HCMV US2 co-opts TRC8 to degrade the endoplasmic reticulum-resident protein LMAN2L.

The human cytomegalovirus (HCMV) pUS2 glycoprotein exploits the host's endoplasmic reticulum (ER)-associated degradation (ERAD) pathway to degrade major histocompatibility complex class I (MHC-I) and prevent antigen presentation. Beyond MHC-I, pUS2 has been shown to target a range of cellular proteins for degradation, preventing their cell surface expression. Here we have identified a novel pUS2 target, ER-resident protein lectin mannose binding 2 like (LMAN2L). pUS2 expression was both necessary and sufficient for the downregulation of LMAN2L, which was dependent on the cellular E3 ligase TRC8. Given the hypothesized role of LMAN2L in the trafficking of glycoproteins, we employed proteomic plasma membrane profiling to measure LMAN2L-dependent changes at the cell surface. A known pUS2 target, integrin alpha-6 (ITGA6), was downregulated from the surface of LMAN2L-deficient cells, but not other integrins. Overall, these results suggest a novel strategy of pUS2-mediated protein degradation whereby pUS2 targets LMAN2L to impair trafficking of ITGA6. Given that pUS2 can directly target other integrins, we propose that this single viral protein may exhibit both direct and indirect mechanisms to downregulate key cell surface molecules.

Human cytomegalovirus degrades DMXL1 to inhibit autophagy, lysosomal acidification, and viral assembly.

Human cytomegalovirus (HCMV) is an important human pathogen that regulates host immunity and hijacks host compartments, including lysosomes, to assemble virions. We combined a quantitative proteomic analysis of HCMV infection with a database of proteins involved in vacuolar acidification, revealing Dmx-like protein-1 (DMXL1) as the only protein that acidifies vacuoles yet is degraded by HCMV. Systematic comparison of viral deletion mutants reveals the uncharacterized 7 kDa US33A protein as necessary and sufficient for DMXL1 degradation, which occurs via recruitment of the E3 ubiquitin ligase Kip1 ubiquitination-promoting complex (KPC). US33A-mediated DMXL1 degradation inhibits lysosome acidification and autophagic cargo degradation. Formation of the virion assembly compartment, which requires lysosomes, occurs significantly later with US33A-expressing virus infection, with reduced viral replication. These data thus identify a viral strategy for cellular remodeling, with the potential to employ US33A in therapies for viral infection or rheumatic conditions, in which inhibition of lysosome acidification can attenuate disease.

Examining teacher approaches to implementation of a classwide SEL program.

Scale-up of universal social-emotional learning (SEL) programs has become a priority in schools to promote positive social outcomes for all students. Although studies have examined student outcomes associated with school-based SEL when extensive training and support are provided, research on teacher SEL implementation practices under typical conditions has received far less attention. As such, this study examined the first-year universal SEL implementation practices of 41 teachers across 13 schools in three states. School personnel made all decisions regarding approaches to training, support, and program implementation of a manualized program delivered at the classroom level to first- and second-grade students. Within this authentic context for implementation, variability was observed in selection of skills units, number of lessons taught, and adherence to lesson activities and scripts. Coding of teacher anecdotal reports revealed that perceived student needs, classroom context demands, and school-level factors informed teachers' implementation of the program. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Human cytomegalovirus protein RL1 degrades the antiviral factor SLFN11 via recruitment of the CRL4 E3 ubiquitin ligase complex.

Human cytomegalovirus (HCMV) is an important human pathogen and a paradigm of viral immune evasion, targeting intrinsic, innate, and adaptive immunity. We have employed two orthogonal multiplexed tandem mass tag-based proteomic screens to identify host proteins down-regulated by viral factors expressed during the latest phases of viral infection. This approach revealed that the HIV-1 restriction factor Schlafen-11 (SLFN11) was degraded by the poorly characterized, late-expressed HCMV protein RL1, via recruitment of the Cullin4-RING E3 Ubiquitin Ligase (CRL4) complex. SLFN11 potently restricted HCMV infection, inhibiting the formation and spread of viral plaques. Overall, we show that a restriction factor previously thought only to inhibit RNA viruses additionally restricts HCMV. We define the mechanism of viral antagonism and also describe an important resource for revealing additional molecules of importance in antiviral innate immunity and viral immune evasion.

Predicting school readiness program implementation in community-based childcare centers.

Introduction: Targeted curricular interventions can increase preschool program quality and boost children's academic and social-emotional readiness skills, but variable funding and weak organizational infrastructure in many community-based childcare centers may reduce the effective implementation of these programs. Method: This study examined individual teacher and workplace predictors of the REDI program implementation, a targeted school readiness program that was adapted to support delivery in childcare centers. REDI was delivered by 63 teachers in 37 community-based childcare centers with center directors serving as local implementation coaches. Results: Results showed that individual teacher factors (e.g., teaching skills and receptivity to intervention consultation) predicted the quality with which REDI activities and teaching strategies were delivered, and workplace factors were important predictors across multiple implementation indicators. Discussion: Practice and policy implications for improving intervention implementation and corresponding program quality in childcare centers are highlighted.

Comparative Cell Surface Proteomic Analysis of the Primary Human T Cell and Monocyte Responses to Type I Interferon.

The cellular response to interferon (IFN) is essential for antiviral immunity, IFN-based therapy and IFN-related disease. The plasma membrane (PM) provides a critical interface between the cell and its environment, and is the initial portal of entry for viruses. Nonetheless, the effect of IFN on PM proteins is surprisingly poorly understood, and has not been systematically investigated in primary immune cells. Here, we use multiplexed proteomics to quantify IFNα2a-stimulated PM protein changes in primary human CD14+ monocytes and CD4+ T cells from five donors, quantifying 606 and 482 PM proteins respectively. Comparison of cell surface proteomes revealed a remarkable invariance between donors in the overall composition of the cell surface from each cell type, but a marked donor-to-donor variability in the effects of IFNα2a. Furthermore, whereas only 2.7% of quantified proteins were consistently upregulated by IFNα2a at the surface of CD4+ T cells, 6.8% of proteins were consistently upregulated in primary monocytes, suggesting that the magnitude of the IFNα2a response varies according to cell type. Among these differentially regulated proteins, we found the viral target Endothelin-converting enzyme 1 (ECE1) to be an IFNα2a-stimulated protein exclusively upregulated at the surface of CD4+ T cells. We therefore provide a comprehensive map of the cell surface of IFNα2a-stimulated primary human immune cells, including previously uncharacterized interferon stimulated genes (ISGs) and candidate antiviral factors.

Implementing a School Readiness Intervention in Community-based Childcare Centers: Director and Teacher Perceptions.

In the U.S., one-third of preschool children attend programs run by childcare centers. Childcare centers are generally isolated and under-resourced businesses, often challenged by high rates of teacher turnover and inconsistent financial support. Correspondingly, childcare centers often struggle to provide high-quality educational experiences for preschool children. This study introduced an evidence-based curriculum (REDI) to improve the educational experiences of preschool children in childcare centers and used a novel professional development (PD) model in which center directors were trained to serve as local coaches. Open-ended interviews with 45 teachers and 31 center directors evaluated the acceptability, feasibility, and effectiveness of the REDI intervention and coaching model. Participants also described their workplace and rated their intentions regarding future program use. Participants described multiple challenges in the workplace but generally positive perceptions of the REDI intervention. Perceptions regarding the feasibility of the coaching model were mixed. Workplace descriptions were associated with intervention experiences which, in turn, predicted intentions for future program use. Findings validate the importance of addressing the unique workplace challenges faced by childcare staff when designing classroom interventions and PD supports. Attending to program acceptability, feasibility, and perceived effectiveness in these settings may be critical to support sustained use.

Temporal Proteomic Analysis of Herpes Simplex Virus 1 Infection Reveals Cell-Surface Remodeling via pUL56-Mediated GOPC Degradation.

Herpesviruses are ubiquitous in the human population and they extensively remodel the cellular environment during infection. Multiplexed quantitative proteomic analysis over the time course of herpes simplex virus 1 (HSV-1) infection was used to characterize changes in the host-cell proteome and the kinetics of viral protein production. Several host-cell proteins are targeted for rapid degradation by HSV-1, including the cellular trafficking factor Golgi-associated PDZ and coiled-coil motif-containing protein (GOPC). We show that the poorly characterized HSV-1 pUL56 directly binds GOPC, stimulating its ubiquitination and proteasomal degradation. Plasma membrane profiling reveals that pUL56 mediates specific changes to the cell-surface proteome of infected cells, including loss of interleukin-18 (IL18) receptor and Toll-like receptor 2 (TLR2), and that cell-surface expression of TLR2 is GOPC dependent. Our study provides significant resources for future investigation of HSV-host interactions and highlights an efficient mechanism whereby a single virus protein targets a cellular trafficking factor to modify the surface of infected cells.

Histone deacetylase inhibitors valproic acid and vorinostat enhance trastuzumab-mediated antibody-dependent cell-mediated phagocytosis.

BACKGROUND: The monoclonal antibody (mAb) trastuzumab is part of the standard of care for patients with human epidermal growth factor receptor 2 (HER2)-overexpressing breast cancer. Antibody-dependent cell-mediated phagocytosis (ADCP) and cytotoxicity (ADCC) are major mechanisms of action of the mAb trastuzumab. Histone deacetylase inhibitors (HDACi), such as valproic acid (VPA) or vorinostat (SAHA), exert several immunostimulatory properties, which contribute at least in part to their anticancer effect. However, the impact of HDACi-induced immunostimulatory effects on trastuzumab-mediated anti-tumor immune response is not well characterized. METHODS: We analyzed the ADCP and ADCC activity of peripheral blood mononuclear cells (PBMCs) from age and gender-matched healthy volunteers (n=5) against HDACi-treated HER2-overexpressing breast cancer cells (SKBR3), using a well-established in vitro three-color imaging flow cytometry and flow cytometry approach. RESULTS: VPA and SAHA enhanced trastuzumab-mediated ADCP and trastuzumab-independent cytotoxicity. Mechanistically, VPA upregulated the activating antibody-binding receptor Fc-gamma receptor (FcγR) IIA (CD32A) on monocytes (CD14+). Moreover, VPA and SAHA downregulated the anti-apoptotic protein myeloid leukemia cell differentiation 1 (MCL1) in breast cancer cells. Additionally, VPA and SAHA induced an immunogenic cell death, characterized by the exposure of calreticulin (CALR), as well as decreased the "do not eat me" signal CD47 on tumor cells. CONCLUSIONS: HDACi VPA and SAHA increase trastuzumab-mediated phagocytosis and trastuzumab-independent cytotoxicity. The immunomodulatory activities of those HDACi support a rationale combined treatment approach with mAb for cancer treatment.

A Novel Approach to Gross Dissection of the Human Pelvis and Perineum.

Progressive curricular changes in medical education over the past two decades have resulted in the diaspora of gross anatomy content into integrated curricula while significantly reducing total contact hours. Despite the development of a wide range of alternative teaching modalities, gross dissection remains a critical component of medical education. The challenge posed to modern anatomists is how to maximize and integrate the time spent dissecting under the current curricular changes. In this study, an alternative approach to the dissection of the pelvis and perineum is presented in an effort to improve content delivery and student satisfaction. The approach involves removal of the perineum en bloc from the cadaver followed by excision of the pubic symphysis, removal and examination of the bladder and associated structures, examination and bisection of the midline pelvic organs in situ, and midsagittal hemisection of the pelvis for identification of the neurovasculature. Results indicate that this novel dissecting approach increases the number of structures identified by 46% ± 14% over current dissecting methods. Survey results indicate that students were better able to integrate lecture and laboratory concepts, understand the concepts, and successfully identify more structures using the new approach (P < 0.05). The concept of anatomic efficiency is introduced and proposed as a standard quantitative measure of gross dissection proficiency across programs and institutions. These findings provide evidence that innovative solutions to anatomy education can be found that help to maintain critical content and student satisfaction in a modern medical curriculum.

At what cost? Examining the cost effectiveness of a universal social-emotional learning program.

Although implementation of universal social-emotional learning programs is becoming more common in schools, few studies have examined the cost-effectiveness of such programs. As such, the purpose of this article is two fold. First, we provide an overview of cost-effectiveness methods for school-based programs, and second, we share results of a cost-effectiveness analysis (CEA) of a universal social-emotional learning (SEL) program, the Social Skills Improvement System-Classwide Intervention Program (SSIS-CIP; Elliott & Gresham, 2007). Specifically, we compared the cost-effectiveness of SSIS-CIP implementation across first- and second-grade classrooms, and results indicated that second grade is the more cost-effective option for implementing the SSIS-CIP. Several considerations are discussed regarding cost-effectiveness analysis of universal SEL programs as well as the importance of using CEA results to inform programming decisions. (PsycINFO Database Record

Assessing Noncognitive Aspects of School Readiness: The Predictive Validity of Brief Teacher Rating Scales of Social-Emotional Competence and Approaches to Learning.

RESEARCH FINDINGS: Head Start teachers completed brief rating scales measuring the social-emotional competence and approaches to learning of preschool children (Total N = 164; 14% Hispanic-American, 30% African-American, 56% Caucasian; 56% girls). Head Start lead and assistant teacher ratings on both scales demonstrated strong internal consistency and moderate inter-rater reliability. When examined longitudinally, preschool teacher-rated approaches to learning made unique contributions to the prediction of kindergarten and first grade academic outcomes, need for supplemental services, and grade retention, even after accounting for preschool academic skills. In contrast, preschool teacher-rated social-emotional competence made unique contributions to the prediction of reduced behavior problems and peer difficulties in kindergarten and first grade. PRACTICE IMPLICATIONS: The findings demonstrate that preschool teachers are able to provide distinct and reliable ratings of child social-emotional competence and approaches to learning using brief rating scales, with validity for predicting elementary school adjustment.

A mixed-methods study of system-level sustainability of evidence-based practices in 12 large-scale implementation initiatives.

BACKGROUND: In recent decades, evidence-based practices (EBPs) have been broadly promoted in community behavioural health systems in the United States of America, yet reported EBP penetration rates remain low. Determining how to systematically sustain EBPs in complex, multi-level service systems has important implications for public health. This study examined factors impacting the sustainability of parent-child interaction therapy (PCIT) in large-scale initiatives in order to identify potential predictors of sustainment. METHODS: A mixed-methods approach to data collection was used. Qualitative interviews and quantitative surveys examining sustainability processes and outcomes were completed by participants from 12 large-scale initiatives. RESULTS: Sustainment strategies fell into nine categories, including infrastructure, training, marketing, integration and building partnerships. Strategies involving integration of PCIT into existing practices and quality monitoring predicted sustainment, while financing also emerged as a key factor. CONCLUSIONS: The reported factors and strategies impacting sustainability varied across initiatives; however, integration into existing practices, monitoring quality and financing appear central to high levels of sustainability of PCIT in community-based systems. More detailed examination of the progression of specific activities related to these strategies may aide in identifying priorities to include in strategic planning of future large-scale initiatives. TRIAL REGISTRATION: ClinicalTrials.gov ID NCT02543359 ; Protocol number PRO12060529.

Enrolling Family Participants in a Statewide Implementation Trial of an Evidence-Based Treatment.

BACKGROUND: Community-based research with families is subject to unique challenges in recruiting and enrolling participants, particularly when researchers do not have face-to-face contact with participants. OBJECTIVES: To identify strategies that overcome challenges associated with the recruitment and enrollment of family participants from community behavioral health settings into a research study. METHODS: We used a case study design to describe lessons learned during the recruitment and enrollment phases of a large-scale study of the implementation of an evidence-based treatment (EBT) across the Commonwealth of Pennsylvania. LESSONS LEARNED: Key lessons identified include the early involvement of stakeholders, engaging clinicians in the research process, establishing a research-community relationship, and using a flexible, ongoing recruitment approach. CONCLUSIONS: In large-scale community-based research, engagement, recruitment, and enrollment strategies are particularly crucial. The strategies included in this paper are relevant to multiple disciplines involving community-based research.