bluuebunny/crossref_metadata_2025 huggingface dataset:
Augmented Antitumor Activity for Novel Dual PI3K/BDR4 Inhibitors, SF2523 and SF1126 in Ewing Sarcoma
Authors: Goldin Amanda N. , Singh Alok , Joshi Shweta , Jamieson Christina , Durden Donald L.
Published: January 2021

Ewing sarcoma (ES) is the second most common pediatric bone cancer. Despite recent advances in the treatment, patients with metastatic tumors have dismal prognosis and hence novel therapies are urgently needed to combat this cancer. A recent study has shown that phosphoinositide-3 kinase (PI3K) inhibitors can synergistically increase sensitivity to bromodomain and extraterminal domain inhibitors in ES cells and therefore combined inhibition of PI3K and bromodomain and extraterminal domain bromodomain proteins might provide benefit in this cancer. Herein, we have investigated the efficacy of dual PI3K/BRD4 inhibitors, SF2523 and SF1126, for their antitumor activity in ES cell lines. The effect of SF1126 and SF2523 on cell viability and PI3K signaling was assessed on a panel of human ES cell lines. To evaluate the antitumor activity of SF1126, A673 cells were injected intrafemorally into RAG-2−/− mice and treated with 50 mg/kg SF1126 6 days per week, for 30 days. Both SF1126 and SF2523 decreased cell survival and inhibited phosphorylation of AKT in human ES cell lines. In vivo, SF1126 showed a significant reduction in tumor volume. These results suggest that dual PI3K/BRD4 inhibitor, SF1126, has antitumor activity in ES models.
Primary Primitive Neuroectodermal Tumor of the Spine With t(11;22): Report of 3 Cases and Review of Literature
Authors: Margol Ashley , Abongwa Chenue , Dhall Girish
Published: January 2021

Intradural extramedullary peripheral primitive neuroectodermal tumor (pPNET) with t(11;22) is a rare clinical finding in the pediatric population with few published cases in the literature. The authors report 3 cases of intradural primary pPNET and discuss the clinical presentation, treatment, and survival of the patients. Clinicians should be vigilant in considering pPNET in the differential diagnosis of extradural masses. The authors also compare the clinical course and outcome of therapy with primary PNET of the central nervous system and Ewing sarcoma family of tumors. In addition, this report highlights the risk for leptomeningeal dissemination at recurrence and discusses the importance of central nervous system-targeted therapy for durable disease control.
The Role of Expectations and Endogenous Opioids in Mindfulness-Based Relief of Experimentally Induced Acute Pain
Authors: Case Laura , Adler-Neal Adrienne L. , Wells Rebecca E. , Zeidan Fadel
Published: January 2021

Objective

Expectations contribute to cognitive pain modulation through opioidergically mediated descending inhibition. Mindfulness meditation reduces pain independent of endogenous opioids, engaging unique corticothalamocortical mechanisms. However, it remains unknown whether expectations for pain relief predict mindfulness-induced analgesia and if these expectations are modified by endogenous opioids.

Methods

In this secondary analysis of previously published work, 78 pain-free participants (mean age, 27 ± 7 years; 50% women) were randomized to a four-session mindfulness meditation or book listening regimen. Expectations for intervention-induced pain relief were assessed before and after each intervention. Pain ratings were examined after meditation or rest (control group) during noxious heat (49°C) and intravenous administration of saline placebo or the opioid antagonist naloxone (0.15 mg/kg bolus + 0.1 mg kg−1 h−1 infusion.

Results

Mindfulness significantly lowered pain during saline and naloxone infusion. Higher expected pain relief from mindfulness predicted lower pain intensity (r(40) = −0.41, p = .009). The relationship between meditation-related expectations and pain intensity reductions was exhibited during naloxone (r(20) = −0.76, p < .001) but not saline (r(20) = −0.22, p = .36). Expectations for book listening–based analgesia did not significantly predict pain changes during saline (r(20) = −0.37, p = .11) or naloxone (r(18) = 0.26, p = .30) in the control group.

Conclusions

These novel findings demonstrate a significant role for expectations in mindfulness-based pain relief. However, this role was minimal during saline and stronger during opioid blockade, despite similar pain reductions. This supports growing evidence that mindfulness engages multiple mechanisms to reduce pain, suggesting that mindfulness might be an effective pain-reducing technique even for individuals with low expectations for pain relief.
Understanding Engagement in and Affective Experiences During Physical Activity: The Role of Meditation Interventions
Authors: Don Brian P. , Van Cappellen Patty , Fredrickson Barbara L.
Published: January 2021

Objective

Meditation interventions promote an array of well-being outcomes. However, the way in which these interventions promote beneficial outcomes is less clear. Here, we expanded on prior work by examining the influence of mindfulness and loving-kindness meditation on a key health behavior: physical activity.

Methods

To test our hypotheses, we drew upon two randomized intervention studies. In the first study, 171 adults (73.0% female) received 6 weeks of training in either mindfulness meditation or loving-kindness meditation, or were assigned to a control condition. In the second study, 124 adults (60.0% female) were assigned to a 6-week mindfulness or loving-kindness meditation group.

Results

Study 1 demonstrated that individuals who received mindfulness training reported sustained levels of physical activity across the intervention period (Pre: mean [standard deviation], or M [SD] = 4.09 [2.07]; Post: M [SD] = 3.68 [2.00]; p = .054), whereas those in the control (Pre: M [SD] = 3.98 [2.25]; Post: M [SD] = 3.01 [2.07]; p < .001) and loving-kindness (Pre: M [SD] = 4.11 [2.26]; Post: M [SD] = 3.45 [1.96]; p < .001) conditions reported lower levels. Study 2 demonstrated those who received mindfulness training experienced increases in positive emotions during physical activity from preintervention to postintervention (Pre: M [SD] = 6.06 [2.51]; Post: M [SD] = 6.54 [2.43]; p = .001), whereas those trained in loving-kindness meditation experienced decreases in positive emotions during physical activity (Pre: M [SD] = 6.45 [2.35]; Post: M [SD] = 6.09 [2.46]; p = .040).

Conclusions

These results suggest mindfulness training (but not loving-kindness training) promotes sustained physical activity, and one plausible reason why this occurs is enhanced positive emotion during physical activity.
Diminished Age-Related Decline of the Amygdala in Long-Term Meditation Practitioners
Authors: Luders Eileen , Jain Felipe A. , Kurth Florian
Published: January 2021

Objective

A growing body of scientific evidence suggests that meditation may slow brain aging. The amygdala—a heterogenous brain region known to decrease in volume with increasing age—seems to be involved in meditation and affected by meditation. Thus, we hypothesized that the age-related decline of the amygdala is diminished in meditation practitioners.

Methods

We investigated whether correlations between age and gray matter volumes of the amygdala are significantly reduced in 50 long-term meditators compared with 50 sex- and age-matched healthy controls. Both the meditator and control groups included 44% women. The age of the participants ranged between 24 and 77 years, with mean (standard deviation) ages of 50.4 (±11.8) years in meditators and 51.4 (±12.8) years in controls. In addition to studying the amygdala as a whole, we investigated its centromedial, laterobasal, and superficial subregions using a well-validated approach combining imaging-based signal intensities and cytoarchitectonically defined probabilities.

Results

We detected significant group-by-age interactions for the whole amygdala and for its subregions. Follow-up analyses indicated negative age-related correlations in both meditators and controls (the older the participants, the smaller the volumes) but with significantly steeper aging trajectories in controls.

Conclusions

Altogether, these findings suggest that the age-related volume loss of the amygdala is less pronounced in long-term meditators. This effect was particularly evident for the laterobasal subregion, which has been functionally linked to aspects of self-focused reflection.
Investigating Hospital Supervision: A Case Study of Regulatory Inspectors’ Roles as Potential Co-creators of Resilience
Authors: Øyri Sina Furnes , Braut Geir Sverre , Macrae Carl , Wiig Siri
Published: January 2021

Objectives

The aim of this study was to explore if, and in what ways, there has been changes in the supervisory approach toward Norwegian hospitals due to the implementation of a new management and quality improvement regulation (Regulation on Management and Quality Improvement in the Healthcare Services, hereinafter referred to as “Quality Improvement Regulation”). Moreover, we aimed to understand how inspectors’ work promotes or hampers resilience potentials of adaptive capacity and learning in hospitals.

Methods

The study design is a case study of implementation and impact of the Quality Improvement Regulation. We performed a document analysis, and conducted and analyzed 3 focus groups and 2 individual interviews with regulatory inspectors, recruited from 3 county governor offices who are responsible for implementation and supervision of the Quality Improvement Regulation in Norwegian regions.

Results

Data analysis resulted in 5 themes. Informants described no substantial change in their approach owing to the Quality Improvement Regulation. Regardless, data pointed to a development in their practices and expectations. Although the Norwegian Board of Health Supervision, at the national level, occasionally provides guidance, supervision is adapted to specific contexts and inspectors balance trade-offs. Informants expressed concern about the impact of supervision on hospital performance. Benefits and disadvantage with positive feedback from inspectors were debated. Inspectors could nurture learning by improving their follow-up and add more hospital self-assessment.

Conclusions

A nondetailed regulatory framework such as the Quality Improvement Regulation provides hospitals with room to maneuver, and self-assessment might reduce resource demands. The impact of supervision is scarce with an unfulfilled potential to learn from supervision. The Government could contribute to a shift in focus by instructing the county governors to actively reflect on and communicate positive experiences from, and smart adaptations in, hospital practice.
The Development of a Surgical Oncology Center During the COVID-19 Pandemic
Authors: Moriarty Peter , Chang Justin , Kayani Babar , Roberts Lois , Bourke Neil , Dann Christopher , MacArthur Emma , Haddad Fares S.
Published: January 2021

Coronavirus disease 2019 (COVID-19) was declared a pandemic by the World Health Organization on March 11, 2020. By mid-March, London had emerged as the epicenter in the United Kingdom, accounting for 45% of the COVID-19–related mortality. A cancer COVID-19–free “cold hub,” through National Health Service partnership with independent institutions, was established to maintain a throughput of surgical oncology patients with an accessible triage system for oncologic specialties. The high population density, commuter volume, and rising COVID-19 incidence heightened the challenge of segregating a vulnerable population that was already at high risk for surgical morbidity and mortality.

The aim of this review is to report the experience of developing a multicenter COVID-19–free cold hub with the aim of providing safe surgery for surgical oncology patients. We discuss the timeline, structure, and infection control policy and suggest practical points that may guide other health care systems.

The surgical oncology hub treated 1542 patients between March 1 and July 1, 2020. There were no cases of COVID-19–related mortality in a 30-day follow-up. Key strategies for the restructuring of the cancer service included the following: (1) formation of an accessible referral pathway, (2) creation and structuring of cold hub hospitals, (3) development of protocols for infection control and preoperative testing, (4) rapid reorganization of services based on initial feedback, and (5) clear communication and leadership.

It has been shown that a surgical oncology cold hub with an accessible referral system and an effective system of preoperative screening system can minimize COVID-19 transmission, morbidity, and mortality, in a region with heavy disease prevalence. This structure represents a safe, ethical, and viable system that can be replicated in other health care systems.
Examining the Relationship Between Perceived Quality of Care and Actual Quality of Care as Measured by 30-Day Readmission Rates
Authors: Salinas Stanley R.
Published: January 2021

Objective:

To test the relationship between patient experience, as measured by the Hospital Consumer Assessment of Healthcare Providers and Systems (HCAHPS), and actual quality of care, as measured by 30-day readmission rates.

Data Sources:

Both HCAHPS data and outcome data reported to the Centers of Medicare & Medicaid Services (CMS).

Study Design:

This secondary, nationwide (N = 4060), hospital-level study focused only on acute care hospitals. HCAHPS question 22 “Would you recommend this hospital to your friends and family?” was used to determine level of overall satisfaction, and 30-day readmission rates, as reported to the CMS, was used as a proxy for actual quality of care.

Principal Findings:

A statistically significant relationship was found between patient experience and actual quality outcomes.

Conclusion:

The results of this study reinforce the inclusion of patient experience in Medicare's Value Based Purchasing program as a matter of good public policy.
Does the Proportion of Same-Day and 24-Hour Appointments Impact Patient Satisfaction?
Authors: Richter Jason P. , Downs Lynn , Beauvais Brad , Huynh Peter V. , Hamilton Jessica E. , Kim Forest , Weigel Fred
Published: January 2021

Background:

The relationship between open access and patient satisfaction is mixed. Our study is the first to assess the relationship between open access appointment scheduling and patient satisfaction in the Military Health System (MHS). It is also unique in that we examine both same-day and 24-hour access through a relationship with satisfaction.

Methods:

We conducted a panel time-series analysis with general estimating equations on the Army population of outpatient facilities (N = 32), with 32 364 957 total observations. Our primary independent variables were the proportion of a facility's appointments within 24 hours and same day from July 2013 to May 2015.

Results:

We identified that a higher proportion of same-day appointments is associated with increased patient satisfaction with the ability to see their provider when needed. We did not find the same result when examining access within 24 hours.

Conclusions:

Open access appointment scheduling appears to have a greater impact on patient satisfaction with timeliness of care if that appointment is made the same day the patient presents to the facility. Facilities should consider opening more of their schedule to accommodate same-day appointments. This can result in less costly primary care instead of emergency department usage.
Promoting Continuous Quality Improvement in the Alabama Child Health Improvement Alliance Through Q-Sort Methodology and Learning Collaboratives
Authors: Fifolt Matthew , Preskitt Julie , Rucks Andrew , Corvey Kathryn , Benton Elizabeth Cason
Published: January 2021

Objective:

Q-sort methodology is an underutilized tool for differentiating among multiple priority measures. The authors describe steps to identify, delimit, and sort potential health measures and use selected priority measures to establish an overall agenda for continuous quality improvement (CQI) activities within learning collaboratives.

Methods:

Through an iterative process, the authors vetted a list of potential child and adolescent health measures. Multiple stakeholders, including payers, direct care providers, and organizational representatives sorted and prioritized measures, using Q-methodology.

Results:

Q-methodology provided the Alabama Child Health Improvement Alliance (ACHIA) an objective and rigorous approach to system improvement. Selected priority measures were used to design learning collaboratives. An open dialogue among stakeholders about state health priorities spurred greater organizational buy-in for ACHIA and increased its credibility as a statewide provider of learning collaboratives.

Conclusions:

The integrated processes of Q-sort methodology, learning collaboratives, and CQI offer a practical yet innovative way to identify and prioritize state measures for child and adolescent health and establish a learning agenda for targeted quality improvement activities.
“Reuma-Check”
Authors: García Salinas Rodrigo , Ruta Santiago , Chichande Jessica Torres , Prado Einer Sanchez , Ruta Alvaro , Salvatori Facundo , Aguerre Dario , Magri Sebastián
Published: January 2021

Objectives

The aim of this study was to evaluate the performance of a comprehensive diagnosis program called “Reuma-check” for the diagnosis of axial spondyloarthritis (SpA) in patients with low back pain (LBP).

Methods

This is a cross-sectional study. Patients with LBP aged 18 years or older were preselected, and those with at least 1 SpA feature completed the circuit. They were referred after 2 strategies: education for orthopedists and a campaign on social networks. All patients underwent a clinical evaluation, laboratory testing, and imaging (including human leukocyte antigen B27 evaluation and magnetic resonance imaging). The diagnosis of axial SpA was established by an expert rheumatologist opinion. Time from onset of symptoms to “Reuma-check,” time from patient referral to admission of the checkup, and time from “Reuma-check” to diagnosis were evaluated.

Results

A total of 175 of 246 patients were included, most of them came from the social media campaign (55%). Seventy-five (43%) of 175 patients were diagnosed as axial SpA. The median time from referral (or self-referral) to access to the program was 1.3 months. The median time from symptoms onset to access to the program was 31.7 months, and the median time from the performance of “Reuma-check” to final diagnosis was 2 weeks. Features associated with a diagnosis of axial SpA were as follows: inflammatory LBP (odds ratio [OR], 6.64; 95% confidence interval [CI], 1.6–28), clinical enthesopathy (OR, 4.56; 95% CI, 1.1–18.4), positive human leukocyte antigen B27 (OR, 23.02; 95% CI, 3.5–58), and positive magnetic resonance imaging (OR, 14.34; 95% CI, 3.5–58).

Conclusions

“Reuma-check” allowed a high frequency of axial SpA diagnosis and improved access to rapid diagnosis, shortening the time from referral to diagnosis with a shorter acquisition time for the ancillary studies. Patients with a final diagnosis of axial SpA presented distinctive features.
A novel partial resuscitative endovascular balloon aortic occlusion device that can be deployed in zone 1 for more than 2 hours with minimal provider titration
Authors: Kemp Michael T. , Wakam Glenn K. , Williams Aaron M. , Biesterveld Ben E. , O’Connell Rachel L. , Vercruysse Claire A. , Chtraklin Kiril , Russo Rachel M. , Alam Hasan B.
Published: January 2021

BACKGROUND

Hemorrhage is a leading cause of mortality in trauma. Resuscitative endovascular balloon occlusion of the aorta (REBOA) can control hemorrhage, but distal ischemia, subsequent reperfusion injury, and the need for frequent balloon titration remain problems. Improved device design can allow for partial REBOA (pREBOA) that may provide hemorrhage control while also perfusing distally without need for significant provider titration.

METHODS

Female Yorkshire swine (N = 10) were subjected to 40% hemorrhagic shock for 1 hour (mean arterial pressure [MAP], 28–32 mm Hg). Animals were then randomized to either complete aortic occlusion (ER-REBOA) or partial occlusion (novel pREBOA-PRO) without frequent provider titration or distal MAP targets. Detection of a trace distal waveform determined partial occlusion in the pREBOA-PRO arm. After 2 hours of zone 1 occlusion, the hemorrhaged whole blood was returned. After 50% autotransfusion, the balloon was deflated over a 10-minute period. Following transfusion, the animals were survived for 2 hours while receiving resuscitation based on objective targets: lactated Ringer’s fluid boluses (goal central venous pressure, ≥ 6 mm Hg), a norepinephrine infusion (goal MAP, 55–60 mm Hg), and acid-base correction (goal pH, >7.2). Hemodynamic variables, arterial lactate, lactate dehydrogenase, aspartate aminotransferase, and creatinine levels were measured.

RESULTS

All animals survived throughout the experiment, with similar increase in proximal MAPs in both groups. Animals that underwent partial occlusion had slightly higher distal MAPs. At the end of the experiment, the partial occlusion group had lower end levels of serum lactate (p = 0.006), lactate dehydrogenase (p = 0.0004) and aspartate aminotransferase (p = 0.004). Animals that underwent partial occlusion required less norepinephrine (p = 0.002), less bicarbonate administration (p = 0.006), and less fluid resuscitation (p = 0.042).

CONCLUSION

Improved design for pREBOA can decrease the degree of distal ischemia and reperfusion injury compared with complete aortic occlusion, while providing a similar increase in proximal MAPs. This can allow pREBOA zone-1 deployment for longer periods without the need for significant balloon titration.
Causes of early mortality in pediatric trauma patients
Authors: Theodorou Christina M. , Galganski Laura A. , Jurkovich Gregory J. , Farmer Diana L. , Hirose Shinjiro , Stephenson Jacob T. , Trappey A. Francois
Published: January 2021

BACKGROUND

Trauma is the leading cause of death in children, and most deaths occur within 24 hours of injury. A better understanding of the causes of death in the immediate period of hospital care is needed.

METHODS

Trauma admissions younger than 18 years from 2009 to 2019 at a Level I pediatric trauma center were reviewed for deaths (n = 7,145). Patients were stratified into ages 0–6, 7–12, and 13–17 years old. The primary outcome was cause of death, with early death defined as less than 24 hours after trauma center arrival.

RESULTS

There were 134 (2%) deaths with a median age of 7 years. The median time from arrival to death was 14.4 hours (interquartile range, 0.5–87.8 hours). Half (54%) occurred within 24 hours. However, most patients who survived initial resuscitation in the emergency department died longer than 24 hours after arrival (69%). Traumatic brain injury was the most common cause of death (66%), followed by anoxia (9.7%) and hemorrhage (8%). Deaths from hemorrhage were most often in patients sustaining gunshot wounds (73% vs. 11% of all other deaths, p < 0.0001), more likely to occur early (100% vs. 50% of all other deaths, p = 0.0009), and all died within 6 hours of arrival. Death from hemorrhage was more common in adolescents (21.4% of children aged 13–17 vs. 6.3% of children aged 0–6, and 0% of children aged 7–12 p = 0.03). The highest case fatality rates were seen in hangings (38.5%) and gunshot wounds (9.6%).

CONCLUSION

Half of pediatric trauma deaths occurred within 24 hours. Death from hemorrhage was rare, but all occurred within 6 hours of arrival. This is a critical time for interventions for bleeding control to prevent death from hemorrhage in children. Analysis of these deaths can focus efforts on the urgent need for development of new hemorrhage control adjuncts in children.

LEVEL OF EVIDENCE

Epidemiological study, level IV.
The effect of the Affordable Care Act on rates on inpatient rehabilitation hospital admission in a mature trauma system
Authors: Winter Mike , Vernon Tawnya , Morgan Madison , Bresz Kellie , Cook Alan , Bradburn Eric , Rogers Frederick
Published: January 2021

BACKGROUND

The beneficial effects of acute rehabilitation for trauma patients are well documented but can be limited because of insurance coverage. The Patient Protection and Affordable Care Act (ACA) went into effect on March 23, 2010. The ACA allowed patients who previously did not have insurance to be fully incorporated into the health system. We sought to analyze the likelihood of discharge to rehab for trauma patients before and after the implementation of the ACA. We hypothesized that there would be a higher rate of inpatient rehabilitation hospital (IRH) admission after the ACA was put into effect.

METHODS

The Pennsylvania Trauma Outcome Study (PTOS) database was retrospectively queried from 2003 to 2017 for all trauma patients admitted to accredited trauma centers in Pennsylvania, who also had a functional status at discharge (FSD). Admission to an IRH was determined using discharge destination. Two categories were created to represent periods before and after ACA was implemented, 2003 to 2009 (pre-ACA) and 2010–2017 (post-ACA). A multilevel mixed-effects logistic regression model controlling for demographics, injury severity, and FSD assessed the adjusted impact of ACA implementation on IRH admissions.

RESULTS

From the Pennsylvania Trauma Outcome Study query, 341,252 patients had FSD scores and of these patients, 47,522 (13.9%) were admitted to IRH. Patients who were severely injured were more likely to be admitted to IRH. Compared with FSD scores signifying complete independence at discharge, those with lower FSD had significantly increased odds of IRH admission. The odds of IRH admission post-ACA implementation significantly increased when compared with pre-ACA years (adjusted odds ratio, 1.14; 95% confidence interval, 1.12–1.17; p < 0.001; area under the receiver operating curve, 0.818).

CONCLUSION

The implementation of the ACA significantly increased the likelihood of discharge to IRH for trauma patients.

LEVEL OF EVIDENCE

Care management, level III
Optimizing Function for the Older Adult With Degenerative Scoliosis Through Pilates Environment Intervention
Authors: Martin Suzanne
Published: January 2021

However, the degenerative spinal changes associated with an adult with scoliosis often confound with the many typical physical degenerative issues associated with aging. Exercise is showing promise in mitigating general age-related physical decline. However, adult degenerative scoliosis and its neurological and biomechanical impact on the rib cage, pelvis, head, and limbs demonstrate a need for a multiplanar, integrative whole-body system of intervention. The Pilates Method is one whole-body approach that is showing success with adult degenerative scoliosis symptom management and with keeping our general adult population active through the aging process.
Cerebral Amyloid Angiopathy-related Inflammation Presenting With a Cystic Lesion in Young-onset Alzheimer Disease
Authors: Ringman John M. , Joe Elizabeth , Sheikh-Bahaei Nasim , Miller Carol , Vinters Harry V. , Guzman Samuel , Chui Helena C.
Published: January 2021

We describe a patient with cerebral amyloid angiopathy-related inflammation (CAA-ri) presenting as Alzheimer disease (AD) with a mass lesion with symptom onset at age 59. He was found to have a nonenhancing lesion in the right temporal lobe on magnetic resonance imaging without evidence of hemorrhage. He underwent a biopsy which showed amyloid beta in blood vessel walls and a perivascular inflammatory infiltrate consistent with CAA-ri. Neurofibrillary tangles were present and a flortaucipir positron emission tomography showed bilateral signal highest in the lateral temporal and parietal cortices. A lumbar puncture showed tau, p-tau, and amyloid beta levels consistent with AD without evidence of inflammation. He was homozygous for the APOE ε4 allele. He died at age 67. A focus of CAA-ri can be present in the context of AD with a mass lesion and without evidence of hemorrhage, significant ischemic changes, or overt inflammation on cerebrospinal fluid examination.
Characterisation of the RNA-interference pathway as a Tool for Genetics in the Nascent Phototrophic Endosymbiosis,Paramecium bursaria
Authors: Jenkins Benjamin H. , Maguire Finlay , Leonard Guy , Eaton Joshua D. , West Steven , Housden Benjamin E. , Milner David S. , Richards Thomas A.
Published: January 2021

Endosymbiosis was fundamental for the evolution of eukaryotic complexity. Endosymbiotic interactions can be dissected through forward and reverse-genetic experiments, such as RNA-interference (RNAi). However, distinguishing small (s)RNA pathways in a eukaryote-eukaryote endosymbiotic interaction is challenging. Here, we investigate the repertoire of RNAi pathway protein-encoding genes in the model nascent endosymbiotic system,Paramecium bursaria–Chlorellaspp. Using comparative genomics and transcriptomics supported by phylogentics, we identify essential proteome components of the small interfering (si)RNA, scan (scn)RNA, and internal eliminated sequence (ies)RNA pathways. Our analyses reveal that copies of these components have been retained throughout successive whole genome duplication (WGD) events in theParameciumclade. We then validate feeding-induced siRNA-based RNAi inP. bursariavia knock-down of the splicing factor,u2af1, which we show to be crucial to host growth. Finally, using simultaneous knock-down paradox controls to rescue the effectu2af1knock-down, we demonstrate that feeding-induced RNAi inP. bursariais dependent upon a core pathway of host-encodedDcr1,PiwiandPds1components. Our experiments confirm the presence of a functional, host-derived RNAi pathway inP. bursariathat generates 23-nt siRNA, validating use of theP. bursaria-Chlorellaspp. system to investigate the genetic basis of a nascent endosymbiosis.
A two-color haploid genetic screen identifies novel host factors involved in HIV latency
Authors: Röling Michael D , Mollapour Sisakht Mahsa , Ne Enrico , Moulos Panagiotis , Stoszko Mateusz , De Crignis Elisa , Bodmer Helen , Kan Tsung Wai , Akbarzadeh Maryam , Harokopos Vaggelis , Hatzis Pantelis , Palstra Robert-Jan , Mahmoudi Tokameh
Published: January 2021

To identify novel host factors as putative targets to reverse HIV latency, we performed an insertional mutagenesis genetic screen in a latently HIV-1-infected pseudo-haploid KBM7 cell line (Hap-Lat). Following mutagenesis, insertions were mapped to the genome and bioinformatic analysis resulted in the identification of 69 candidate host genes involved in maintaining HIV-1 latency. A select set of candidate genes was functionally validated using shRNA mediated depletion in latent HIV-1 infected J-Lat A2 and 11.1 T cell lines. We confirmed ADK, CHD9, CMSS1, EVI2B, EXOSC8, FAM19A, GRIK5, IRF2BP2, NF1, and USP15 as novel host factors involved in the maintenance of HIV latency. Chromatin immunoprecipitation assays indicated that CHD9, a Chromodomain Helicase DNA-binding protein, maintains HIV latency via direct association with the HIV 5’LTR, and its depletion results in increased histone acetylation at the HIV-1 promoter, concomitant with HIV-1 latency reversal. FDA-approved inhibitors 5-Iodotubercidin, Trametinib, and Topiramate, targeting ADK, NF1, and GRIK5, respectively were characterized for their latency reversal potential. While 5-Iodotubercidin exhibited significant cytotoxicity in both J-Lat and primary CD4+ T cells, Trametinib reversed latency in J-Lat cells but not in latently HIV-1-infected primary CD4+ T cells. Crucially, Topiramate reversed latency in cell-line models and latently infected primary CD4+ T cells, without inducing T cell activation or significant toxicity. Thus, using an adaptation of a haploid forward genetic screen, we identified novel and druggable host factors contributing to HIV-1 latency.

Importance

A reservoir of latent HIV-1-infected cells persists in the presence of combination antiretroviral therapy (cART), representing a major obstacle for viral eradication. Reactivation of the latent HIV-1 provirus is part of curative strategies which aim to promote clearance of the infected cells. Using a two-color haploid screen, we identified 69 candidate genes as latency maintaining host factors and functionally validated a subset of 10 of those in additional T-cell based cell line models of HIV-1 latency. We further demonstrated that CHD9 is associated with HIV-1’s promoter, the 5’LTR while this association is lost upon reactivation. Additionally, we characterized the latency reversal potential of FDA compounds targeting ADK, NF1, and GRIK5 and identify the GRIK5 inhibitor Topiramate as a viable latency reversal agent with clinical potential.
Filamentation inCandida albicansis modulated by adaptive translation of farnesol signalling genes
Authors: Oliveira Carla , Guimarães Ana Rita , Correia Inês , Sousa Inês , Poim Ana , Rocha Sílvia M. , Moura Gabriela , Santos Manuel A. S. , Bezerra Ana Rita
Published: January 2021

The complex biology of the human pathogenCandida albicansis reflected in its remarkable ability to proliferate in numerous body sites, adapt to drastic changes in the environment, form various types of colonies and grow in yeast, pseudo-hyphal and hyphal forms. Much has been learnt in recent years about the relevance of this phenotypic plasticity, but the mechanisms that support it are still not fully understood. We have demonstrated that atypical translation of the CUG codon is a source of unexpected morphological diversity. The CUG codon is translated as both leucine (Leu) (~3%) and serine (Ser) (~97%) in normal growth conditions, but Ser/Leu levels change in response to stress. Remarkably, recombinantC. albicansstrains incorporating between 20% and 99% of Leu at CUG sites display a diverse array of phenotypes and produce colonies of variable morphology containing a mixture of yeast, pseudohyphal and hyphal cells. In this work we investigate the role of the CUG codon in the yeast-hypha transition. Our data show that increasing incorporation levels of Leu at CUG sites trigger hyphal initiation under non-inducing conditions by reducing farnesol production, and increasing the degradation of the Nrg1 hyphal repressor. We propose that dual CUG Ser/Leu translation triggers filamentation via the Nrg1 pathway.

Importance

The unique translation of the CUG codon as both Ser (~97%) and Leu (~3%) plays a key role in the production of high genomic and phenotypic diversity inC. albicans. The molecular mechanisms that support such diversity are poorly understood. Here, we show that increased Leu incorporation at CUG sites induce hyphae formation in media whereC. albicansnormally grows in the yeast form. The data show that increasing Leu at CUG sites triggers the degradation of the hyphal repressor Nrg1, allowing for full expression of hyphal genes. Since filamentation is important for invasion of host tissues, this work shows how the atypical translation of a single codon may play a critical role in the virulence of all fungi of the CTG clade.
iGEM comes of age: trends in its research output
Authors: Jainarayanan Ashwin K. , Galanis Anastasios , Nakara Amatullah Mustafa , Kundlatsch Guilherme E. , Rubio-Sánchez Roger
Published: January 2021

The international Genetically Engineered Machine (iGEM) is an educational benchmark in synthetic biology. Eighteen years after its inception, it has also catalysed the infusion of synthetic biology with interdisciplinary fundamental and translational research as well as with inspired young scientists. Here, we communicate a quantitative analysis of compiled published work associated to iGEM projects, highlighting trends in their dissemination and versatility. As iGEM comes of age, we anticipate it will continue to revolutionise, alongside SynBio, several disciplines of science and industries through the development of synthetic biological systems towards a sustainable future.
Independent control of mean and noise by convolution of gene expression distributions
Authors: Gerhardt Karl P. , Rao Satyajit D. , Olson Evan J. , Igoshin Oleg A. , Tabor Jeffrey J.
Published: January 2021

Gene expression noise can reduce cellular fitness or facilitate processes such as alternative metabolism, antibiotic resistance, and differentiation. Unfortunately, efforts to study the impacts of noise have been hampered by a scaling relationship between noise and expression level from a single promoter. Here, we use theory to demonstrate that mean and noise can be controlled independently by expressing two copies of a gene from separate inducible promoters in the same cell. We engineer low and high noise inducible promoters to validate this result inEscherichia coli, and develop a model that predicts the experimental distributions. Finally, we use our method to reveal that the response of a promoter to a repressor is less sensitive with higher repressor noise and explain this result using a law from probability theory. Our approach can be applied to investigate the effects of noise on diverse biological pathways or program cellular heterogeneity for synthetic biology applications.
Prophage-encoded hotspots of bacterial immune systems
Authors: Rousset François , Dowding Julien , Bernheim Aude , Rocha Eduardo P.C. , Bikard David
Published: January 2021

The arms race between bacteria and phages led to the emergence of a variety of genetic systems used by bacteria to defend against viral infection, some of which were repurposed as powerful biotechnological tools. While numerous defense systems have been identified in genomic regions termed defense islands, it is believed that many more remain to be discovered. Here, we show that P2- like prophages and their P4-like satellites have genomic hotspots that represent a significant source of novel anti-phage systems. We validate the defense activity of 14 systems spanning various protein domains and describe PARIS, an abortive infection system triggered by a phage-encoded anti-restriction protein. Immunity hotspots are present across prophages of distant bacterial species, highlighting their biological importance in the competition between bacteria and phages.
A new method for determining ribosomal DNA copy number shows differences betweenSaccharomyces cerevisiaepopulations
Authors: Sharma Diksha , Hermann-Le Denmat Sylvie , Matzke Nicholas J. , Hannan Katherine , Hannan Ross D. , O’Sullivan Justin M. , Ganley Austen R. D.
Published: January 2021

Ribosomal DNA genes (rDNA) encode the major ribosomal RNAs (rRNA) and in eukaryotic genomes are typically present as one or more arrays of tandem repeats. Species have characteristic rDNA copy numbers, ranging from tens to thousands of copies, with the number thought to be redundant for rRNA production. However, the tandem rDNA repeats are prone to recombination-mediated changes in copy number, resulting in substantial intra-species copy number variation. There is growing evidence that these copy number differences can have phenotypic consequences. However, we lack a comprehensive understanding of what determines rDNA copy number, how it evolves, and what the consequences are, in part because of difficulties in quantifying copy number. Here, we developed a genomic sequence read approach that estimates rDNA copy number from the modal coverage of the rDNA and whole genome to help overcome limitations in quantifying copy number with existing mean coverage-based approaches. We validated our method using strains of the yeastSaccharomyces cerevisiaewith previously-determined rDNA copy numbers, and then applied our pipeline to investigate rDNA copy number in a global sample of 788 yeast isolates. We found that wild yeast have a mean copy number of 92, consistent with what is reported for other fungi but much lower than in laboratory strains. We also show that different populations have different rDNA copy numbers. These differences can partially be explained by phylogeny, but other factors such as environment are also likely to contribute to population differences in copy number. Our results demonstrate the utility of the modal coverage method, and highlight the high level of rDNA copy number variation within and between populations.

Author summary

The ribosomal RNA gene repeats (rDNA) form large tandem repeat arrays in most eukaryote genomes. Their tandem arrangement makes the rDNA prone to copy number variation, and there is increasing evidence that this copy number variation has phenotypic consequences. However, difficulties in measuring rDNA copy number hamper investigation into rDNA copy number dynamics and their significance. Here we developed a novel bioinformatics method for measuring rDNA copy number from whole genome sequence data that is based on the modal sequence read coverage. We established parameters for optimal performance of the method and validated it using yeast strains of known rDNA copy numbers. We then applied the method to a dataset of almost 800 global yeast isolates and demonstrate that yeast populations have different rDNA copy numbers that partially correlate with phylogeny. Our work provides a simple and accurate method for determining rDNA copy number that leverages the growing number of whole genome datasets, and highlights the dynamic nature of rDNA copy number.
The fitness costs and benefits of trisomy of eachCandida albicanschromosome
Authors: Yang Feng , Jiang Yuan-ying , Cao Yong-bing , Berman Judith
Published: January 2021

Candida albicansis a prevalent human fungal pathogen. Rapid genomic change, due to aneuploidy, is a common mechanism that facilitates survival from multiple types of stresses including the few classes of available antifungal drugs. The stress survival of aneuploids occurs despite the fitness costs attributed to most aneuploids growing under idealized lab conditions. Systematic study of the aneuploid state inC. albicanshas been hindered by the lack of a comprehensive collection of aneuploid strains. Here, we describe a collection of diploidC. albicansaneuploid strains, each carrying one extra copy of each chromosome, all from the same genetic background. We tested the fitness of this collection under several physiological conditions including shifts in pH, low glucose, oxidative stress, temperature, high osmolarity, membrane stress and cell wall stress. We found that, most aneuploids, under most conditions, were less fit than their euploid parent, yet there were specific conditions under which specific aneuploid isolates provided a fitness benefit relative to the euploid parent strain. Importantly, this fitness benefit was attributable to the change in the copy number of specific chromosomes. Thus,C. albicanscan tolerate aneuploidy of each chromosome and some aneuploids confer improved growth under conditions that the yeast encounters in its host niches.
Structure of an Hsp90-immunophilin complex reveals cochaperone recognition of the client-maturation state
Authors: Lee Kanghyun , Thwin Aye C. , Tse Eric , Gates Stephanie N. , Southworth Daniel R.
Published: January 2021

The Hsp90 chaperone promotes the folding and activation of hundreds of client proteins in the cell through an ATP-dependent conformational cycle guided by distinct cochaperone regulators. The FKBP51 immunophilin binds Hsp90 with its tetratricopeptide repeat (TPR) domain and catalyzes peptidyl-prolyl isomerase (PPIase) activity during the folding of kinases, nuclear receptors and tau. Here we have determined the cryo-EM structure of the human Hsp90:FKBP51:p23 complex to 3.3 Å that, together with mutagenesis and crosslinking analysis, reveals the basis for cochaperone binding to Hsp90 during client maturation. A helix extension in the TPR functions as a key recognition element, interacting across the Hsp90 C-terminal dimer interface presented in the closed, ATP conformation. The PPIase domain is positioned along the middle domain, adjacent Hsp90 client binding sites, while a single p23 makes stabilizing interactions with the N-terminal dimer. With this architecture, FKBP51 could thereby act on specific client residues presented during Hsp90-catalyzed remodeling.
Stress fiber strain recognition by the LIM protein testin is cryptic and mediated by RhoA
Authors: Sala Stefano , Oakes Patrick W.
Published: January 2021

The actin cytoskeleton is a key regulator of mechanical processes in cells. The family of LIM domain proteins have recently emerged as important mechanoresponsive cytoskeletal elements capable of sensing strain in the actin cytoskeleton. The mechanisms regulating this mechanosensitive behavior, however, remain poorly understood. Here we show that the LIM domain protein testin is peculiar in that despite the full-length protein primarily appearing diffuse in the cytoplasm, the C-terminal LIM domains alone recognize focal adhesions and strained actin while the N-terminal domains alone recognize stress fibers. Phosphorylation mutations in the dimerization regions of testin, however, reveal its mechanosensitivity and cause it to relocate to focal adhesions and sites of strain in the actin cytoskeleton. Finally, we demonstrate activated RhoA causes testin to adorn stress fibers and become mechanosensitive. Together, our data show that testin’s mechanoresponse is regulated in cells and provide new insights into LIM domain protein recognition of the actin cytoskeleton mechanical state.
Different resource allocation in aBacillus subtilispopulation displaying bimodal motility
Authors: Syvertsson Simon , Wang Biwen , Staal Jojet , Gao Yongqiang , Kort Remco , Hamoen Leendert W.
Published: January 2021

To cope with sudden changes in their environment, bacteria can use a bet-hedging strategy by dividing the population into cells with different properties. This so-called bimodal or bistable cellular differentiation is generally controlled by positive feedback regulation of transcriptional activators. Due to the continuous increase in cell volume, it is difficult for these activators to reach an activation threshold concentration when cells are growing exponentially. This is one reason why bimodal differentiation is primarily observed from the onset of the stationary phase when exponential growth ceases. An exception is the bimodal induction of motility inBacillus subtilis, which occurs early during exponential growth. Several mechanisms have been put forward to explain this, including double negative-feedback regulation and the stability of the mRNA molecules involved. In this study, we used fluorescence-assisted cell sorting to compare the transcriptome of motile and non-motile cells and noted that expression of ribosomal genes is lower in motile cells. This was confirmed using an unstable GFP reporter fused to the strong ribosomalrpsDpromoter. We propose that the reduction in ribosomal gene expression in motile cells is the result of a diversion of cellular resources to the synthesis of the chemotaxis and motility systems. In agreement, single-cell microscopic analysis showed that motile cells are slightly shorter than non-motile cells, an indication of slower growth. We speculate that this growth rate reduction can contribute to the bimodal induction of motility during exponential growth.

IMPORTANCE

To cope with sudden environmental changes, bacteria can use a bet-hedging strategy and generate different types of cells within a population, so called bimodal differentiation. For example, aBacillus subtilisculture can contain both motile and non-motile cells. In this study we compared the gene expression between motile and non-motile cells. It appeared that motile cells express less ribosomes. To confirm this, we constructed a ribosomal promoter fusion that enabled us to measure expression of this promoter in individual cells. This reporter fusion confirmed our initial finding. The re-allocation of cellular resources from ribosome synthesis towards synthesis of the motility apparatus results in a reduction in growth. Interestingly, this growth reduction has been shown to stimulate bimodal differentiation.