A visual screen of protein localization during sporulation identifies new components of prospore membrane-associated complexes in budding yeast.

During ascospore formation in Saccharomyces cerevisiae, the secretory pathway is reorganized to create new intracellular compartments, termed prospore membranes. Prospore membranes engulf the nuclei produced by the meiotic divisions, giving rise to individual spores. The shape and growth of prospore membranes are constrained by cytoskeletal structures, such as septin proteins, that associate with the membranes. Green fluorescent protein (GFP) fusions to various proteins that associate with septins at the bud neck during vegetative growth as well as to proteins encoded by genes that are transcriptionally induced during sporulation were examined for their cellular localization during prospore membrane growth. We report localizations for over 100 different GFP fusions, including over 30 proteins localized to the prospore membrane compartment. In particular, the screen identified IRC10 as a new component of the leading-edge protein complex (LEP), a ring structure localized to the lip of the prospore membrane. Localization of Irc10 to the leading edge is dependent on SSP1, but not ADY3. Loss of IRC10 caused no obvious phenotype, but an ady3 irc10 mutant was completely defective in sporulation and displayed prospore membrane morphologies similar to those of an ssp1 strain. These results reveal the architecture of the LEP and provide insight into the evolution of this membrane-organizing complex.

Metabolic Costs of Walking with Weighted Vests.

INTRODUCTION: The US Army Load Carriage Decision Aid (LCDA) metabolic model is used by militaries across the globe and is intended to predict physiological responses, specifically metabolic costs, in a wide range of dismounted warfighter operations. However, the LCDA has yet to be adapted for vest-borne load carriage, which is commonplace in tactical populations, and differs in energetic costs to backpacking and other forms of load carriage. PURPOSE: The purpose of this study is to develop and validate a metabolic model term that accurately estimates the effect of weighted vest loads on standing and walking metabolic rate for military mission-planning and general applications. METHODS: Twenty healthy, physically active military-age adults (4 women, 16 men; age, 26 ± 8 yr old; height, 1.74 ± 0.09 m; body mass, 81 ± 16 kg) walked for 6 to 21 min with four levels of weighted vest loading (0 to 66% body mass) at up to 11 treadmill speeds (0.45 to 1.97 m·s -1 ). Using indirect calorimetry measurements, we derived a new model term for estimating metabolic rate when carrying vest-borne loads. Model estimates were evaluated internally by k -fold cross-validation and externally against 12 reference datasets (264 total participants). We tested if the 90% confidence interval of the mean paired difference was within equivalence limits equal to 10% of the measured walking metabolic rate. Estimation accuracy, precision, and level of agreement were also evaluated by the bias, standard deviation of paired differences, and concordance correlation coefficient (CCC), respectively. RESULTS: Metabolic rate estimates using the new weighted vest term were statistically equivalent ( P < 0.01) to measured values in the current study (bias, -0.01 ± 0.54 W·kg -1 ; CCC, 0.973) as well as from the 12 reference datasets (bias, -0.16 ± 0.59 W·kg -1 ; CCC, 0.963). CONCLUSIONS: The updated LCDA metabolic model calculates accurate predictions of metabolic rate when carrying heavy backpack and vest-borne loads. Tactical populations and recreational athletes that train with weighted vests can confidently use the simplified LCDA metabolic calculator provided as Supplemental Digital Content to estimate metabolic rates for work/rest guidance, training periodization, and nutritional interventions.

Carbohydrate or Electrolyte Rehydration Recovers Plasma Volume but Not Post-immersion Performance Compared to Water After Immersion Diuresis.

INTRODUCTION: We tested the hypothesis that a carbohydrate (CHO: 6.5%) or carbohydrate-electrolyte (CHO + E: 6.5% + 50 mmol/L NaCl) drink would better recover plasma volume (PV) and exercise performance compared to water (H2O) after immersion diuresis. METHODS: Twelve men (24 ± 2 years; 82.4 ± 15.5 kg; and V̇O2max: 49.8 ± 5.1 mL · kg-1 · min-1) completed four experimental visits: a no-immersion control (CON) and three 4-h cold-water (18.0 °C) immersion trials (H2O, CHO, and CHO + E) followed by exercise in a warm environment (30 °C, 50% relative humidity). The exercise was a 60-minute loaded march (20.4 kg; 55% VO2max) followed by a 10-minute intermittent running protocol. After immersion, subjects were rehydrated with 100% of body mass loss from immersion diuresis during the ruck march. PV is reported as a percent change after immersion, after the ruck march, and after the intermittent running protocol. The intermittent running protocol distance provided an index of exercise performance. Data are reported as mean ± SD. RESULTS: After immersion, body mass loss was 2.3 ± 0.7%, 2.3 ± 0.5%, and 2.3 ± 0.6% for H2O, CHO, and CHO + E. PV loss after immersion was 19.8 ± 8.5% in H2O, 18.2 ± 7.0% in CHO, and 13.9 ± 9.3% in CHO + E, which was reduced after the ruck march to 14.7 ± 4.7% (P = .13) in H2O, 8.8 ± 8.3% (P < .01) in CHO, and 4.4 ± 10.9% (P = .02) in CHO + E. The intermittent running protocol distance was 1.4 ± 0.1 km in CON, 1.4 ± 0.2 km in H2O, 1.4 ± 0.1 km in CHO, and 1.4 ± 0.2 km in CHO + E (P = .28). CONCLUSIONS: Although CHO and CHO + E better restored PV after immersion, post-immersion exercise performance was not augmented compared to H2O, highlighting that fluid replacement following immersion diuresis should focus on restoring volume lost rather than fluid constituents.

Peak performance and cardiometabolic responses of modern US army soldiers during heavy, fatiguing vest-borne load carriage.

INTRODUCTION: Physiological limits imposed by vest-borne loads must be defined for optimal performance monitoring of the modern dismounted warfighter. PURPOSE: To evaluate how weighted vests affect locomotion economy and relative cardiometabolic strain during military load carriage while identifying key physiological predictors of exhaustion limits. METHODS: Fifteen US Army soldiers (4 women, 11 men; age, 26 ± 8 years; height, 173 ± 10 cm; body mass (BM), 79 ± 16 kg) performed four incremental walking tests with different vest loads (0, 22, 44, or 66% BM). We examined the effects of vest-borne loading on peak walking speed, the physiological costs of transport, and relative work intensity. We then sought to determine which of the cardiometabolic indicators (oxygen uptake, heart rate, respiration rate) was most predictive of task failure. RESULTS: Peak walking speed significantly decreased with successively heavier vest loads (p < 0.01). Physiological costs per kilometer walked were significantly higher with added vest loads for each measure (p < 0.05). Relative oxygen uptake and heart rate were significantly higher during the loaded trials than the 0% BM trial (p < 0.01) yet not different from one another (p > 0.07). Conversely, respiration rate was significantly higher with the heavier load in every comparison (p < 0.01). Probability modeling revealed heart rate as the best predictor of task failure (marginal R2, 0.587, conditional R2, 0.791). CONCLUSION: Heavy vest-borne loads cause exceptional losses in performance capabilities and increased physiological strain during walking. Heart rate provides a useful non-invasive indicator of relative intensity and task failure during military load carriage.

Physical and Physiological Characterization of Female Elite Warfighters.

INTRODUCTION: This study characterized a sample of the first women to complete elite United States (US) military training. METHODS: Twelve female graduates of the US Army Ranger Course and one of the first Marine Corps Infantry Officers Course graduates participated in 3 d of laboratory testing including serum endocrine profiles, aerobic capacity, standing broad jump, common soldiering tasks, Army Combat Fitness Test, and body composition (dual-energy x-ray absorptiometry, three-dimensional body surface scans, and anthropometry). RESULTS: The women were 6 months to 4 yr postcourse graduation, 30 ± 6 yr (mean ± SD); height, 1.67 ± 0.07 m; body mass, 69.4 ± 8.2 kg; body mass index, 25.0 ± 2.3 kg·m -2 . Dual-energy x-ray absorptiometry relative fat was 20.0% ± 2.0%; fat-free mass, 53.0 ± 5.9 kg; fat-free mass index, 20.0 ± 1.7 kg·m -2 ; bone mineral content, 2.75 ± 0.28 kg; bone mineral density, 1.24 ± 0.07 g·cm -2 ; aerobic capacity, 48.2 ± 4.8 mL·kg -1 ·min -1 ; total Army Combat Fitness Test score 505 ± 27; standing broad jump 2.0 ± 0.2 m; 123 kg casualty drag 0.70 ± 0.20 m·s -1 , and 4 mile 47 kg ruck march 64 ± 6 min. All women were within normal healthy female range for circulating androgens. Physique from three-dimensional scan demonstrated greater circumferences at eight of the 11 sites compared with the standard military female. CONCLUSIONS: These pioneering women possessed high strength and aerobic capacity, low %BF; high fat-free mass, fat-free mass index, and bone mass and density; and they were not virilized based on endocrine measures as compared with other reference groups. This group is larger in body size and leaner than the average Army woman. These elite physical performers seem most comparable to female competitive strength athletes.

Modeling the Metabolic Costs of Heavy Military Backpacking.

INTRODUCTION: Existing predictive equations underestimate the metabolic costs of heavy military load carriage. Metabolic costs are specific to each type of military equipment, and backpack loads often impose the most sustained burden on the dismounted warfighter. PURPOSE: This study aimed to develop and validate an equation for estimating metabolic rates during heavy backpacking for the US Army Load Carriage Decision Aid (LCDA), an integrated software mission planning tool. METHODS: Thirty healthy, active military-age adults (3 women, 27 men; age, 25 ± 7 yr; height, 1.74 ± 0.07 m; body mass, 77 ± 15 kg) walked for 6-21 min while carrying backpacks loaded up to 66% body mass at speeds between 0.45 and 1.97 m·s-1. A new predictive model, the LCDA backpacking equation, was developed on metabolic rate data calculated from indirect calorimetry. Model estimation performance was evaluated internally by k-fold cross-validation and externally against seven historical reference data sets. We tested if the 90% confidence interval of the mean paired difference was within equivalence limits equal to 10% of the measured metabolic rate. Estimation accuracy and level of agreement were also evaluated by the bias and concordance correlation coefficient (CCC), respectively. RESULTS: Estimates from the LCDA backpacking equation were statistically equivalent (P < 0.01) to metabolic rates measured in the current study (bias, -0.01 ± 0.62 W·kg-1; CCC, 0.965) and from the seven independent data sets (bias, -0.08 ± 0.59 W·kg-1; CCC, 0.926). CONCLUSIONS: The newly derived LCDA backpacking equation provides close estimates of steady-state metabolic energy expenditure during heavy load carriage. These advances enable further optimization of thermal-work strain monitoring, sports nutrition, and hydration strategies.

Microparticles formed during storage of red blood cell units support thrombin generation.

BACKGROUND: Intact red blood cells (RBCs) appear to support thrombin generation in in vitro models of blood coagulation. During storage of RBC units, biochemical, structural, and physiological changes occur including alterations to RBC membranes and release of microparticles, which are collectively known as storage lesion. The clinical consequences of microparticle formation in RBC units are unclear. This study was performed to assess thrombin generation via the prothrombinase complex by washed RBCs and RBC-derived microparticles as a function of RBC unit age. METHODS: Well-characterized kinetic and flow cytometric assays were used to quantify and characterize microparticles isolated from leukocyte-reduced RBC units during storage for 42 days under standard blood banking conditions. RESULTS: Stored RBCs exhibited known features of storage lesion including decreasing pH, cell lysis, and release of microparticles demonstrated by scanning electron microscopy. The rate of thrombin formation by RBC units linearly increased during storage, with the microparticle fraction accounting for approximately 70% of the prothrombinase activity after 35 days. High-resolution flow cytometric analyses of microparticle isolates identified phosphatidylserine-positive RBC-derived microparticles; however, their numbers over time did not correlate with thrombin formation in that fraction. CONCLUSION: Red blood cell-derived microparticles capable of supporting prothrombinase function accumulate during storage, suggesting an increased potential of transfused units as they age to interact in unplanned ways with ongoing hemostatic processes in injured individuals, especially given the standard blood bank practice of using the oldest units available.