Selection of a promiscuous minimalist cAMP phosphodiesterase from a library of de novo designed proteins.

The ability of unevolved amino acid sequences to become biological catalysts was key to the emergence of life on Earth. However, billions of years of evolution separate complex modern enzymes from their simpler early ancestors. To probe how unevolved sequences can develop new functions, we use ultrahigh-throughput droplet microfluidics to screen for phosphoesterase activity amidst a library of more than one million sequences based on a de novo designed 4-helix bundle. Characterization of hits revealed that acquisition of function involved a large jump in sequence space enriching for truncations that removed >40% of the protein chain. Biophysical characterization of a catalytically active truncated protein revealed that it dimerizes into an α-helical structure, with the gain of function accompanied by increased structural dynamics. The identified phosphodiesterase is a manganese-dependent metalloenzyme that hydrolyses a range of phosphodiesters. It is most active towards cyclic AMP, with a rate acceleration of ~109 and a catalytic proficiency of >1014 M-1, comparable to larger enzymes shaped by billions of years of evolution.

Engagement with Adolescent Substance Prevention Messages: Impact and Dissemination of REAL Media.

This article reviews the development and testing of a youth substance prevention program, REAL media. The contributions of this body of research include theoretical development, measurement, and dissemination of an efficacious independently evaluated program. Special attention is given to the impact of the program through collaborations with multiple community groups and multiple phases of development and testing.

Present-day thermal and water activity environment of the Mars Sample Return collection.

The Mars Sample Return mission intends to retrieve a sealed collection of rocks, regolith, and atmosphere sampled from Jezero Crater, Mars, by the NASA Perseverance rover mission. For all life-related research, it is necessary to evaluate water availability in the samples and on Mars. Within the first Martian year, Perseverance has acquired an estimated total mass of 355 g of rocks and regolith, and 38 µmoles of Martian atmospheric gas. Using in-situ observations acquired by the Perseverance rover, we show that the present-day environmental conditions at Jezero allow for the hydration of sulfates, chlorides, and perchlorates and the occasional formation of frost as well as a diurnal atmospheric-surface water exchange of 0.5-10 g water per m2 (assuming a well-mixed atmosphere). At night, when the temperature drops below 190 K, the surface water activity can exceed 0.5, the lowest limit for cell reproduction. During the day, when the temperature is above the cell replication limit of 245 K, water activity is less than 0.02. The environmental conditions at the surface of Jezero Crater, where these samples were acquired, are incompatible with the cell replication limits currently known on Earth.

Crystal structure and activity of a de novo enzyme, ferric enterobactin esterase Syn-F4.

Producing novel enzymes that are catalytically active in vitro and biologically functional in vivo is a key goal of synthetic biology. Previously, we reported Syn-F4, the first de novo protein that meets both criteria. Syn-F4 hydrolyzed the siderophore ferric enterobactin, and expression of Syn-F4 allowed an inviable strain of Escherichia coli (Δfes) to grow in iron-limited medium. Here, we describe the crystal structure of Syn-F4. Syn-F4 forms a dimeric 4-helix bundle. Each monomer comprises two long α-helices, and the loops of the Syn-F4 dimer are on the same end of the bundle (syn topology). Interestingly, there is a penetrated hole in the central region of the Syn-F4 structure. Extensive mutagenesis experiments in a previous study showed that five residues (Glu26, His74, Arg77, Lys78, and Arg85) were essential for enzymatic activity in vivo. All these residues are located around the hole in the central region of the Syn-F4 structure, suggesting a putative active site with a catalytic dyad (Glu26-His74). The complete inactivity of purified proteins with mutations at the five residues supports the putative active site and reaction mechanism. Molecular dynamics and docking simulations of the ferric enterobactin siderophore binding to the Syn-F4 structure demonstrate the dynamic property of the putative active site. The structure and active site of Syn-F4 are completely different from native enterobactin esterase enzymes, thereby demonstrating that proteins designed de novo can provide life-sustaining catalytic activities using structures and mechanisms dramatically different from those that arose in nature.

Developing the Opioid Rapid Response System™ for Lay Citizen Response to the Opioid Overdose Crisis: a Randomized Controlled Trial.

Emergency responders face challenges in arriving timely to administer naloxone in opioid overdoses. Therefore, interest in having lay citizens administer naloxone nasal spray has emerged. These citizens, however, must be recruited and trained, and be in proximity to the overdose. This study aimed to develop the Opioid Rapid Response System (ORRS)tm to meet this need by developing a system to recruit and train citizen responders and evaluate outcomes in a randomized clinical trial. ORRS recruitment messages and training platform were developed iteratively and then outcomes for each were evaluated in a randomized, unblinded two-arm waitlist-controlled trial. ORRS was field tested in 5 Indiana counties, recruiting adult citizen responders (age 18 or older) who did not self-identity as a certified first responder. Participants were recruited using either personal or communal messages and then randomly assigned to online naloxone training and waitlisted-control conditions. Pre- and post-surveys were administered online to measure the exposure to recruitment messages and training effects on knowledge of opioid overdose, confidence responding, concerns about responding, and intent to respond. Of the 220 randomized participants (114 training, 106 waitlisted-control), 140 were analyzed (59 training, 81 waitlisted-control). Recruited participants more frequently identified with communal appeal than with the personal appeal (chi-square = 53.5; p < 0.0001). Between-group differences for intervention effects were significant for knowledge of overdose signs (Cohen's d = 1.17), knowledge of overdose management (d = 1.72), self-efficacy (d = 1.39), and concerns (d = 1.31), but not for intent (d = 0.17), which suffered from a ceiling effect. ORRS provides stronger support for efficacy than that reported for other training interventions and the digital modality eases rapid dissemination.Trial Registration: NCT04589676.

Does Narrative Quality Matter During Implementation of a School-Based Prevention Intervention? A Test of Narrative Engagement Theory.

Narratives play a powerful role in sharing meaning and making sense of experiences. Specifically, health narratives convey storylines, characters, and messages about health-related behaviors and provide audiences with models for healthy behaviors, prompting audiences' health-related reflections and decision-making. Narrative engagement theory (NET) explains how personal narratives can be integrated into interventions to promote health. This study utilizes NET to test direct and indirect effects of teachers' narrative quality on adolescent outcomes during a school-based substance use prevention intervention that includes narrative pedagogy and an implementation strategy. Observational coding of teacher narratives in video-recorded lessons along with self-report student surveys (N = 1,683) were subjected to path analysis. Findings showed significant direct effects of narrative quality on student engagement, norms (i.e. personal, best-friend injunctive, and descriptive norms), and substance use behavior. The analysis also yielded support for indirect effects of narrative quality on adolescent substance use behavior via student engagement, personal norms, and descriptive norms. Findings highlight important issues related to teacher-student interaction during implementation and contributes implications for adolescent substance use prevention research.

Digital HPV education to increase vaccine uptake among low income women.

Objective: The objective of this formative study was to gather women's perspectives in the design and communication modalities of a health kiosk set within a Planned Parenthood setting to promote patient education about the Human papillomavirus (HPV) and to motivate uptake of the HPV vaccine. Methods: Twenty-four women aged 18-35 participated in in-depth one-on-one interviews at a Planned Parenthood health center, which were analyzed in code-associated categories using NVivo11 Pro. Results: Most women showed receptivity to using an on-site health kiosk, as well as QR codes linked to text messages, to receive HPV-related health information outside of the clinic setting and reminders. Participants provided suggestions for kiosk design and communication modalities. Conclusions: Among low-income women we interviewed at Planned Parenthood, increasing HPV vaccination rates necessitates engaging digital health tools which incorporate both the preferences and needs of vulnerable populations. Innovation: Designing a point-of-service health kiosk that 1) draws on user preferences early in the design phase, 2) integrates multiple communication technologies, and 3) disseminates culturally grounded HPV vaccination decisions narratives that are tailored to vaccination awareness level is a promising approach in reducing barriers to HPV vaccine education and vaccine uptake among low-income women at safety-net clinics.

Roadmap on Deep Learning for Microscopy.

Through digital imaging, microscopy has evolved from primarily being a means for visual observation of life at the micro- and nano-scale, to a quantitative tool with ever-increasing resolution and throughput. Artificial intelligence, deep neural networks, and machine learning are all niche terms describing computational methods that have gained a pivotal role in microscopy-based research over the past decade. This Roadmap is written collectively by prominent researchers and encompasses selected aspects of how machine learning is applied to microscopy image data, with the aim of gaining scientific knowledge by improved image quality, automated detection, segmentation, classification and tracking of objects, and efficient merging of information from multiple imaging modalities. We aim to give the reader an overview of the key developments and an understanding of possibilities and limitations of machine learning for microscopy. It will be of interest to a wide cross-disciplinary audience in the physical sciences and life sciences.

Examining engagement with and acceptability and usability of REAL Parenting: A brief online parent-based intervention to reduce alcohol use and consequences among high school students.

This study is a formative assessment of REAL Parenting (RP): a brief, digital intervention for parents of high school students that encourages parent-teen communication about alcohol and, in turn, aims to prevent teen alcohol use. The aims of this study were to describe engagement with, and acceptability and usability of RP; and to explore the relationship of these measures with each other and with short-term outcomes. Participants were 160 parents randomly assigned to the treatment group who received RP as part of a randomized pilot trial (Mage = 45.43[SD = 7.26], 59.3% female, 56% White, 19% Hispanic). App-based program analytics captured real-time engagement with RP. Parents completed self-report measures of acceptability, usability, perceived communication effectiveness, perceived self-efficacy to communicate, and frequency of communication post-intervention. Descriptive statistics were calculated to describe engagement, acceptability and usability, and zero-order correlations were calculated to examine associations between these and self-report variables. About 75% (n = 118) of parents accessed the intervention and two-thirds (n = 110) accessed at least one module. Self-report ratings of acceptability and usability were neutral to positive, and mothers liked RP more than fathers. Self-report, but not program analytic indicators were associated with short-term outcomes. Findings suggest that, with little incentive, most parents will access an app focused on parent-teen communication about alcohol. While parent feedback was positive, it also highlighted areas for improvement with app content and design. Correlations suggest that analytic metrics of engagement are useful to discern who is and is not using interventions, and self-report measures are important for understanding pathways by which interventions are associated with short-term outcomes.

A de novo protein catalyzes the synthesis of semiconductor quantum dots.

De novo proteins constructed from novel amino acid sequences are distinct from proteins that evolved in nature. Construct K (ConK) is a binary-patterned de novo designed protein that rescues Escherichia coli from otherwise toxic concentrations of copper. ConK was recently found to bind the cofactor PLP (pyridoxal phosphate, the active form of vitamin B6). Here, we show that ConK catalyzes the desulfurization of cysteine to H2S, which can be used to synthesize CdS nanocrystals in solution. The CdS nanocrystals are approximately 3 nm, as measured by transmission electron microscope, with optical properties similar to those seen in chemically synthesized quantum dots. The CdS nanocrystals synthesized using ConK have slower growth rates and a different growth mechanism than those synthesized using natural biomineralization pathways. The slower growth rate yields CdS nanocrystals with two desirable properties not observed during biomineralization using natural proteins. First, CdS nanocrystals are predominantly of the zinc blende crystal phase; this is in stark contrast to natural biomineralization routes that produce a mixture of zinc blende and wurtzite phase CdS. Second, in contrast to the growth and eventual precipitation observed in natural biomineralization systems, the CdS nanocrystals produced by ConK stabilize at a final size. Future optimization of CdS nanocrystal growth using ConK-or other de novo proteins-may help to overcome the limits on nanocrystal quality typically observed from natural biomineralization by enabling the synthesis of more stable, high-quality quantum dots at room temperature.

The Student Athlete Wellness Portal: Translating Student Athletes' Prescription Opioid Use Narratives into a Targeted Public Health Intervention.

Background and Objectives: The opioid epidemic has permeated all strata of society over the last two decades, especially within the adolescent student athletic environment, a group particularly at risk and presenting their own challenges for science and practice. This paper (a) describes the development of a web-based intervention called the Student Athlete Wellness Portal that models effective opioid misuse resistance strategies and (b) details the findings of a single-group design to test its effectiveness. Materials and Methods: Formative research included 35 student athletes residing in the United States, ages 14 to 21, who had been injured in their school-based sport. They participated in in-depth qualitative interviews to explore narratives relating to their injuries and pain management plans. Inductive analyses of interview transcripts revealed themes of the challenges of being a student athlete, manageable vs. unmanageable pain, and ways to stay healthy. These themes were translated into prevention messages for the portal, which was then tested in a single-group design. Results: Users of the portal (n = 102) showed significant decreases in their willingness to misuse opioids and increases in their perceptions of opioid risks. Conclusions: This manuscript illuminates the processes involved in translating basic research knowledge into intervention scripts and reveals the promising effects of a technology-based wellness portal.

The impact of HPV vaccine narratives on social media: Testing narrative engagement theory with a diverse sample of young adults.

Rates of human papillomavirus (HPV) infection are highest in young adults, who can be vaccinated against HPV if they were not vaccinated as adolescents. Since young adults increasingly access health information on social media, we tested the impact of a social media campaign with narrative-based health information on intentions related to HPV vaccination. We also aimed to understand which ads resonated most with young adults and led to higher survey completion rates. We created social media posts featuring videos promoting HPV vaccination. We launched a sponsored ad campaign on Facebook to reach young women, ages 18-26, across the country. Participants were randomly assigned one of 6 videos and then completed a brief survey about video engagement and intentions to: talk with a health care professional, talk with friends or family, and vaccinate against HPV. A descriptive correlational design and a test for moderation were used to explore hypothesized relationships. Across all ads, 1332 link clicks led to 991 completed surveys that were reduced to 607 surveys (95 % ages 18-26, 63 % non-Caucasian; 58 % sexually active). Higher video engagement was associated with stronger intentions to talk with a health care professional (r = 0.44, p =.01), talk with friends/family (r = 0.52, p =.01), and vaccinate against HPV (r = 0.43, p =.01). Young adults were receptive to watching narrative-based health information videos on social media. When promoting HPV vaccination, more engaging information leads to greater intentions to talk about the vaccine and get vaccinated.

Mars Oxygen ISRU Experiment (MOXIE)-Preparing for human Mars exploration.

MOXIE [Mars Oxygen In Situ Resource Utilization (ISRU) Experiment] is the first demonstration of ISRU on another planet, producing oxygen by solid oxide electrolysis of carbon dioxide in the martian atmosphere. A scaled-up MOXIE would contribute to sustainable human exploration of Mars by producing on-site the tens of tons of oxygen required for a rocket to transport astronauts off the surface of Mars, instead of having to launch hundreds of tons of material from Earth's surface to transport the required oxygen to Mars. MOXIE has produced oxygen seven times between landing in February 2021 and the end of 2021 and will continue to demonstrate oxygen production during night and day throughout all martian seasons. This paper reviews what MOXIE has accomplished and the implications for larger-scale oxygen-producing systems.

Motivators and Barriers to HPV Vaccination: A Qualitative Study of Underserved Women Attending Planned Parenthood.

Human papilloma virus (HPV) is the most common sexually transmitted infection in the United States. Disease-associated strains of HPV can cause genital warts and six cancer types. HPV-associated cervical cancer disproportionately impacts medically underserved women including Black and Latina women with respect to incidence, prevalence, and mortality rates. Although safe and effective vaccines are available, HPV vaccination rates remain low among low-income individuals and women of color. The current study examined individual and structural motivators and barriers to HPV vaccination among medically underserved women utilizing a Planned Parenthood health center in Southeast Pennsylvania. Guided by narrative engagement theory (NET), qualitative interviews (N = 24) were used to elicit HPV vaccine decision stories from both vaccinated and unvaccinated women. Using a phronetic iterative data analysis approach, we identified three motivators to vaccinate against HPV: (1) receiving an explicit vaccine recommendation from a healthcare provider (a structural determinant), (2) feeling empowered to take control of one's health (an individual determinant), and (3) knowing someone infected with HPV (an individual determinant). Among unvaccinated participants, barriers to HPV vaccination included: (1) not receiving an explicit vaccine recommendation from a healthcare provider (a structural determinant), (2) low perceived risk for acquiring HPV or that HPV is not severe (an individual determinant), and (3) lack of maternal support to vaccinate (a structural determinant). Healthcare providers are optimally positioned to fill the gap in prior missed vaccine opportunities and empower women by recommending HPV vaccination.

A snapshot of parenting practices useful for preventing adolescent vaping.

INTRODUCTION: Smoking research demonstrates that parents can influence their adolescent's tobacco smoking perceptions and behaviors, but little is known about the protective effects of different parenting practices on adolescent vaping. In this study we investigate how adolescent perceptions of parents' knowledge of their activities and parental media mediation are associated with adolescents' perceptions of vaping and adolescent vaping behaviors. METHOD: Six hundred thirty-nine youth (65.7% female, average age: 14.71 years old) recruited through 4-H clubs in nine states participated in a study evaluating a substance use intervention program. Because the evaluation design could influence participants, we used only baseline data. An online self-reported survey was administrated. Most youth self-identified as White (87.3%) and only handful youth indicated Asian (3.4%), African American (3.4%), American Indian (1.1%), and other or unreported (4.8%). Approximately 60% of youth lived in small town or rural areas in US. RESULTS: Analyses revealed that parental knowledge was positively related with adolescent perceived harm of vaping and perceived prevalence of vaping, but was negatively related with perceived acceptability of vaping and social expectancy of vaping. In addition, youth who reported greater parental media mediation were more likely to perceive the harm of vaping and less likely to vape compared with youth with lower parental media mediation. CONCLUSION: These findings suggest that parental education about vaping, including those promoting conversations regarding vaping and vaping ads, may be important to the prevention of adolescent vaping.

Public Health Implications of Cannabis Legalization: An Exploration of Adolescent Use and Evidence-Based Interventions.

This article examines the relaxation of state marijuana laws, changes in adolescent use of marijuana, and implications for drug education. Under federal law, use of marijuana remains illegal. In spite of this federal legislation, as of 1 June 2021, 36 states, four territories and the District of Columbia have enacted medical marijuana laws. There are 17 states, two territories and the District of Columbia that have also passed recreational marijuana laws. One of the concerns regarding the enactment of legislation that has increased access to marijuana is the possibility of increased adolescent use of marijuana. While there are documented benefits of marijuana use for certain medical conditions, we know that marijuana use by young people can interfere with brain development, so increased marijuana use by adolescents raises legitimate health concerns. A review of results from national survey data, including CDC's YRBS, Monitoring the Future, and the National Household Survey on Drug Use, allows us to document changes in marijuana use over time. Increased legal access to marijuana also has implications for educational programming. A "Reefer Madness" type educational approach no longer works (if it ever did). We explore various strategies, including prevention programs for education about marijuana, and make recommendations for health educators.

Development of opioid rapid response system: Protocol for a randomized controlled trial.

BACKGROUND: Opioid overdoses require a rapid response, but emergency responders are limited in how quickly they can arrive at the scene for administering naloxone. If laypersons are trained to administer naloxone and are notified of overdoses, more lives can be saved. OBJECTIVE: This study aimed to examine the feasibility of the Opioid Rapid Response System (ORRS) that recruits, trains, and links citizen responders to overdose events in their community in real-time to administer naloxone. Aim of this paper is to present the protocols for recruiting participants through multiple communication channels; developing and evaluating the online training which has both interactive and asynchronous modules; randomly assigning laypersons to either online naloxone training or waitlist control group; measuring participants' knowledge, skills, and attitudes before and after the training; and distributing intranasal naloxone kits to participants for use in events of overdose in their community. METHODS: Sampling: Utilizing a combination of purposive sampling methods, laypersons from across five Indiana counties who did not self-identify as current first responders were invited to participate. DESIGN: In this two-arm randomized waitlist-controlled study (N = 220), individuals were assigned into either online training or waitlist control that received the training two weeks later. ANALYSIS: A linear mixed model will be used for determining the changes in targeted outcomes in the training group and accommodate for fixed and random effects. IMPLICATIONS: While ORRS can become a community-engaged, cost-effective model for technology-based emergency response for opioid overdoses, study protocols can be useful for other emergency response programs that involve laypersons. CLINICALTRIALS: gov Registration Number: NCT04589676.

Multicultural Adaptation of Mighty Girls for Widespread Dissemination: Pilot Study, App Development and Usability Testing, and Gauging Parent Support With Focus Groups.

BACKGROUND: Taking evidence-based interventions to scale is a challenge for prevention science. Mighty Girls is an evidence-based sexual health intervention program that combines classroom sessions with novel, cutting-edge technology (digital puppetry). The program was developed for 7th grade Latinas, but US school and community demographics rarely allow interventions targeting a single ethnic group. Additionally, digital puppetry is costly to scale up, and parent disapproval often prevents successful dissemination of adolescent sexual health programs. Intervening steps along the scaling-up pathway are needed to adapt the program prior to scaling up for dissemination. OBJECTIVE: The aims of this study were to create a multicultural adaptation of the Mighty Girls program using a mobile app that is less costly to disseminate and is acceptable to parents of 7th grade girls. METHODS: This study used a three-phase process to adapt Mighty Girls into Mighty Teens. All phases used purposive (nonprobability) sampling of low-income, multicultural, urban metropolitan groups (7th grade girls and their parents) within central Florida. Phase 1 involved two videotaped implementations of a multicultural adaptation of the classroom sessions, one involving focus groups (N=14) and the other serving as a single-group pretest-posttest pilot study (N=23). Phase 2 involved development of a narrative cell phone app prototype, which was subjected to usability testing (N=25). App usability and engagement were assessed qualitatively (observation, focus group, open-ended questions) and quantitatively. Phase 3 used focus groups to assess parent support for the program (N=6). Qualitative data were analyzed using descriptive content analysis. Quantitative data were analyzed using descriptive statistics and paired t tests. RESULTS: Qualitative findings supported classroom sessions being multicultural, and identified simple changes to improve engagement and learning. Quantitative findings from the second classroom session implementation pilot study indicated a significant pre-post difference in intention to delay sexual intercourse (P=.04). App usability and appeal were supported by a System Usability Scale score of 76 (exceeding 68 per the industry standard) and 83% (20/24) of participants agreeing they would recommend the app to friends. Parents (mothers) expressed only positive regard for program goals, and classroom session and app activities. CONCLUSIONS: This study adapted Mighty Girls into an engaging, easier-to-disseminate, multicultural program, termed Mighty Teens, that uses a narrative-generating app to support behavior change, and is likely to be accepted by parents of 7th grade girls. This study also provides evidence of the preliminary effectiveness of Mighty Teens classroom sessions. The sampling method and sample size were appropriate for adaptation, but research involving a more representative US sample is needed to confirm multicultural fit, parent receptivity, and program effectiveness. Study implications include integrating app use throughout the classroom sessions to build narrative-generating skills across the program and increasing the number of narratives produced, which should in turn increase the program's behavior change potency.

Identifying as a Good Parent: Considering the Communication Theory of Identity for Parents of Children Receiving Palliative Care.

Background: Parents of seriously ill children are at risk of psychosocial morbidity, which may be mitigated by competent family-centered communication and role-affirming conversations. Parent caregivers describe a guiding desire to do a good job in their parenting role but also depict struggling under the intense weight of parental duty. Objectives and Design: Through this case study, the Communication Theory of Identity (CTI) provides a framework for conceptualizing how palliative care teams can help parents cope with this reality. CTI views communication with care teams as formative in the development and enablement of parental perceptions of their "good parenting" role. Results: Palliative care teams may consider the four frames of identity (personal, enacted, relational, and communal) as meaningful dimensions of the parental pursuit to care well for an ill child. Conclusion: Palliative care teams may consider compassionate communication about parental roles to support the directional virtues of multilayered dynamic parental identity.

The Role of Engagement in Effective, Digital Prevention Interventions: the Function of Engagement in the REAL Media Substance Use Prevention Curriculum.

Prevention curricula rely on audience engagement to effectively communicate their messages. However, to date, measurement of engagement has primarily focused on self-report that is often an indicator of liking or satisfaction. Emerging technologies for intervention delivery hold promise not only for additional engagement indicators but also for dissemination outside of traditional vehicles such as classroom delivery. The present study, grounded in the theory of active involvement (Greene 2013), explores the role of engagement (as measured by self-report, program analytics, and observation) with short-term substance use prevention outcomes such as self-efficacy to counter-argue and descriptive and injunctive norms. The study tracks 4-H youth (N = 310) engaged with a media literacy focused e-learning substance prevention curriculum, REAL media. Results indicate that self-reports of engagement predicted self-efficacy to counter-argue, but a program-analytic indicator of dosage predicted lower injunctive and descriptive norms, all at 3 months. The observational indicator was correlated with self-efficacy to counter-argue but not significant in the predictive models. The implications and directions for future research regarding how engagement is measured in prevention and included in studying program effects are discussed. Clinical trial: NCT03157700, May 2017.