attacked and consumed by fungal parasites of the genus Escovopsis (Ascomycota, Pezizomycotina: anamorphic Hypocreales) (Fig. 1A) (7), which have evolved in association with the ants and their cultivated fungi (8). Escovopsis infection can have detrimental impacts on garden health and, consequently, on the survival of ant colonies (9, 10). Such mycoparasitism, the phenomenon whereby one fungus is parasitic on another fungus, is rare. It is most well-known for species from the genus Trichoderma, some of which are used as biocontrol agents for fungal diseases and others of which attack human-cultivated fungi (11–13). In contrast to Trichoderma species, however, Escovopsis species grow poorly in their hosts’ absence (SI Appendix, Figs. S1 and S2).Escovopsis species have never been isolated outside of fungus-growing ant colonies, and different strains of Escovopsis are capable of attacking the fungi grown by different fungus-growing ant species (8, 14, 15). The long-term, specialized evolutionary history of the association between Escovopsis and their hosts provides a unique venue to explore the consequences of host specialization on pathogen genome evolution. Here, we assemble and annotate the genome of a strain of Escovopsis weberi. Consistent with expectations under an evolutionary transition toward using a narrow host range, and similar to many other specialized, host-associated microbes (16, 17), E. weberi exhibits gene loss. Contrasting other fungal pathogens, the large genomes of which are expanded with genetic elements that influence host adaptation (18), the genome size of Escovopsis is small compared with those of its closest sequenced relatives.     

Neoadjuvant Therapy for Rectal Cancer: Histologic Response of the Primary Tumor Predicts Nodal Status

PURPOSE: This study was designed to compare histologic T and N stages in patients with rectal adenocarcinoma undergoing various neoadjuvant radiotherapy regimens and proctectomy, in an attempt to determine if final histologic stage of the mural tumor predicts nodal status.METHODS: Data were collected from computerized databases at two institutions on 649 consecutive patients who underwent neoadjuvant radiotherapy or chemoradiotherapy and proctectomy for primary adenocarcinoma of the rectum from 1990 to 2002.RESULTS: Five patients were excluded because of incomplete pathology data sets, leaving a study population of 644. Patients underwent neoadjuvant radiotherapy alone (2,000 cGy in 5 fractions, n = 191; or 4,500 cGy in 25 fractions, n = 259) or chemoradiation (4,500 cGy in 25 fractions with concurrent 5-fluorouracil, n = 194). Histologic stage of the remaining mural tumor (ypT) correlated with nodal status (ypN). Lymph nodes harboring metastatic tumor were found in 1 of 42 (2 percent) ypT0 patients, 2 of 45 (4 percent) ypT1 patients, 43 of 186 (23 percent) ypT2 patients, 158 of 338 (47 percent) ypT3 patients, and 16 of 33 (48 percent) ypT4 patients (P < 0.001, chi-squared test). The probability of finding ypN+ disease was 3 of 87 (3 percent) in patients with ypT0-1 residual primary tumors vs. 220 of 557 (39 percent) in patients with ypT2-4 residual primary tumors (P < 0.0001; Fishers exact test).CONCLUSIONS: Nodal metastases are rare in patients whose mural tumor burden shrinks to ypT0-1 after neoadjuvant radiotherapy. If transanal excision is offered to select patients with distal rectal cancer, it is reasonable to select those who have an excellent clinical response to neoadjuvant therapy for transanal excision, and then reserve proctectomy for patients proven to have residual ypT2-4 disease.Read at the meeting of The American Society of Colon and Rectal Surgeons, New Orleans, Louisiana, June 21 to 26, 2003.