Structural Analysis of the CDK-Cyclin Complex of Pho85-Pho80 and Genome-Wide Characterization of the Phosphate Starvation Response in Schizosaccharomyces pombe

Inorganic phosphate is an essential nutrient required by all organisms for optimal growth. During phosphate starvation, Saccharomyces cerevisiae induces a set of genes responsible for the regulation of inorganic phosphate acquisition. The phosphate-responsive signaling (PHO) pathway controls this response, with the CDK-cyclin complex Pho85-Pho80 playing a prominent role. Here we report the X-ray structure of the Pho85-Pho80 complex, identifying the unique structural features that distinguish it from other cell cycle associated CDK-cyclin complexes. The structure reveals a specific salt bridge between a Pho85 arginine and a Pho80 aspartate that maintains a Pho80 loop confirmation important for substrate recognition and makes phosphorylation of the Pho85 activation loop dispensible. We show that a cluster of residues distal to the kinase active site are involved in a high affinity interaction between the Pho80 cyclin and the transcription factor substrate (Pho4). The structure also reveals a separate high affinity binding site for the CDK inhibitor (Pho81). The fission yeast, Schizosaccharomyces pombe, regulates expression of the secreted acid phosphatase $(pho1^+)$ via a non-orthologous PHO pathway. The genes induced by phosphate limitation and the molecular mechanism by which the genetically identified positive $(pho7^+)$ and negative $(csk1^+)$ regulators function are not known. Here we use a combination of molecular biology, expression microarrays, chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-Seq), and global transcriptome sequencing (RNA-Seq) to characterize the role of $pho7^+$ and $csk1^+$ in the PHO response. We show that there is a fast and slow response to phosphate starvation, each with defined regulatory roles. We use ChIP-Seq to identify members of the Pho7 regulon and characterize Pho7 binding dynamics in response to phosphate-limitation and Csk1 activity. We identify a conserved PHO response for the PHO5 $(pho1^+)$, PHO84 $(spbc8e4.01c^+)$, and GIT1 $(spbc1271.09^+)$ orthologs. We show that activation of $pho1^+$ requires Pho7 binding to a UAS in the $pho1^+$ promoter and that a URS is necessary for Csk1 repression. We find that Pho7-dependent activation is not limited to phosphate-starvation, as additional environmental stress response pathways require $pho7^+$ for maximal induction. Using RNA-Seq we show that Pho7 is also involved in regulating non-coding transcription and is a bi-functional transcription factor

Genome-Wide Characterization of the Phosphate Starvation Response in Schizosaccharomyces pombe

Background: Inorganic phosphate is an essential nutrient required by organisms for growth. During phosphate starvation, Saccharomyces cerevisiae activates the phosphate signal transduction (PHO) pathway, leading to expression of the secreted acid phosphatase, PHO5. The fission yeast, Schizosaccharomyces pombe, regulates expression of the ScPHO5 homolog (pho1+) via a non-orthologous PHO pathway involving genetically identified positive (pho7+) and negative (csk1+) regulators. The genes induced by phosphate limitation and the molecular mechanism by which pho7+ and csk1+ function are unknown. Here we use a combination of molecular biology, expression microarrays, and chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-Seq) to characterize the role of pho7+ and csk1+ in the PHO response. Results: We define the set of genes that comprise the initial response to phosphate starvation in S. pombe. We identify a conserved PHO response that contains the ScPHO5 (pho1+), ScPHO84 (SPBC8E4.01c), and ScGIT1 (SPBC1271.09) orthologs. We identify members of the Pho7 regulon and characterize Pho7 binding in response to phosphate-limitation and Csk1 activity. We demonstrate that activation of pho1+ requires Pho7 binding to a UAS in the pho1+ promoter and that Csk1 repression does not regulate Pho7 enrichment. Further, we find that Pho7-dependent activation is not limited to phosphate-starvation, as additional environmental stress response pathways require pho7+ for maximal induction. Conclusions: We provide a global analysis of the transcriptional response to phosphate limitation in S. pombe. Our results elucidate the conserved core regulon induced in response to phosphate starvation in this ascomycete distantly related to S. cerevisiae and provide a better understanding of flexibility in environmental stress response networks.Chemistry and Chemical BiologyMolecular and Cellular Biolog

Prefrontal modulation of the sustained attention network in ageing, a tDCS-EEG co-registration approach

The ability to sustain attention is integral to healthy cognition in aging. The right PFC (rPFC) is critical for maintaining high levels of attentional focus. Whether plasticity of this region can be harnessed to support sustained attention in older adults is unknown. We used transcranial direct current stimulation to increase cortical excitability of the rPFC, while monitoring behavioral and electrophysiological markers of sustained attention in older adults with suboptimal sustained attention capacity. During rPFC transcranial direct current stimulation, fewer lapses of attention occurred and electroencephalography signals of frontal engagement and early visual attention were enhanced. To further verify these results, we repeated the experiment in an independent cohort of cognitively typical older adults using a different sustained attention paradigm. Again, prefrontal stimulation was associated with better sustained attention. These experiments suggest the rPFC can be manipulated in later years to increase top–down modulation over early sensory processing and improve sustained attention performance. This holds valuable information for the development of neurorehabilitation protocols to ameliorate age-related deficits in this capacity

Causal effect of plasminogen activator inhibitor type 1 on coronary heart disease

Background--Plasminogen activator inhibitor type 1 (PAI-1) plays an essential role in the fibrinolysis system and thrombosis. Population studies have reported that blood PAI-1 levels are associated with increased risk of coronary heart disease (CHD). However, it is unclear whether the association reflects a causal influence of PAI-1 on CHD risk. Methods and Results--To evaluate the association between PAI-1 and CHD, we applied a 3-step strategy. First, we investigated the observational association between PAI-1 and CHD incidence using a systematic review based on a literature search for PAI-1 and CHD studies. Second, we explored the causal association between PAI-1 and CHD using a Mendelian randomization approach using summary statistics from large genome-wide association studies. Finally, we explored the causal effect of PAI-1 on cardiovascular risk factors including metabolic and subclinical atherosclerosis measures. In the systematic meta-analysis, the highest quantile of blood PAI-1 level was associated with higher CHD risk comparing with the lowest quantile (odds ratio=2.17; 95% CI: 1.53, 3.07) in an age- and sex-adjusted model. The effect size was reduced in studies using a multivariable-adjusted model (odds ratio=1.46; 95% CI: 1.13, 1.88). The Mendelian randomization analyses suggested a causal effect of increased PAI-1 level on CHD risk (odds ratio=1.22 per unit increase of log-transformed PAI-1; 95% CI: 1.01, 1.47). In addition, we also detected a causal effect of PAI-1 on elevating blood glucose and high-density lipoprotein cholesterol. Conclusions--Our study indicates a causal effect of elevated PAI-1 level on CHD risk, which may be mediated by glucose dysfunction

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

Prevalence, associated factors and outcomes of pressure injuries in adult intensive care unit patients: the DecubICUs study

Funder: European Society of Intensive Care Medicine; doi: http://dx.doi.org/10.13039/501100013347Funder: Flemish Society for Critical Care NursesAbstract: Purpose: Intensive care unit (ICU) patients are particularly susceptible to developing pressure injuries. Epidemiologic data is however unavailable. We aimed to provide an international picture of the extent of pressure injuries and factors associated with ICU-acquired pressure injuries in adult ICU patients. Methods: International 1-day point-prevalence study; follow-up for outcome assessment until hospital discharge (maximum 12 weeks). Factors associated with ICU-acquired pressure injury and hospital mortality were assessed by generalised linear mixed-effects regression analysis. Results: Data from 13,254 patients in 1117 ICUs (90 countries) revealed 6747 pressure injuries; 3997 (59.2%) were ICU-acquired. Overall prevalence was 26.6% (95% confidence interval [CI] 25.9–27.3). ICU-acquired prevalence was 16.2% (95% CI 15.6–16.8). Sacrum (37%) and heels (19.5%) were most affected. Factors independently associated with ICU-acquired pressure injuries were older age, male sex, being underweight, emergency surgery, higher Simplified Acute Physiology Score II, Braden score 3 days, comorbidities (chronic obstructive pulmonary disease, immunodeficiency), organ support (renal replacement, mechanical ventilation on ICU admission), and being in a low or lower-middle income-economy. Gradually increasing associations with mortality were identified for increasing severity of pressure injury: stage I (odds ratio [OR] 1.5; 95% CI 1.2–1.8), stage II (OR 1.6; 95% CI 1.4–1.9), and stage III or worse (OR 2.8; 95% CI 2.3–3.3). Conclusion: Pressure injuries are common in adult ICU patients. ICU-acquired pressure injuries are associated with mainly intrinsic factors and mortality. Optimal care standards, increased awareness, appropriate resource allocation, and further research into optimal prevention are pivotal to tackle this important patient safety threat

RAD51B in familial breast cancer

Common variation on 14q24.1, close to RAD51B, has been associated with breast cancer: rs999737 and rs2588809 with the risk of female breast cancer and rs1314913 with the risk of male breast cancer. The aim of this study was to investigate the role of RAD51B variants in breast cancer predisposition, particularly in the context of familial breast cancer in Finland. We sequenced the coding region of RAD51B in 168 Finnish breast cancer patients from the Helsinki region for identification of possible recurrent founder mutations. In addition, we studied the known rs999737, rs2588809, and rs1314913 SNPs and RAD51B haplotypes in 44,791 breast cancer cases and 43,583 controls from 40 studies participating in the Breast Cancer Association Consortium (BCAC) that were genotyped on a custom chip (iCOGS). We identified one putatively pathogenic missense mutation c.541C>T among the Finnish cancer patients and subsequently genotyped the mutation in additional breast cancer cases (n = 5259) and population controls (n = 3586) from Finland and Belarus. No significant association with breast cancer risk was seen in the meta-analysis of the Finnish datasets or in the large BCAC dataset. The association with previously identified risk variants rs999737, rs2588809, and rs1314913 was replicated among all breast cancer cases and also among familial cases in the BCAC dataset. The most significant association was observed for the haplotype carrying the risk-alleles of all the three SNPs both among all cases (odds ratio (OR): 1.15, 95% confidence interval (CI): 1.11-1.19, P = 8.88 × 10-16) and among familial cases (OR: 1.24, 95% CI: 1.16-1.32, P = 6.19 × 10-11), compared to the haplotype with the respective protective alleles. Our results suggest that loss-of-function mutations in RAD51B are rare, but common variation at the RAD51B region is significantly associated with familial breast cancer risk.</p