A supramodular FHA/BRCT-repeat architecture mediates Nbs1 adaptor function in response to DNA damage.

The Mre11/Rad50/Nbs1 protein complex plays central enzymatic and signaling roles in the DNA-damage response. Nuclease (Mre11) and scaffolding (Rad50) components of MRN have been extensively characterized, but the molecular basis of Nbs1 function has remained elusive. Here, we present a 2.3A crystal structure of the N-terminal region of fission yeast Nbs1, revealing an unusual but conserved architecture in which the FHA- and BRCT-repeat domains structurally coalesce. We demonstrate that diphosphorylated pSer-Asp-pThr-Asp motifs, recently identified as multicopy docking sites within Mdc1, are evolutionarily conserved Nbs1 binding targets. Furthermore, we show that similar phosphomotifs within Ctp1, the fission yeast ortholog of human CtIP, promote interactions with the Nbs1 FHA domain that are necessary for Ctp1-dependent resistance to DNA damage. Finally, we establish that human Nbs1 interactions with Mdc1 occur through both its FHA- and BRCT-repeat domains, suggesting how their structural and functional interdependence underpins Nbs1 adaptor functions in the DNA-damage response.

Effects of a Rehabilitation-at-Home Program Compared to Post-acute Skilled Nursing Facility Care on Safety, Readmission, and Community Dwelling Status: A Matched Cohort Analysis.

OBJECTIVES: To evaluate the effectiveness and safety of Rehabilitation-at-Home (RaH), which provides high-frequency, multidisciplinary post-acute rehabilitative services in patients' homes. DESIGN: Comparative effectiveness analysis. SETTING AND PARTICIPANTS: Medicare Fee-For-Service patients who received RaH in a Center for Medicare and Medicaid Innovation Center Demonstration during 2016-2017 (N=173) or who received Medicare Skilled Nursing Facility (SNF) care in 2016-2017 within the same geographic service area with similar inclusion and exclusion criteria (N=5535). METHODS: We propensity-matched RaH participants to a cohort of SNF patients using clinical and demographic characteristics with exact match on surgical and non-surgical hospitalizations. Outcomes included hospitalization within 30 days of post-acute admission, death within 30 days of post-acute discharge, length of stay, falls, use of antipsychotic medication, and discharge to community. RESULTS: The majority of RaH participants were older than or equal to 85 years (57.8%) and non-Hispanic white (72.2%) with mean hospital length of stay of 8.1 (SD 7.6) days. In propensity-matched analyses, 10.1% (95% CI: 0.5%, 19.8) and 4.2% (95% CI: 0.1%, 8.5%) fewer RaH participants experienced hospital readmission and death, respectively. RaH participants had, on average, 2.8 fewer days (95% CI 1.4, 4.3) of post-acute care; 11.4% (95% CI: 5.2%, 17.7%) fewer RaH participants experienced fall; and 25.8% (95% CI: 17.8%, 33.9%) more were discharged to the community. Use of antipsychotic medications was no different. CONCLUSIONS AND IMPLICATIONS: RaH is a promising alternative to delivering SNF-level post-acute RaH. The program seems to be safe, readmissions are lower, and transition back to the community is improved.

Benchmarking Site of Death and Hospice Use: A Case Study of Veterans Cared by Department of Veterans Affairs Home-based Primary Care.

OBJECTIVE: The objective of this study was to examine site of death and hospice use, identifying potential disparities among veterans dying in Department of Veterans Affairs (VA) Home Based Primary Care (VA-HBPC). METHODS: Administrative data (2008, 2012, and 2016) were compiled using the VA Residential-History-File which tracks health care service location, daily. Outcomes were site of death [home, nursing home (NH), hospital, inpatient hospice]; and hospice use on the day of death. We compared VA-HBPC rates to rates of 2 decedent benchmarks: VA patients and 5% Traditional Medicare non-veteran males. Potential age, race, urban/rural residence and living alone status disparities in rates among veterans dying in VA-HBPC in 2016 were examined by multinomial logistic regression. RESULTS: In 2016, 7796 veterans died in VA-HBPC of whom 62.1% died at home, 11.8% in NHs, 14.7% in hospitals and 11.4% in inpatient hospice. Hospice was provided to 60.9% of veterans dying at home and 63.9% of veterans dying in NH. Over the 2008-2012-2016 period, rates of VA-HBPC veterans who died at home and rates of home death with hospice increased and were higher than both benchmarks. Among VA-HBPC decedents, younger/older veterans were more/less likely to die at home and less/more likely to die with hospice. Race/ethnicity and urban/rural residence were unrelated to death at home but veterans living alone were less likely to die at home. CONCLUSIONS: Results reflect VA-HBPC's primary goal of supporting its veterans at home, including at the end-of-life, surpassing other population benchmarks with some potential disparities remaining.

Mobility Improvements are Found in Older Veterans After 6-Months of Gerofit Regardless of BMI Classification.

Veterans represent a unique population of older adults as they are more likely to self-report disability and be overweight or obese compared to the general population. We sought to compare changes in mobility function across the obesity spectrum in older Veterans participating in six-months of Gerofit, a clinical exercise program. 270 Veterans completed baseline, three, and six-month functional assessment and were divided post-hoc into groups: normal weight, overweight, and obese. Physical function assessment included: ten-meter walk time, six-minute walk distance, 30-second chair stands, and eight-foot up-and-go time. No significant weight x time interactions were found for any measure. However, significant (P<0.02) improvements were found for all mobility measures from baseline to three-months and maintained at six-months. Six-months of participation in Gerofit, if enacted nationwide, appears to be one way to improve mobility and function in older Veterans at high risk for disability regardless of weight status.

The Impact of Chain Standardization on Nursing Home Staffing.

BACKGROUND: Standardization in production is common in multientity chain organizations. Although chains are prominent in the nursing home sector, standardization in care has not been studied. One way nursing home chains may standardize is by controlling the level and mix of staffing in member homes. OBJECTIVES: To examine the extent to which standardization occurred in staffing, its relative presence across different types of chains, and whether facilities became more standardized following acquisition by a chain. RESEARCH DESIGN: We estimated predictors of the difference between facility and chain staffing using Generalized Estimating Equations with 2000-2010 data. SUBJECTS: This study included nursing homes nationally, excluding hospital-based homes and homes in Alaska, Hawaii, and the District of Columbia. MEASURES: Chain ownership was coded from text identifying chain names. Two nurse staffing measures were used: staff hours per resident day and staff mix. RESULTS: Very large for-profit chain nursing homes and large nonprofits had less variation in staff hours per resident day (P<0.001) but greater variation in staffing mix (P<0.001) compared with the chain average nationally. Large for-profit chains and medium nonprofit chains had greater dispersion on staff hours per resident day (P<0.001), while large nonprofit chains had greater dispersion in staffing mix (P<0.001). The difference between facility and chain staffing decreased over time. CONCLUSIONS: The largest chains (for-profit and nonprofit) had less staffing variation compared with national standards, suggesting they were best at implementing corporate practices. Following ownership changes, staffing converged towards chain averages over time, suggesting standardization takes time to implement.

HF-Free Boc Synthesis of Peptide Thioesters for Ligation and Cyclization.

We have developed a convenient method for the direct synthesis of peptide thioesters, versatile intermediates for peptide ligation and cyclic peptide synthesis. The technology uses a modified Boc SPPS strategy that avoids the use of anhydrous HF. Boc in situ neutralization protocols are used in combination with Merrifield hydroxymethyl resin and TFA/TMSBr cleavage. Avoiding HF extends the scope of Boc SPPS to post-translational modifications that are compatible with the milder cleavage conditions, demonstrated here with the synthesis of the phosphorylated protein CHK2. Peptide thioesters give easy, direct, access to cyclic peptides, illustrated by the synthesis of cyclorasin, a KRAS inhibitor.

The molecular basis of ATM-dependent dimerization of the Mdc1 DNA damage checkpoint mediator.

Mdc1 is a large modular phosphoprotein scaffold that maintains signaling and repair complexes at double-stranded DNA break sites. Mdc1 is anchored to damaged chromatin through interaction of its C-terminal BRCT-repeat domain with the tail of γH2AX following DNA damage, but the role of the N-terminal forkhead-associated (FHA) domain remains unclear. We show that a major binding target of the Mdc1 FHA domain is a previously unidentified DNA damage and ATM-dependent phosphorylation site near the N-terminus of Mdc1 itself. Binding to this motif stabilizes a weak self-association of the FHA domain to form a tight dimer. X-ray structures of free and complexed Mdc1 FHA domain reveal a 'head-to-tail' dimerization mechanism that is closely related to that seen in pre-activated forms of the Chk2 DNA damage kinase, and which both positively and negatively influences Mdc1 FHA domain-mediated interactions in human cells prior to and following DNA damage.

Factors Associated With Hospitalization by Veterans in Home-Based Primary Care.

OBJECTIVES: This study examined the extent to which program site-based and Veteran characteristics were associated with potentially avoidable hospitalizations or other hospitalization of Veterans enrolled in the Veterans Affairs (VA) Home-Based Primary Care (HBPC). DESIGN: Retrospective claims-based study. SETTING AND PARTICIPANTS: HBPC programs that responded to a national survey of HBPC programs (n = 189) in fiscal year (FY) 2016 were studied. Veterans in the analysis cohort were identified as having been enrolled in VA-HBPC in FY2016 who had not received care by VA-HBPC within 1 year prior to their first HBPC enrollment in FY2016 (N = 8497). METHODS: Multinomial logistic regression analysis with 5 outcome categories within the 6 months following the first HBPC enrollment date: (1) any potentially avoidable hospitalizations for ambulatory care-sensitive conditions (ACSC) as identified by AHRQ Prevention Quality Indicator (PQI), (2) any other hospitalizations for non-ACSC conditions, (3) died during study period, (4) discharged from HBPC, or (5) remained at home with HBPC. Average marginal effects (AME) of veteran-level and VA-HBPC-level covariates are reported for each of the outcome categories. RESULTS: More frail Veterans and Veterans 85 years old or older were more likely to have potentially preventable ACSC hospitalizations (AME = 5.4%, 1.8%, respectively). Veterans who were younger than 75 years, functionally impaired, bed-bound, or frail were more likely to have non-ACSC hospitalization (AME = 3.0%, 2.2%, 3.5%, and 9.0%, respectively). Veterans with low frailty index scores were less likely to have non-ACSC hospitalizations (AME = -17.1%). Six-month hospitalization patterns were not associated with reported HBPC site characteristics. CONCLUSIONS AND IMPLICATIONS: Within the framework of the national VA HBPC program, variations in the structural model used at HBPC sites are not significantly associated with hospitalizations. Tailoring of HBPC care, based on individual patient factors and clinical judgment rather than standard protocols, may be central to the success of HBPC in reducing ACSC hospitalizations.

Metabolic Switch and Cytotoxic Effect of Metformin on Burkitt Lymphoma.

Altered cellular energetic metabolism has recently emerged as important feature of neoplastic cells. Indeed, interfering with cancer cell metabolism might represent a suitable therapeutic strategy. In this study, we aimed to assess glucose metabolism activation in human lymphomas and evaluate how metformin can exert its action on lymphoma cells. We studied a large series of human lymphomas (N = 252) and an in vitro model of Burkitt lymphoma (BL) cells. We combined molecular biology techniques, including global gene expression profiling (GEP) analysis, quantitative PCR (qPCR) and Western blotting, and biochemical assays, aimed to assess pentose phosphate pathway, tricarboxylic acid (TCA) cycle, and aerobic glycolysis rates. We found that glucose metabolism is overall enhanced in most lymphoma subtypes, based on gene expression profiling (GEP), with general shift to aerobic glycolysis. By contrast, normal B cells only showed an overall increase in glucose usage during germinal center transition. Interestingly, not only highly proliferating aggressive lymphomas but also indolent ones, like marginal zone lymphomas, showed the phenomenon. Consistently, genes involved in glycolysis were confirmed to be overexpressed in BL cells by qPCR. Biochemical assays showed that while aerobic glycolysis is increased, TCA cycle is reduced. Finally, we showed that metformin can induce cell death in BL cells by stressing cellular metabolism through the induction of GLUT1, PKM2, and LDHA. In conclusion, we unveiled glucose metabolism abnormalities in human lymphomas and characterized the mechanism of action of metformin in Burkitt lymphoma model.

Access to phosphoproteins and glycoproteins through semi-synthesis, Native Chemical Ligation and N→S acyl transfer.

Peptide thioesters are important tools for protein synthesis and semi-synthesis through their use in Native Chemical Ligation (NCL). NCL can be employed to assemble site-specifically modified proteins that can help elucidate the mechanisms of biomolecular processes. In this article we explore the compatibility of phosphopeptide synthesis and glycopeptide synthesis with thioester production through N→S acyl transfer.

Effects of full-length borealin on the composition and protein-protein interaction activity of a binary chromosomal passenger complex.

The chromosomal passenger complex (CPC) comprises at least four protein components and functions at various cellular localizations during different mitotic stages to ensure correct chromosome segregation and completion of cytokinesis. Borealin, the most recently identified member of the CPC, is an intrinsically unstructured protein of low solubility and stability. Recent reports have demonstrated the formation of binary or ternary CPC subcomplexes incorporating short Borealin fragments in vitro. Using isothermal titration calorimetry, we show that full-length Borealin, instead of a Borealin fragment possessing the complete Survivin and INCENP recognition sequence, is required for the composition of a Borealin-Survivin complex competent to interact with INCENP. In addition, we show evidence that full-length Borealin, which forms high-order oligomers in its isolated form, is a monomer in the Borealin-Survivin CPC subcomplex.

Telehealth in Home-Based Primary Care: Factors and Challenges Associated With Integration Into Veteran Care.

OBJECTIVES: To describe the structural characteristics and challenges associated with home telehealth (HT) use in the US Department of Veterans Affairs (VA) Home-Based Primary Care (HBPC) program. DESIGN: We designed a national survey to collect information about HBPC program structural characteristics. The survey included eight organizational and service domains, one of which was HT. HBPC program directors were surveyed online using REDCap. PARTICIPANTS: We received 232 surveys from 394 HBPC sites (59% response rate). METHODS: HBPC structural domains were compared between sites using and not using HT technology. Open-ended responses were analyzed using content analysis. RESULTS: A total of 127 sites (76%) used HT, which was more likely when HBPC sites were aligned organizationally with the VA's Geriatrics and Extended Care Services division, when there were more disciplines on the HBPC team, and when primary care providers made home visits. Program directors overwhelmingly viewed HT as contributing to managing veterans' complex chronic conditions (81%), yet HT data were not readily integrated into care planning (24%). Challenges to HT use included veterans' acceptance and adherence, device issues, and collaboration between HBPC teams and HT staff. CONCLUSION: Corresponding to HBPC's complexity, HT use is primarily a self-organizing process that shapes the patterns of integration at each site. Although HT technology is compatible with core structures of the HBPC model, usability varies, and overall is low. To optimize HT use in HBPC, there are opportunities to redesign systems to mitigate challenges to adoption. As the Centers for Medicare and Medicaid Services' strives to increase access to both HBPC and telehealth benefits, evidenced by the continuation of its successful Independence at Home demonstration and the final changes in the proposed rule in April 2019 incorporating additional telehealth benefits for beneficiaries, this information will be relevant to VA and non-VA alike. J Am Geriatr Soc 67:1928-1933, 2019.

Auxiliary-assisted chemical ubiquitylation of NEMO and linear extension by HOIP.

The ubiquitylation of NF-κB essential modulator (NEMO) is part of the intracellular immune signalling pathway. Monoubiquitylated NEMO is required for exploring the mechanism of NEMO linear ubiquitylation by LUBAC (linear ubiquitin chain assembly complex), but is not accessible by biological techniques. Here we perform the chemical ubiquitylation of NEMO using a ligation auxiliary, which only requires a two-step synthesis, and is easily installed onto the lysine side-chain. Chemical ligation occurs directly on the lysine ε amine and remains efficient below pH 7. We show that ubiquitylated NEMO has similar affinity to linear diubiquitin chains as unmodified NEMO. The proximal ubiquitin of chemically synthesised NEMOCoZi-Ub is accepted as a substrate for linear extension by the (RING-Between-RING) RBR domain of HOIL-1-interacting protein (HOIP) alone. Our results indicate that NEMO linear ubiquitylation consists of two-steps, an initial priming event and a separate extension step requiring different LUBAC components.

Should Structured Exercise Be Promoted As a Model of Care? Dissemination of the Department of Veterans Affairs Gerofit Program.

Exercise provides a wide range of health-promoting benefits, but support is limited for clinical programs that use exercise as a means of health promotion. This stands in contrast to restorative or rehabilitative exercise, which is considered an essential medical service. We propose that there is a place for ongoing, structured wellness and health promotion programs, with exercise as the primary therapeutic focus. Such programs have long-lasting health benefits, are easily implementable, and are associated with high levels of participant satisfaction. We describe the dissemination and implementation of a long-standing exercise and health promotion program, Gerofit, for which significant gains in physical function that have been maintained over 5 years of follow-up, improvements in well-being, and a 10-year 25% survival benefit among program adherents have been documented. The program has been replicated at 6 Veterans Affairs Medical Centers. The pooled characteristics of enrolled participants (n = 691) demonstrate substantial baseline functional impairment (usual gait speed 1.05 ± 0.3 m/s, 8-foot up and go 8.7 ± 6.7 seconds, 30-second chair stands 10.7 ± 5.1, 6-minute walk distance 404.31 ± 141.9 m), highlighting the need for such programs. Change scores over baseline for 3, 6, and 12 months of follow-up are clinically and statistically significant (P < .05 all measures) and replicate findings from the parent program. Patient satisfaction ratings of high ranged from 88% to 94%. We describe the implementation process and present 1-year outcomes. We suggest that such programs be considered essential elements of healthcare systems.

National Structural Survey of Veterans Affairs Home-Based Primary Care Programs.

OBJECTIVES: To describe the current structural and practice characteristics of the Department of Veterans Affairs (VA) Home-Based Primary Care (HBPC) program. DESIGN: We designed a national survey and surveyed HBPC program directors on-line using REDCap. PARTICIPANTS: We received 236 surveys from 394 identified HBPC sites (60% response rate). MEASUREMENTS: HBPC site characteristics were quantified using closed-ended formats. RESULTS: HBPC program directors were most often registered nurses, and HBPC programs primarily served veterans with complex chronic illnesses that were at high risk of hospitalization and nursing home care. Primary care was delivered using interdisciplinary teams, with nurses, social workers, and registered dietitians as team members in more than 90% of the sites. Most often, nurse practitioners were the principal primary care providers (PCPs), typically working with nurse case managers. Nearly 60% of the sites reported dual PCPs involving VA and community-based physicians. Nearly all sites provided access to a core set of comprehensive services and programs (e.g., case management, supportive home health care). At the same time, there were variations according to site (e.g., size, location (urban, rural), use of non-VA hospitals, primary care models used). CONCLUSION: HBPC sites reflected the rationale and mission of HBPC by focusing on complex chronic illness of home-based veterans and providing comprehensive primary care using interdisciplinary teams. Our next series of studies will examine how HBPC site structural characteristics and care models are related to the processes and outcomes of care to determine whether there are best practice standards that define an optimal HBPC structure and care model or whether multiple approaches to HBPC better serve the needs of veterans.

FAN1 activity on asymmetric repair intermediates is mediated by an atypical monomeric virus-type replication-repair nuclease domain.

FAN1 is a structure-selective DNA repair nuclease with 5' flap endonuclease activity, involved in the repair of interstrand DNA crosslinks. It is the only eukaryotic protein with a virus-type replication-repair nuclease ("VRR-Nuc") "module" that commonly occurs as a standalone domain in many bacteria and viruses. Crystal structures of three representatives show that they structurally resemble Holliday junction resolvases (HJRs), are dimeric in solution, and are able to cleave symmetric four-way junctions. In contrast, FAN1 orthologs are monomeric and cleave 5' flap structures in vitro, but not Holliday junctions. Modeling of the VRR-Nuc domain of FAN1 reveals that it has an insertion, which packs against the dimerization interface observed in the structures of the viral/bacterial VRR-Nuc proteins. We propose that these additional structural elements in FAN1 prevent dimerization and bias specificity toward flap structures.

Migration and Remittances: Evidence from a Poor Province in China.

This paper examines patterns of remittances among migrants from Guizhou province of China. Our research is motivated by three lines of theoretical arguments, namely the new economics of migration, a translocal perspective linking remittances and development, and the culture of remittances. Taking individual, household, and village-level characteristics into account, we estimated multilevel logistic models of the decision to remit and multilevel models of the amount of remittances. Our results show that migrant remittance behaviour is responsive to family needs as well as household economic position in the village.. Migrants who come from entrepreneurial households are more likely to remit a large amount than other types of households. We find some evidence of "culture of remittances" in these villages. Consistent with our expectations, migrants who are from villages with higher amount of average remittances are likely to remit a larger amount than otherwise.

Structural and functional analysis of phosphothreonine-dependent FHA domain interactions.

FHA domains are well established as phospho-dependent binding modules mediating signal transduction in Ser/Thr kinase signaling networks in both eukaryotic and prokaryotic species. Although they are unique in binding exclusively to phosphothreonine, the basis for this discrimination over phosphoserine has remained elusive. Here, we attempt to dissect overall binding specificity at the molecular level. We first determined the optimal peptide sequence for Rv0020c FHA domain binding by oriented peptide library screening. This served as a basis for systematic mutagenic and binding analyses, allowing us to derive relative thermodynamic contributions of conserved protein and peptide residues to binding and specificity. Structures of phosphopeptide-bound and uncomplexed Rv0020c FHA domain then directed molecular dynamics simulations which show how the extraordinary discrimination in favor of phosphothreonine occurs through formation of additional hydrogen-bonding networks that are ultimately stabilized by van der Waals interactions of the phosphothreonine γ-methyl group with a conserved pocket on the FHA domain surface.

An intramolecular switch regulates phosphoindependent FHA domain interactions in Mycobacterium tuberculosis.

Forkhead-associated (FHA) domains have gained considerable prominence as ubiquitous phosphothreonine-dependent binding modules; however, their precise roles in serine and threonine kinase (STK) pathways and mechanisms of regulation remain unclear. From experiments with Rv1827, an FHA domain-containing protein from Mycobacterium tuberculosis, we derived a complete molecular description of an FHA-mediated STK signaling process. First, binding of the FHA domain to each of three metabolic enzyme complexes regulated their catalytic activities but did not require priming phosphorylation. However, phosphorylation of a threonine residue within a conserved amino-terminal motif of Rv1827 triggered its intramolecular association with the FHA domain of Rv1827, thus blocking its interactions with each of the three enzymes. The solution structure of this inactivated form and further mutagenic studies showed how a previously unidentified intramolecular phosphoswitch blocked the access of the target enzymes to a common FHA interaction surface and how this shared surface accommodated three functionally related, but structurally diverse, binding partners. Thus, our data reveal an unsuspected versatility in the FHA domain that allows for the transformation of multiple kinase inputs into various downstream regulatory signals.

Chk2 oligomerization studied by phosphopeptide ligation: implications for regulation and phosphodependent interactions.

Chk2/CHEK2/hCds1 is a modular serine-threonine kinase involved in transducing DNA damage signals. Phosphorylation by ataxia telangiectasia-mutated kinase (ATM) promotes Chk2 self-association, autophosphorylation, and activation. Here we use expressed protein ligation to generate a Chk2 N-terminal regulatory region encompassing a fork-head-associated (FHA) domain, a stoichiometrically phosphorylated Thr-68 motif and intervening linker. Hydrodynamic analysis reveals that Thr-68 phosphorylation stabilizes weak FHA-FHA interactions that occur in the unphosphorylated species to form a high affinity dimer. Although clearly a prerequisite for Chk2 activation in vivo, we show that dimerization modulates potential phosphodependent interactions with effector proteins and substrates through either the pThr-68 site, or the canonical FHA phosphobinding surface with which it is tightly associated. We further show that the dimer-occluded pThr-68 motif is released by intra-dimer autophosphorylation of the FHA domain at the highly conserved Ser-140 position, a major pThr contact in all FHA-phosphopeptide complex structures, revealing a mechanism of Chk2 dimer dissociation following kinase domain activation.