Economical Evolution: Microbes Reduce the Synthetic Cost of Extracellular Proteins

Protein evolution is not simply a race toward improved function. Because organisms compete for limited resources, fitness is also affected by the relative economy of an organism’s proteome. Indeed, many abundant proteins contain relatively high percentages of amino acids that are metabolically less taxing for the cell to make, thus reducing cellular cost. However, not all abundant proteins are economical, and many economical proteins are not particularly abundant. Here we examined protein composition and found that the relative synthetic cost of amino acids constrains the composition of microbial extracellular proteins. In Escherichia coli, extracellular proteins contain, on average, fewer energetically expensive amino acids independent of their abundance, length, function, or structure. Economic pressures have strategically shaped the amino acid composition of multicomponent surface appendages, such as flagella, curli, and type I pili, and extracellular enzymes, including type III effector proteins and secreted serine proteases. Furthermore, in silico analysis of Pseudomonas syringae, Mycobacterium tuberculosis, Saccharomyces cerevisiae, and over 25 other microbes spanning a wide range of GC content revealed a broad bias toward more economical amino acids in extracellular proteins. The synthesis of any protein, especially those rich in expensive aromatic amino acids, represents a significant investment. Because extracellular proteins are lost to the environment and not recycled like other cellular proteins, they present a greater burden on the cell, as their amino acids cannot be reutilized during translation. We hypothesize that evolution has optimized extracellular proteins to reduce their synthetic burden on the cell

Polymerizing the fibre between bacteria and host cells: the biogenesis of functional amyloid fibres

Amyloid fibres are proteinaceous aggregates associated with several human diseases, including Alzheimer's, Huntington's and Creutzfeldt Jakob's. Disease-associated amyloid formation is the result of proteins that misfold and aggregate into β sheet-rich fibre polymers. Cellular toxicity is readily associated with amyloidogenesis, although the molecular mechanism of toxicity remains unknown. Recently, a new class of ‘functional’ amyloid fibres was discovered that demonstrates that amyloids can be utilized as a productive part of cellular biology. These functional amyloids will provide unique insights into how amyloid formation can be controlled and made less cytotoxic. Bacteria produce some of the best-characterized functional amyloids, including a surface amyloid fibre called curli. Assembled by enteric bacteria, curli fibres mediate attachment to surfaces and host tissues. Some bacterial amyloids, like harpins and microcinE492, have exploited amyloid toxicity in a directed and functional manner. Here, we review and discuss the functional amyloids assembled by bacteria. Special emphasis will be paid to the biology of functional amyloid synthesis and the connections between bacterial physiology and pathology.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/75549/1/j.1462-5822.2008.01148.x.pd

Much Ado about Nothing - Why Desert Palace Neither Murdered McDonnell Douglas nor Transformed All Employment Discrimination Cases to Mixed-Motive Essay.

Abstract Forthcoming

Much Ado about Nothing - Why Desert Palace Neither Murdered McDonnell Douglas nor Transformed All Employment Discrimination Cases to Mixed-Motive Essay.

Abstract Forthcoming

Reeves v. Sanderson Plumbing Products: The Emperor Has No Clothes - Pretext Plus Is Alive and Kicking.

Before the Supreme Court’s decision in Reeves v. Sanderson Plumbing Products, Inc., the Fifth Circuit’s en banc decision in Rhodes v. Guiberson Oil Tools established the proper standard of causation in employment discrimination cases. The plaintiff must prove his or her protected trait was the “determinative reason” for the challenged employment action. Following Reeves, which appeared to overrule Rhodes and the doctrine of pretext plus, the Fifth Circuit struggled with the causation question. Despite the apparent confusion, the Fifth Circuit has largely reaffirmed not only its commitment to the Rhodes pretext-plus analysis, but also the determinative-reason standard for pretext cases. In order to avoid summary judgment, employment plaintiffs are required to introduce sufficient evidence from which a reasonable jury could find employment discrimination. Reeves is not the watershed case many have proclaimed it to be, it merely disapproved of one circuit court panel’s review of the sufficiency of the evidence supporting a jury’s verdict in a single case. Reeves is merely a recitation of St. Mary’s Honor Center v. Hicks concept of “pretext plus,” as such, Rhodes remains good law in the Fifth Circuit. Furthermore, its “determinative-reason” standard of causation in discrimination cases also continues to be valid. Yet, the Fifth Circuit has reconfirmed both concepts. In light of this analysis, the only thing which can truly be said of Reeves is that it was poorly written, as it does nothing to articulate any new theory of law and offers little to clarify any existing law. Notwithstanding Reeves’s alleged overruling of pretext plus, the Court bungled the concept of pretext plus, and by association clearly established employment law concerning plaintiff’s burden of proof. Therefore, as implicitly recognized by subsequent opinions, the Court’s confusing opinion in Reeves adds nothing to the fabric of employment law

Reeves v. Sanderson Plumbing Products: The Emperor Has No Clothes - Pretext Plus Is Alive and Kicking.

Before the Supreme Court’s decision in Reeves v. Sanderson Plumbing Products, Inc., the Fifth Circuit’s en banc decision in Rhodes v. Guiberson Oil Tools established the proper standard of causation in employment discrimination cases. The plaintiff must prove his or her protected trait was the “determinative reason” for the challenged employment action. Following Reeves, which appeared to overrule Rhodes and the doctrine of pretext plus, the Fifth Circuit struggled with the causation question. Despite the apparent confusion, the Fifth Circuit has largely reaffirmed not only its commitment to the Rhodes pretext-plus analysis, but also the determinative-reason standard for pretext cases. In order to avoid summary judgment, employment plaintiffs are required to introduce sufficient evidence from which a reasonable jury could find employment discrimination. Reeves is not the watershed case many have proclaimed it to be, it merely disapproved of one circuit court panel’s review of the sufficiency of the evidence supporting a jury’s verdict in a single case. Reeves is merely a recitation of St. Mary’s Honor Center v. Hicks concept of “pretext plus,” as such, Rhodes remains good law in the Fifth Circuit. Furthermore, its “determinative-reason” standard of causation in discrimination cases also continues to be valid. Yet, the Fifth Circuit has reconfirmed both concepts. In light of this analysis, the only thing which can truly be said of Reeves is that it was poorly written, as it does nothing to articulate any new theory of law and offers little to clarify any existing law. Notwithstanding Reeves’s alleged overruling of pretext plus, the Court bungled the concept of pretext plus, and by association clearly established employment law concerning plaintiff’s burden of proof. Therefore, as implicitly recognized by subsequent opinions, the Court’s confusing opinion in Reeves adds nothing to the fabric of employment law

MRI radiomic features are independently associated with overall survival in soft tissue sarcoma

Purpose: Soft tissue sarcomas (STS) represent a heterogeneous group of diseases, and selection of individualized treatments remains a challenge. The goal of this study was to determine whether radiomic features extracted from magnetic resonance (MR) images are independently associated with overall survival (OS) in STS. Methods and Materials: This study analyzed 2 independent cohorts of adult patients with stage II-III STS treated at center 1 (N = 165) and center 2 (N = 61). Thirty radiomic features were extracted from pretreatment T1-weighted contrast-enhanced MR images. Prognostic models for OS were derived on the center 1 cohort and validated on the center 2 cohort. Clinical-only (C), radiomics-only (R), and clinical and radiomics (C+R) penalized Cox models were constructed. Model performance was assessed using Harrell\u27s concordance index. Results: In the R model, tumor volume (hazard ratio [HR], 1.5) and 4 texture features (HR, 1.1-1.5) were selected. In the C+R model, both age (HR, 1.4) and grade (HR, 1.7) were selected along with 5 radiomic features. The adjusted c-indices of the 3 models ranged from 0.68 (C) to 0.74 (C+R) in the derivation cohort and 0.68 (R) to 0.78 (C+R) in the validation cohort. The radiomic features were independently associated with OS in the validation cohort after accounting for age and grade (HR, 2.4; Conclusions: This study found that radiomic features extracted from MR images are independently associated with OS when accounting for age and tumor grade. The overall predictive performance of 3-year OS using a model based on clinical and radiomic features was replicated in an independent cohort. Optimal models using clinical and radiomic features could improve personalized selection of therapy in patients with STS

Oxycodone Ingestion Patterns in Acute Fracture Pain: a Pilot Study Using a Digital Pill

Background: Opioids are prescribed for acute pain as needed, but no data exists on how patients take opioids after discharge from the ED. This places the onus of dosing on the patient and contributes to variable prescribing by ED physicians. ED opioid prescriptions serve as a source for unintentional exposure and contribute to the opioid epidemic. We deployed a digital pill to measure opioid ingestion patterns in individuals discharged after acute fractures. Methods: This pilot study involved individuals without chronic opioid use (i.e. prescribed opioids \u3e 1 week) who were discharged from the ED following acute fracture. Participants were trained to use a digital pill system comprising a single pill (5 mg oxycodone tablet + radiofrequency emitter) and a hip mounted receiver. Upon contact with gastric contents, the digital pill transmitted a radio signal to the receiver, which relayed time of ingestion via cellular 3G signal in real-time to a cloud based server. Participants were instructed to take 1-2 oxycodone digital pills as needed every 8 hours for pain. Participants returned unused medication at orthopedic follow up or 1 week post discharge where any discrepancies between digital pill data and pill counts were reconciled. Results: We enrolled 10 participants (mean age 42). 50% of fractures were managed operatively and 50% were managed nonoperatively. The system recorded ingestions with 85% accuracy. Participants ingested a mean 43 mg oxycodone during the 1 week study period with dose de-escalation occurring after 24 hours. Participants ingested a mean 75.8% of their 1 week total dose in the first 72 hours. 40% of participants stopped taking opioids by 96 hours. 40% of participants remained on opioids 1 week after injury; all required operative treatment. Conclusions: This is the first study to determine opioid ingestion patterns in ED patients discharged with acute fracture pain. Participants self-tapered opioids after 24 hours, most ingestion occurred in the first 72 hours, and a substantial proportion (40%) stopped ingesting oxycodone by 96 hours. Our data shows individuals may require less opioid analgesics than previously considered for acute fracture pain. Additional studies should address ingestion patterns in other painful conditions and development of ED-based interventions to minimize outpatient opioid use while controlling pain

Development and assessment of a clinical calculator for estimating the likelihood of recurrence and survival among patients with locally advanced rectal cancer treated with chemotherapy, radiotherapy, and surgery

Importance: Predicting outcomes in patients receiving neoadjuvant therapy for rectal cancer is challenging because of tumor downstaging. Validated clinical calculators that can estimate recurrence-free survival (RFS) and overall survival (OS) among patients with rectal cancer who have received multimodal therapy are needed. Objective: To develop and validate clinical calculators providing estimates of rectal cancer recurrence and survival that are better for individualized decision-making than the American Joint Committee on Cancer (AJCC) staging system or the neoadjuvant rectal (NAR) score. Design, Setting, and Participants: This prognostic study developed risk models, graphically represented as nomograms, for patients with incomplete pathological response using Cox proportional hazards and multivariable regression analyses with restricted cubic splines. Because patients with complete pathological response to neoadjuvant therapy had uniformly favorable outcomes, their predictions were obtained separately. The study included 1400 patients with stage II or III rectal cancer who received treatment with chemotherapy, radiotherapy, and surgery at 2 comprehensive cancer centers (Memorial Sloan Kettering [MSK] Cancer Center and Siteman Cancer Center [SCC]) between January 1, 1998, and December 31, 2017. Patients from the MSK cohort received chemoradiation, surgery, and adjuvant chemotherapy from January 1, 1998, to December 31, 2014; these patients were randomly assigned to either a model training group or an internal validation group. Models were externally validated using data from the SCC cohort, who received either chemoradiation, surgery, and adjuvant chemotherapy (chemoradiotherapy group) or short-course radiotherapy, consolidation chemotherapy, and surgery (total neoadjuvant therapy with short-course radiotherapy group) from January 1, 2009, to December 31, 2017. Data were analyzed from March 1, 2020, to January 10, 2021. Exposures: Chemotherapy, radiotherapy, chemoradiotherapy, and surgery. Main Outcomes and Measures: Recurrence-free survival and OS were the outcome measures, and the discriminatory performance of the clinical calculators was measured with concordance index and calibration plots. The ability of the clinical calculators to predict RFS and OS was compared with that of the AJCC staging system and the NAR score. The models for RFS and OS among patients with incomplete pathological response included postoperative pathological tumor category, number of positive lymph nodes, tumor distance from anal verge, and large- and small-vessel venous and perineural invasion; age was included in the risk model for OS. The final clinical calculators provided RFS and OS estimates derived from Kaplan-Meier curves for patients with complete pathological response and from risk models for patients with incomplete pathological response. Results: Among 1400 total patients with locally advanced rectal cancer, the median age was 57.8 years (range, 18.0-91.9 years), and 863 patients (61.6%) were male, with tumors at a median distance of 6.7 cm (range, 0-15.0 cm) from the anal verge. The MSK cohort comprised 1069 patients; of those, 710 were assigned to the model training group and 359 were assigned to the internal validation group. The SCC cohort comprised 331 patients; of those, 200 were assigned to the chemoradiotherapy group and 131 were assigned to the total neoadjuvant therapy with short-course radiotherapy group. The concordance indices in the MSK validation data set were 0.70 (95% CI, 0.65-0.76) for RFS and 0.73 (95% CI, 0.65-0.80) for OS. In the external SCC data set, the concordance indices in the chemoradiotherapy group were 0.71 (95% CI, 0.62-0.81) for RFS and 0.72 (95% CI, 0.59-0.85) for OS; the concordance indices in the total neoadjuvant therapy with short-course radiotherapy group were 0.62 (95% CI, 0.49-0.75) for RFS and 0.67 (95% CI, 0.46-0.84) for OS. Calibration plots confirmed good agreement between predicted and observed events. These results compared favorably with predictions based on the AJCC staging system (concordance indices for MSK validation: RFS = 0.69 [95% CI, 0.64-0.74]; OS = 0.67 [95% CI, 0.58-0.75]) and the NAR score (concordance indices for MSK validation: RFS = 0.56 [95% CI, 0.50-0.63]; OS = 0.56 [95% CI, 0.46-0.66]). Furthermore, the clinical calculators provided more individualized outcome estimates compared with the categorical schemas (eg, estimated RFS for patients with AJCC stage IIIB disease ranged from 7% to 68%). Conclusions and Relevance: In this prognostic study, clinical calculators were developed and validated; these calculators provided more individualized estimates of the likelihood of RFS and OS than the AJCC staging system or the NAR score among patients with rectal cancer who received multimodal treatment. The calculators were easy to use and applicable to both short- and long-course radiotherapy regimens, and they may be used to inform surveillance strategies and facilitate future clinical trials and statistical power calculations

The UbiI (VisC) aerobic ubiquinone synthase is required for expression of type 1 pili, biofilm formation, and pathogenesis in uropathogenic Escherichia coli

Uropathogenic Escherichia coli (UPEC), which causes the majority of urinary tract infections (UTI), uses pilus-mediated adherence to initiate biofilm formation in the urinary tract. Oxygen gradients within E. coli biofilms regulate expression and localization of adhesive type 1 pili. A transposon mutant screen for strains defective in biofilm formation identified the ubiI (formerly visC) aerobic ubiquinone synthase gene as critical for UPEC biofilm formation. In this study, we characterized a nonpolar ubiI deletion mutant and compared its behavior to that of wild-type bacteria grown under aerobic and anoxic conditions. Consistent with its function as an aerobic ubiquinone-8 synthase, deletion of ubiI in UPEC resulted in reduced membrane potential, diminished motility, and reduced expression of chaperone-usher pathway pili. Loss of aerobic respiration was previously shown to negatively impact expression of type 1 pili. To determine whether this reduction in type 1 pili was due to an energy deficit, wild-type UPEC and the ubiI mutant were compared for energy-dependent phenotypes under anoxic conditions, in which quinone synthesis is undertaken by anaerobic quinone synthases. Under anoxic conditions, the two strains exhibited wild-type levels of motility but produced diminished numbers of type 1 pili, suggesting that the reduction of type 1 pilus expression in the absence of oxygen is not due to a cellular energy deficit. Acute- and chronic-infection studies in a mouse model of UTI revealed a significant virulence deficit in the ubiI mutant, indicating that UPEC encounters enough oxygen in the bladder to induce aerobic ubiquinone synthesis during infection. IMPORTANCE The majority of urinary tract infections are caused by uropathogenic E. coli, a bacterium that can respire in the presence and absence of oxygen. The bladder environment is hypoxic, with oxygen concentrations ranging from 4% to 7%, compared to 21% atmospheric oxygen. This work provides evidence that aerobic ubiquinone synthesis must be engaged during bladder infection, indicating that UPEC bacteria sense and use oxygen as a terminal electron acceptor in the bladder and that this ability drives infection potential despite the fact that UPEC is a facultative anaerobe