The pathogenic T42A mutation in SHP2 rewires the interaction specificity of its N-terminal regulatory domain.

Mutations in the tyrosine phosphatase Src homology-2 domain-containing protein tyrosine phosphatase-2 (SHP2) are associated with a variety of human diseases. Most mutations in SHP2 increase its basal catalytic activity by disrupting autoinhibitory interactions between its phosphatase domain and N-terminal SH2 (phosphotyrosine recognition) domain. By contrast, some disease-associated mutations located in the ligand-binding pockets of the N- or C-terminal SH2 domains do not increase basal activity and likely exert their pathogenicity through alternative mechanisms. We lack a molecular understanding of how these SH2 mutations impact SHP2 structure, activity, and signaling. Here, we characterize five SHP2 SH2 domain ligand-binding pocket mutants through a combination of high-throughput biochemical screens, biophysical and biochemical measurements, and molecular dynamics simulations. We show that while some of these mutations alter binding affinity to phosphorylation sites, the T42A mutation in the N-SH2 domain is unique in that it also substantially alters ligand-binding specificity, despite being 8 to 10 Å from the specificity-determining region of the SH2 domain. This mutation exerts its effect on sequence specificity by remodeling the phosphotyrosine-binding pocket, altering the mode of engagement of both the phosphotyrosine and surrounding residues on the ligand. The functional consequence of this altered specificity is that the T42A mutant has biased sensitivity toward a subset of activating ligands and enhances downstream signaling. Our study highlights an example of a nuanced mechanism of action for a disease-associated mutation, characterized by a change in protein-protein interaction specificity that alters enzyme activation.

Heterogenous thinning of peripapillary tissues occurs early during high myopia development in juvenile tree shrews.

Myopia is an independent risk factor for glaucoma, but the link between both conditions remains unknown. Both conditions induce connective tissue remodeling at the optic nerve head (ONH), including the peripapillary tissues. The purpose of this study was to investigate the thickness changes of the peripapillary tissues during experimental high myopia development in juvenile tree shrews. Six juvenile tree shrews experienced binocular normal vision, while nine received monocular -10D lens treatment starting at 24 days of visual experience (DVE) to induce high myopia in one eye and the other eye served as control. Daily refractive and biometric measurements and weekly optical coherence tomography scans of the ONH were obtained for five weeks. Peripapillary sclera (Scl), choroid-retinal pigment epithelium complex (Ch-RPE), retinal nerve fiber layer (RNFL), and remaining retinal layers (RRL) were auto-segmented using a deep learning algorithm after nonlinear distortion correction. Peripapillary thickness values were quantified from 3D reconstructed segmentations. All lens-treated eyes developed high myopia (-9.8 ± 1.5 D), significantly different (P < 0.001) from normal (0.69 ± 0.45 D) and control eyes (0.76 ± 1.44 D). Myopic eyes showed significant thinning of all peripapillary tissues compared to both, normal and control eyes (P < 0.001). At the experimental end point, the relative thinning from baseline was heterogeneous across tissues and significantly more pronounced in the Scl (-8.95 ± 3.1%) and Ch-RPE (-16.8 ± 5.8%) when compared to the RNFL (-5.5 ± 1.6%) and RRL (-6.7 ± 1.8%). Furthermore, while axial length increased significantly throughout the five weeks of lens wear, significant peripapillary tissue thinning occurred only during the first week of the experiment (until a refraction of -2.5 ± 1.9 D was reached) and ceased thereafter. A sectorial analysis revealed no clear pattern. In conclusion, our data show that in juvenile tree shrews, experimental high myopia induces significant and heterogeneous thinning of the peripapillary tissues, where the retina seems to be protected from profound thickness changes as seen in Ch-RPE and Scl. Peripapillary tissue thinning occurs early during high myopia development despite continued progression of axial elongation. The observed heterogeneous thinning may contribute to the increased risk for pathological optic nerve head remodeling and glaucoma later in life.

CD4 receptor diversity represents an ancient protection mechanism against primate lentiviruses.

Infection with human and simian immunodeficiency viruses (HIV/SIV) requires binding of the viral envelope glycoprotein (Env) to the host protein CD4 on the surface of immune cells. Although invariant in humans, the Env binding domain of the chimpanzee CD4 is highly polymorphic, with nine coding variants circulating in wild populations. Here, we show that within-species CD4 diversity is not unique to chimpanzees but found in many African primate species. Characterizing the outermost (D1) domain of the CD4 protein in over 500 monkeys and apes, we found polymorphic residues in 24 of 29 primate species, with as many as 11 different coding variants identified within a single species. D1 domain amino acid replacements affected SIV Env-mediated cell entry in a single-round infection assay, restricting infection in a strain- and allele-specific fashion. Several identical CD4 polymorphisms, including the addition of N-linked glycosylation sites, were found in primate species from different genera, providing striking examples of parallel evolution. Moreover, seven different guenons (Cercopithecus spp.) shared multiple distinct D1 domain variants, pointing to long-term trans-specific polymorphism. These data indicate that the HIV/SIV Env binding region of the primate CD4 protein is highly variable, both within and between species, and suggest that this diversity has been maintained by balancing selection for millions of years, at least in part to confer protection against primate lentiviruses. Although long-term SIV-infected species have evolved specific mechanisms to avoid disease progression, primate lentiviruses are intrinsically pathogenic and have left their mark on the host genome.

Hif1α and Wnt are required for posterior gene expression during Xenopus tropicalis tail regeneration.

Regeneration of complex tissues is initiated by an injury-induced stress response, eventually leading to activation of developmental signaling pathways such as Wnt signaling. How early injury cues are interpreted and coupled to activation of these developmental signals and their targets is not well understood. Here, we show that Hif1α, a stress induced transcription factor, is required for tail regeneration in Xenopus tropicalis. We find that Hif1α is required for regeneration of differentiated axial tissues, including axons and muscle. Using RNA-sequencing, we find that Hif1α and Wnt converge on a broad set of genes required for posterior specification and differentiation, including the posterior hox genes. We further show that Hif1α is required for transcription via a Wnt-responsive element, a function that is conserved in both regeneration and early neural patterning. Our findings indicate that Hif1α has regulatory roles in Wnt target gene expression across multiple tissue contexts.

Localization of Spiropyran Activation.

Functionalization of planar and curved glass surfaces with spiropyran (SP) molecules and localized UV-induced activation of the mechanophore are demonstrated. Fluorescence spectra of UV-irradiated SP-functionalized surfaces reveal that increases in surface roughness or curvature produce more efficient conversion of the mechanophore to the open merocyanine (MC) form. Further, force-induced activation of the mechanophore is achieved at curved glass-polymer interfaces and not planar interfaces. Minimal fluorescence signal from UV-irradiated SP-functionalized planar glass surfaces precluded mechanical activation testing. Curved glass-polymer interfaces are prepared by SP functionalization of E-glass fibers, which are subsequently embedded in a poly(methyl methacrylate) (PMMA) matrix. Mechanical activation is induced through shear loading by a single fiber microbond testing protocol. In situ detection of SP activation at the interface is monitored by fluorescence spectroscopy. The fluorescence increase during interfacial testing suggests that attachment of the interfacial SP molecule to both fiber surface and polymer matrix is present and able to achieve significant activation of SP at the fiber-polymer matrix interface. Unlike previous studies for bulk polymers, SP activation is detected at relatively low levels of applied shear stress. By linking SP at the glass-polymer interface and transferring load directly to that interface, a more efficient mechanism for eliciting the SP response is achieved.

Comparable, but distinct: Perceptions of primary care provided by physicians and nurse practitioners in full and restricted practice authority states.

AIMS: To understand patients' and providers' perceptions of primary care delivered by nurse practitioners (NPs) in the Veterans Affairs Healthcare System. DESIGN: Qualitative exploratory study (in convergent mixed-methods design). METHODS: Semi-structured interviews in 2016 with primary care providers and patients from facilities in states with full and restricted practice authority for NPs. Patient sample based on reassignment to: (a) a NP; or (b) a different physician following an established physician relationship. Data were analysed using content analysis. RESULTS: We interviewed 28 patients, 17 physicians and 14 NPs. We found: (a) NPs provided more holistic care than physicians; (b) patients were satisfied with NPs; and (c) providers' professional experience outweighed provider type. CONCLUSIONS: Patients' preferences for NPs (compared with prior physicians) contributed to perceptions of patient centredness. Similarities in providers' perceptions suggest NPs and physicians are both viable providers for primary care. IMPACT: Nurse Practitioners (NPs): practice authority Veterans Affairs Health care: nurse practitioners will continue to be a viable resource for primary care delivery United States Health care: challenges notions patients may not be satisfied with care provided by NPs and supports expanding their use to provide much-needed access to primary care services; expanding Full Practice Authority would allow states to provide acceptable primary care without diminishing patient or provider experiences.

Preadaptation of Simian Immunodeficiency Virus SIVsmm Facilitated Env-Mediated Counteraction of Human Tetherin by Human Immunodeficiency Virus Type 2.

The host restriction factor tetherin inhibits virion release from infected cells and poses a significant barrier to successful zoonotic transmission of primate lentiviruses to humans. While most simian immunodeficiency viruses (SIV), including the direct precursors of human immunodeficiency virus type 1 (HIV-1) and HIV-2, use their Nef protein to counteract tetherin in their natural hosts, they fail to antagonize the human tetherin ortholog. Pandemic HIV-1 group M and epidemic group O strains overcame this hurdle by adapting their Vpu and Nef proteins, respectively, whereas HIV-2 group A uses its envelope (Env) glycoprotein to counteract human tetherin. Whether or how the remaining eight groups of HIV-2 antagonize this antiviral factor has remained unclear. Here, we show that Nef proteins from diverse groups of HIV-2 do not or only modestly antagonize human tetherin, while their ability to downmodulate CD3 and CD4 is highly conserved. Experiments in transfected cell lines and infected primary cells revealed that not only Env proteins of epidemic HIV-2 group A but also those of a circulating recombinant form (CRF01_AB) and rare groups F and I decrease surface expression of human tetherin and significantly enhance progeny virus release. Intriguingly, we found that many SIVsmm Envs also counteract human as well as smm tetherin. Thus, Env-mediated tetherin antagonism in different groups of HIV-2 presumably stems from a preadaptation of their SIVsmm precursors to humans. In summary, we identified a phenotypic trait of SIVsmm that may have facilitated its successful zoonotic transmission to humans and the emergence of HIV-2.IMPORTANCE HIV-2 groups A to I resulted from nine independent cross-species transmission events of SIVsmm to humans and differ considerably in their prevalence and geographic spread. Thus, detailed characterization of these viruses offers a valuable means to elucidate immune evasion mechanisms and human-specific adaptations determining viral spread. In a systematic comparison of rare and epidemic HIV-2 groups and their simian SIVsmm counterparts, we found that the ability of Nef to downmodulate the primary viral entry receptor CD4 and the T cell receptor CD3 is conserved, while effects on CD28, CD74, and major histocompatibility complex class I surface expression vary considerably. Furthermore, we show that not only the Env proteins of HIV-2 groups A, AB, F, and I but also those of some SIVsmm isolates antagonize human tetherin. This finding helps to explain why SIVsmm has been able to cross the species barrier to humans on at least nine independent occasions.

Mindfulness-based cognitive therapy for the treatment of current depressive symptoms: a meta-analysis.

Mindfulness-based cognitive therapy (MBCT) appears to be a promising intervention for the prevention of relapse in major depressive disorder, but its efficacy in patients with current depressive symptoms is less clear. Randomized clinical trials of MBCT for adult patients with current depressive symptoms were included (k = 13, N = 1046). Comparison conditions were coded based on whether they were intended to be therapeutic (specific active controls) or not (non-specific controls). MBCT was superior to non-specific controls at post-treatment (k = 10, d = 0.71, 95% confidence interval [CI] [0.47, 0.96]), although not at longest follow-up (k = 2, d = 1.47, [-0.71, 3.65], mean follow-up = 5.70 months across all studies with follow-up). MBCT did not differ from other active therapies at post-treatment (k = 6, d = 0.002, [-0.43, 0.44]) and longest follow-up (k = 4, d = 0.26, [-0.24, 0.75]). There was some evidence that studies with higher methodological quality showed smaller effects at post-treatment, but no evidence that effects varied by inclusion criterion. The impact of publication bias appeared minimal. MBCT seems to be efficacious for samples with current depressive symptoms at post-treatment, although a limited number of studies tested the long-term effects of this therapy.

Challenges to Educating Smokers About Lung Cancer Screening: a Qualitative Study of Decision Making Experiences in Primary Care.

We sought to qualitatively explore how those at highest risk for lung cancer, current smokers, experienced, understood, and made decisions about participation in lung cancer screening (LCS) after being offered in the target setting for implementation, routine primary care visits. Thirty-seven current smokers were identified within 4 weeks of being offered LCS at seven sites participating in the Veterans Health Administration Clinical Demonstration Project and interviewed via telephone using semi-structured qualitative interviews. Transcripts were coded by two raters and analyzed thematically using iterative inductive content analysis. Five challenges to smokers' decision-making lead to overestimated benefits and minimized risks of LCS: fear of lung cancer fixated focus on inflated screening benefits; shame, regret, and low self-esteem stemming from continued smoking situated screening as less averse and more beneficial; screening was mistakenly believed to provide general evaluation of lungs and reassurance was sought about potential damage caused by smoking; decision-making was deferred to providers; and indifference about numerical educational information that was poorly understood. Biased understanding of risks and benefits was complicated by emotion-driven, uninformed decision-making. Emotional and cognitive biases may interfere with educating and supporting smokers' decision-making and may require interventions tailored for their unique needs.

The Effects of Different Spectrophotometric Modes on Colour Measurement of Resin Composite and Porcelain Materials.

OBJECTIVES: To investigate the influence of a number of controlled modes on the spectrophotometric analysis of the colour of resin composite and porcelain materials. METHODS: A total of 20 samples of commercially available resin composite, and 20 samples of commercially available porcelain materials in four different shades were produced (five samples for each shade). Colour was measured using a spectrophotometer (CM2600-d, Minolta Konica) set with different colour measuring modes namely, small aperture size (SAV) or large aperture size (MAV); specular component included (SCI), or excluded (SCE); 0% (UV-) or 100% UV illumination (UV+). Colour data were then compared using paired T-test. RESULTS: Colour coordinates measured with spectrophotometric modes set as 2° observation angle, SAV, SCI, and UV- were significantly different from those measured with 10° observation angle, MAV, SCE, and UV+ respectively in most cases for both materials. CONCLUSION: Different spectrophotometric modes (2 or 10 degrees observation angle, SAV or MAV aperture size, 0% or 100% UV, and SCI or SCE) significantly influenced the absolute colour measurements of resin composite and porcelain samples. CLINICAL SIGNIFICANCE: Measuring modes should be taken into consideration when comparing the results of absolute colour measurements of resin composite and dental porcelain materials.

The Effect of Try-In Paste and Resin Cement Shade on Colour Properties of Dental Veneers.

OBJECTIVES: The purpose of this study was to investigate how different shades of try-in pastes, uncured and cured resin cements affect the overall colour of porcelain veneer restorations. METHODS: A total of 90 porcelain veneers of shade Vita 1M1 VM7 and 0.6 mm thick were applied to bovine teeth using 3 shades of resin cement and their try-in paste produced by 3 manufacturers. RESULTS: Colour differences produced between the try-in pastes and the corresponding shades of cured resin cements ranged from (ΔE* 1.18-3.1). The colour differences between uncured and cured resins of the same shade ranged from (ΔE* 0.78 - 1.41). CONCLUSIONS: Different shades of try-in pastes and resin cements produced colour changes which are clinically useful in changing the colour of veneer restorations and therefore assist in colour matching to adjacent teeth. Clinically significant differences were noticed between try-in pastes and the cured resin of the same shade, however, there were relatively small changes measured between un-cured and cured resins. The colour match obtained by the try-in paste has to be treated with caution and further evaluation of the restoration made with the resin in place before curing is recommended.

Zoonotic Potential of Simian Arteriviruses.

Wild nonhuman primates are immediate sources and long-term reservoirs of human pathogens. However, ethical and technical challenges have hampered the identification of novel blood-borne pathogens in these animals. We recently examined RNA viruses in plasma from wild African monkeys and discovered several novel, highly divergent viruses belonging to the family Arteriviridae. Close relatives of these viruses, including simian hemorrhagic fever virus, have caused sporadic outbreaks of viral hemorrhagic fever in captive macaque monkeys since the 1960s. However, arterivirus infection in wild nonhuman primates had not been described prior to 2011. The arteriviruses recently identified in wild monkeys have high sequence and host species diversity, maintain high viremia, and are prevalent in affected populations. Taken together, these features suggest that the simian arteriviruses may be "preemergent" zoonotic pathogens. If not, this would imply that biological characteristics of RNA viruses thought to facilitate zoonotic transmission may not, by themselves, be sufficient for such transmission to occur.

Derivation and Characterization of a CD4-Independent, Non-CD4-Tropic Simian Immunodeficiency Virus.

UNLABELLED: CD4 tropism is conserved among all primate lentiviruses and likely contributes to viral pathogenesis by targeting cells that are critical for adaptive antiviral immune responses. Although CD4-independent variants of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) have been described that can utilize the coreceptor CCR5 or CXCR4 in the absence of CD4, these viruses typically retain their CD4 binding sites and still can interact with CD4. We describe the derivation of a novel CD4-independent variant of pathogenic SIVmac239, termed iMac239, that was used to derive an infectious R5-tropic SIV lacking a CD4 binding site. Of the seven mutations that differentiate iMac239 from wild-type SIVmac239, a single change (D178G) in the V1/V2 region was sufficient to confer CD4 independence in cell-cell fusion assays, although other mutations were required for replication competence. Like other CD4-independent viruses, iMac239 was highly neutralization sensitive, although mutations were identified that could confer CD4-independent infection without increasing its neutralization sensitivity. Strikingly, iMac239 retained the ability to replicate in cell lines and primary cells even when its CD4 binding site had been ablated by deletion of a highly conserved aspartic acid at position 385, which, for HIV-1, plays a critical role in CD4 binding. iMac239, with and without the D385 deletion, exhibited an expanded host range in primary rhesus peripheral blood mononuclear cells that included CCR5(+) CD8(+) T cells. As the first non-CD4-tropic SIV, iMac239-ΔD385 will afford the opportunity to directly assess the in vivo role of CD4 targeting on pathogenesis and host immune responses. IMPORTANCE: CD4 tropism is an invariant feature of primate lentiviruses and likely plays a key role in pathogenesis by focusing viral infection onto cells that mediate adaptive immune responses and in protecting virions attached to cells from neutralizing antibodies. Although CD4-independent viruses are well described for HIV and SIV, these viruses characteristically retain their CD4 binding site and can engage CD4 if available. We derived a novel CD4-independent, CCR5-tropic variant of the pathogenic molecular clone SIVmac239, termed iMac239. The genetic determinants of iMac239's CD4 independence provide new insights into mechanisms that underlie this phenotype. This virus remained replication competent even after its CD4 binding site had been ablated by mutagenesis. As the first truly non-CD4-tropic SIV, lacking the capacity to interact with CD4, iMac239 will provide the unique opportunity to evaluate SIV pathogenesis and host immune responses in the absence of the immunomodulatory effects of CD4(+) T cell targeting and infection.

Screening for Housing Instability and Homelessness Among Families Undergoing Child Maltreatment Investigation.

This paper reports results of mixed methods, population survey of housing instability, and homelessness. Child welfare personnel conducted the Quick Risks and Assets for Family Triage (QRAFT), a three-question screening tool intended to identify housing instability and homelessness. The QRAFT requires users to assess family housing history, current housing arrangement, and current housing condition, on a four-point scale from "asset/not a risk" to "severe risk." The QRAFT was completed among 6828 families undergoing new child maltreatment investigations. Approximately 5.4% of families demonstrated significant to severe housing problems; approximately one-third exhibited moderate housing risk. Housing problems and homelessness were significantly associated with the outcome of child welfare investigations; among families with substantiated child welfare determinations, 21% demonstrated significant to severe housing risk, a significantly higher proportion than among families where the investigation outcome was unsubstantiated or differential response (i.e., voluntary services). Of significant to severe housing risk families, 15.7% later met eligibility criteria for a supportive housing intervention, suggesting that housing concerns combined with substantial parent and child functional difficulties. Qualitative data indicated the QRAFT was perceived as easy to administer, effective as a screening tool, and useful to "apply the housing lens" early in child welfare involvement.

Bordetella parapertussis PagP mediates the addition of two palmitates to the lipopolysaccharide lipid A.

Bordetella bronchiseptica PagP (PagPBB) is a lipid A palmitoyl transferase that is required for resistance to antibody-dependent complement-mediated killing in a murine model of infection. B. parapertussis contains a putative pagP homolog (encoding B. parapertussis PagP [PagPBPa]), but its role in the biosynthesis of lipid A, the membrane anchor of lipopolysaccharide (LPS), has not been investigated. Mass spectrometry analysis revealed that wild-type B. parapertussis lipid A consists of a heterogeneous mixture of lipid A structures, with penta- and hexa-acylated structures containing one and two palmitates, respectively. Through mutational analysis, we demonstrate that PagPBPa is required for the modification of lipid A with palmitate. While PagPBB transfers a single palmitate to the lipid A C-3' position, PagPBPa transfers palmitates to the lipid A C-2 and C-3' positions. The addition of two palmitate acyl chains is unique to B. parapertussis. Mutation of pagPBPa resulted in a mutant strain with increased sensitivity to antimicrobial peptide killing and decreased endotoxicity, as evidenced by reduced proinflammatory responses via Toll-like receptor 4 (TLR4) to the hypoacylated LPS. Therefore, PagP-mediated modification of lipid A regulates outer membrane function and may be a means to modify interactions between the bacterium and its human host during infection.

Facilitators and barriers to weight loss and weight loss maintenance: a qualitative exploration.

BACKGROUND: The present study aimed to explore facilitators and barriers to weight loss (WL) and weight loss maintenance (WLM) in women who participated in a primary, 18-week comparative trial that promoted WL with an energy-restricted diet. METHODS: Twenty-three women participated in seven focus groups conducted by a moderator and co-facilitator using open-ended questions and probes. Focus groups were held in a private room and audio tape-recorded. Tapes were transcribed verbatim and thematic analysis was used to evaluate transcripts for common themes. RESULTS: Accountability to others, social support, planning ahead, awareness and mindfulness of food choices, basic nutrition education, portion control, exercise, and self-motivation were perceived as key facilitators for WL and WLM by women. Identified barriers included life transitions, health status changes, internal factors, environmental pressures, lack of accountability and an absence of social support. CONCLUSIONS: Future interventions should address these salient facilitators and barriers to promote sustainable changes in women across their WL and WLM journeys.

Influences of acute alcohol consumption, sexual precedence, and relationship motivation on women's relationship and sex appraisals and unprotected sex intentions.

Guided by the cognitive mediation model of sexual decision making (Norris, Masters, & Zawacki, 2004. Cognitive mediation of women's sexual decision making: The influence of alcohol, contextual factors, and background variables. Annual Review of Sex Research, 15, 258-296), we examined female social drinkers' (N = 162) in-the-moment risky sexual decision making by testing how individual differences (relationship motivation) and situational factors (alcohol consumption and sexual precedence conditions) influenced cognitive appraisals and sexual outcomes in a hypothetical sexual scenario. In a path model, acute intoxication, sexual precedence, and relationship motivation interactively predicted primary relationship appraisals and independently predicted primary sex appraisals. Primary appraisals predicted secondary appraisals related to relationship and unprotected sex, which predicted unprotected sex intentions. Sexual precedence directly increased unprotected sex intentions. Findings support the cognitive mediation model and suggest that sexual risk reduction interventions should address alcohol, relationship, sexual, and cognitive factors.