AlphaFold: Research accelerator and hypothesis generator.

To celebrate the 50th anniversary of Cell Press and the Cell focus issue on structural biology, we discussed with scientists working across diverse fields how AlphaFold has changed their research and brought structural biology to the masses.

Incomplete transcripts dominate the Mycobacterium tuberculosis transcriptome.

Mycobacterium tuberculosis (Mtb) is a bacterial pathogen that causes tuberculosis (TB), an infectious disease that is responsible for major health and economic costs worldwide1. Mtb encounters diverse environments during its life cycle and responds to these changes largely by reprogramming its transcriptional output2. However, the mechanisms of Mtb transcription and how they are regulated remain poorly understood. Here we use a sequencing method that simultaneously determines both termini of individual RNA molecules in bacterial cells3 to profile the Mtb transcriptome at high resolution. Unexpectedly, we find that most Mtb transcripts are incomplete, with their 5' ends aligned at transcription start sites and 3' ends located 200-500 nucleotides downstream. We show that these short RNAs are mainly associated with paused RNA polymerases (RNAPs) rather than being products of premature termination. We further show that the high propensity of Mtb RNAP to pause early in transcription relies on the binding of the σ-factor. Finally, we show that a translating ribosome promotes transcription elongation, revealing a potential role for transcription-translation coupling in controlling Mtb gene expression. In sum, our findings depict a mycobacterial transcriptome that prominently features incomplete transcripts resulting from RNAP pausing. We propose that the pausing phase constitutes an important transcriptional checkpoint in Mtb that allows the bacterium to adapt to environmental changes and could be exploited for TB therapeutics.

Compensatory evolution in NusG improves fitness of drug-resistant M. tuberculosis.

Drug-resistant bacteria are emerging as a global threat, despite frequently being less fit than their drug-susceptible ancestors1-8. Here we sought to define the mechanisms that drive or buffer the fitness cost of rifampicin resistance (RifR) in the bacterial pathogen Mycobacterium tuberculosis (Mtb). Rifampicin inhibits RNA polymerase (RNAP) and is a cornerstone of modern short-course tuberculosis therapy9,10. However, RifR Mtb accounts for one-quarter of all deaths due to drug-resistant bacteria11,12. We took a comparative functional genomics approach to define processes that are differentially vulnerable to CRISPR interference (CRISPRi) inhibition in RifR Mtb. Among other hits, we found that the universally conserved transcription factor NusG is crucial for the fitness of RifR Mtb. In contrast to its role in Escherichia coli, Mtb NusG has an essential RNAP pro-pausing function mediated by distinct contacts with RNAP and the DNA13. We find this pro-pausing NusG-RNAP interface to be under positive selection in clinical RifR Mtb isolates. Mutations in the NusG-RNAP interface reduce pro-pausing activity and increase fitness of RifR Mtb. Collectively, these results define excessive RNAP pausing as a molecular mechanism that drives the fitness cost of RifR in Mtb, identify a new mechanism of compensation to overcome this cost, suggest rational approaches to exacerbate the fitness cost, and, more broadly, could inform new therapeutic approaches to develop drug combinations to slow the evolution of RifR in Mtb.

Risk prevalence, readiness and confidence to change lifestyle risk factors among clients of community mental health services.

OBJECTIVE: People with mental health conditions have high rates of chronic physical diseases, partially attributable to lifestyle risks factors. This study examined risk prevalence among community mental health service clients, their readiness and confidence to change, and associations with participant characteristics. METHODS: Cross-sectional survey of adult clients from 12 community mental health services across 3 local health districts in New South Wales, Australia, collected from 2021 to 2022. Participants (n = 486) completed a telephone interview determining five risk factors, and readiness and confidence to change these. Multiple binary logistic regression models determined associations between readiness and confidence (for each risk), and participant characteristics (demographics and diagnosis). RESULTS: Participants most commonly reported a diagnosis of schizophrenia (36.7%) or depression (21.1%). Risk factors were prevalent: ranging from 26% (harmful alcohol use) to 97% (poor nutrition). High readiness was greatest for smoking (68%), weight (66%) and physical inactivity (63%), while confidence was highest for changing alcohol use (67%). Two significant associations were identified; females were more likely than males to have high readiness to change nutrition (odds ratio = 1.14, confidence interval = [1.13, 2.34], p = 0.0092), with males more likely to have high confidence to change physical activity (odds ratio = 0.91, confidence interval = [0.45, 0.99], p = 0.0109). CONCLUSIONS: Many participants were ready and confident to change risk factors. Gender influenced readiness to change nutrition and physical activity confidence. Training to upskill mental health clinicians in provision of preventive care that builds confidence and readiness levels may aid in supporting positive behaviour change.

Process Evaluation of a Scaled-Up School-Based Physical Activity Program for Adolescents: Physical Activity 4 Everyone.

BACKGROUND: Physical Activity 4 Everyone (PA4E1) is a whole-school physical activity program, with demonstrated efficacy (2012-2014). PA4E1 was adapted (scaled-up) and tested in a scale-up trial (2017-2020). This process evaluation study of the scale-up trial had 2 aims. First, to describe the acceptability, appropriateness, and feasibility of PA4E1 in the scale-up trial, from the perspective of school staff involved in the program management and delivery. Second, to generate themes that may explain school staff assessments of acceptability, appropriateness, and feasibility. METHODS: Data were collected at various time points throughout the 2-year implementation phase. Online surveys were collected from In-School Champions, Head Physical Education teachers, Principals, and Physical Education teachers (quantitative data). Focus groups and interviews were conducted with In-School Champions, Principals, and Physical Education teachers (qualitative data). Existing published data on website engagement, adaptations, modifications, and the scale-up trial primary outcome (implementation of physical activity practices) were triangulated with the quantitative and qualitative during analysis, to generate themes. RESULTS: School staff delivering PA4E1 reported it was highly acceptable, appropriate, and feasible. Seven themes were generated relating to acceptability, appropriateness, and feasibility. The themes related to how the program was funded, the delivery modes of implementation support, the identification of easy-wins, the recruitment of the right in-school champion, facilitating principal buy-in, mitigating the impact of school staff turnover, and engaging the whole school. CONCLUSIONS: Recommendations are made to inform future adaptations for PA4E1 and potentially school-based physical activity programs more generally. The findings may inform future scalability assessments of the suitability of programs for scale-up.

Growth trends clustering: A novel method for detecting forest disturbances and extracting climate signals in tree rings.

Tree-ring widths contain valuable historical information related to both forest disturbances and climate variability and changes within forests. However, current methods are still unable to accurately distinguish between disturbances and climate signals in tree rings, especially in the case of climate anomalies. To address this issue, we developed a novel method, called Growth Trends Clustering (GTC) that uses the distribution characteristics of tree-ring widths within a stand to distinguish the effects of climate and other forest disturbances. GTC employed a Gaussian mixture model to fit the probability density distribution of annual ring-width index (RWI) in a stand. Discriminative criteria were established to cluster diverse sub-distributions from the Gaussian mixture model into categories of growth release, suppression, or normal trends. This approach allowed us to identify the occurrence, duration, and severity of forest disturbances based on percentage changes in the growth release or suppression categories of trees. And the effect of climate on tree growth was assessed according to the mean statistics of the growth normal categories. Using common forest disturbances such as defoliating insects and thinning as examples, we validated our method using tree-ring collections from six sites in British Columbia and Quebec, Canada. We found that the GTC method was superior to traditional time-series analysis methods (e.g., Radial Growth Averaging, Boundary Line, Absolute Increase, and Curve Intervention Detection) for detecting past forest disturbances and was able to significantly enhance climate signals. In summary, the GTC method presented in this study introduces a novel statistical approach for accurately distinguishing between forest disturbances and climate signals in tree rings. This is particularly important for understanding forest disturbance regimes under climate change and for developing future disturbance mitigation strategies.

A randomised trial of real-time video counselling for smoking cessation among rural and remote residents.

INTRODUCTION: Despite its reach, very limited evidence exists on the effectiveness of real-time video counselling for smoking cessation (e.g. via Skype). This study compared the effectiveness of real-time video counselling for smoking cessation to (a) telephone counselling; and (b) a control among rural and remote residents. METHODS: Between 25 May 2017 and 3 March 2020, a three-arm, parallel group, randomised trial, randomised 1244 rural and remote residents from New South Wales, Australia who smoked tobacco to: video counselling (4-6 video sessions); telephone counselling (4-6 telephone calls); or a control (printed materials). The primary outcome was 7-day point prevalence abstinence at 13 months post-baseline. Secondary outcomes were point prevalence abstinence at 4 months and 7-months post-baseline, prolonged abstinence, quit attempts, anxiety and depression. RESULTS: For the primary outcome of 7-day point prevalence abstinence at 13 months post-baseline, there was no significant difference between video counselling and telephone counselling (14.6% vs 13.3%; (OR = 1.11, 95% CI (0.75-1.64), P = 0.61) or video counselling and control (14.6% vs 13.9%; (OR = 1.06, 95% CI (0.71-1.57), P = 0.77). For secondary outcomes at 4 months post-baseline, the video counselling group had significantly higher odds than the control of 7-day point prevalence abstinence (14.3% vs 8.2%; OR = 1.88, 95% CI (1.20-2.95), P = 0.006) and 3-month prolonged abstinence (4.9% vs 2.2%; OR = 2.28, 95% CI (1.03-5.07), P = 0.04). There were no significant differences for other secondary outcomes. DISCUSSION: Video counselling increased smoking cessation in the short-term compared to a control although strategies to improve its long-term effectiveness are needed. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry, https://www.anzctr.org.au ACTRN12617000514303.

Effectiveness of a healthy lifestyle intervention for low back pain and osteoarthritis of the knee: protocol and statistical analysis plan for two randomised controlled trials

ABSTRACT Background These trials are the first randomised controlled trials of telephone-based weight management and healthy lifestyle interventions for low back pain and knee osteoarthritis. This article describes the protocol and statistical analysis plan. Method These trials are parallel randomised controlled trials that investigate and compare the effect of a telephone-based weight management and healthy lifestyle intervention for improving pain intensity in overweight or obese patients with low back pain or knee osteoarthritis. The analysis plan was finalised prior to initiation of analyses. All data collected as part of the trial were reviewed, without stratification by group, and classified by baseline characteristics, process of care and trial outcomes. Trial outcomes were classified as primary and secondary outcomes. Appropriate descriptive statistics and statistical testing of between-group differences, where relevant, have been planned and described. Conclusions A protocol for standard analyses was developed for the results of two randomised controlled trials. This protocol describes the data, and the pre-determined statistical tests of relevant outcome measures. The plan demonstrates transparent and verifiable use of the data collected. This a priori protocol will be followed to ensure rigorous standards of data analysis are strictly adhered to.