Utilization of extracellular information before ligand-receptor binding reaches equilibrium expands and shifts the input dynamic range

Cell signaling systems sense and respond to ligands that bind cell surface receptors. These systems often respond to changes in the concentration of extracellular ligand more rapidly than the ligand equilibrates with its receptor. We demonstrate, by modeling and experiment, a general “systems level” mechanism cells use to take advantage of the information present in the early signal, before receptor binding reaches a new steady state. This mechanism, pre-equilibrium sensing and signaling (PRESS), operates in signaling systems in which the kinetics of ligand-receptor binding are slower than the downstream signaling steps, and it typically involves transient activation of a downstream step. In the systems where it operates, PRESS expands and shifts the input dynamic range, allowing cells to make different responses to ligand concentrations so high as to be otherwise indistinguishable. Specifically, we show that PRESS applies to the yeast directional polarization in response to pheromone gradients. Consideration of preexisting kinetic data for ligand-receptor interactions suggests that PRESS operates in many cell signaling systems throughout biology. The same mechanism may also operate at other levels in signaling systems in which a slow activation step couples to a faster downstream step.Detecting and responding to a chemical gradient is a central feature of a multitude of biological processes (1). For this behavior, organisms use signaling systems that sense information about the extracellular world, transmit this information into the cell, and orchestrate a response. Measurements of the direction and proximity of the extracellular stimuli usually rely on the binding of diffusing chemical particles (ligands) to specific cell surface receptors. Different organisms have evolved different strategies to make use of this information. Small motile organisms, including certain bacteria, use a temporal sensing strategy, measuring and comparing concentration signals over time along their swimming tracks (2). In contrast, some eukaryotic cells, including Saccharomyces cerevisiae, are sufficiently large to implement a spatial sensing mechanism, measuring concentration differences across their cell bodies (3).The observation that some eukaryotes that use spatial sensing exhibit remarkable precision in response to shallow gradients (1–2% differences in ligand concentration between front and rear) (4, 5) has led to several proposed models in which large amplification is achieved by positive feedback loops in the signaling pathways triggered by the ligand-receptor binding (6, 7). Here, we describe a different mechanism, dependent on ligand-receptor binding dynamics, which improves gradient sensing when the concentration of external ligand is close to saturation. We use the budding yeast S. cerevisiae to study the efficiency of this mechanism.Haploid yeast cells exist in two mating types, MATa and MATα (also referred to as a and α cells). Mating occurs when a and α cells sense each other’s secreted mating pheromones: a-factor and α-factor (αF) (8). The pheromone secreted by the nearby mating partner diffuses, forming a gradient surrounding the sensing cell. Pheromone binds a membrane receptor, Ste2, in MATa yeast (9) that activates a pheromone response system (PRS), which cells use to decide whether to fuse with a mating partner or not. At high enough αF concentrations, cells develop a polarized chemotropic growth toward the pheromone source (4). To do that, the nonmotile yeast determines the direction of the potential mating partner measuring on which side there are more bound pheromone receptors, which are initially distributed homogeneously on the cell surface (10). However, this sensing modality can only work when external pheromone is nonsaturating: If all receptors are bound, cells should not be able to determine the direction of the gradient. Surprisingly, even at high pheromone concentrations, yeast tend to polarize in the correct direction (4, 11). Different amplification mechanisms have been proposed to account for the conversion of small differences in ligand concentration across the yeast cell, as is the case for dense mating mixtures, into chemotropic growth (6).We previously studied induction of reporter gene output by the PRS after step increases in the concentration of αF. We found large cell-to-cell variability, the bulk of which was due to large differences in the ability of individual cells to send signal through the system and in their general capacity to express proteins (12). The level of induced gene expression matches well the equilibrium binding curve of αF to receptor (13, 14), a phenomenon known as dose–response alignment (DoRA), common to many other signaling systems (14). In the PRS, DoRA persists for several hours of stimulation.During these studies, we realized that the binding dynamics of αF to its receptor is remarkably slow: At concentrations near the dissociation constant (K_d), binding takes about 20 min to reach 90% of the equilibrium level (15, 16). This dynamics is slow relative not only to the 90-min cell division cycle but also to the pheromone-dependent activation of the mitogen-activated protein kinase (MAPK) Fus3, which takes 2 to 5 min to reach steady-state levels (14). An unavoidable conclusion is that the machinery downstream of the αF receptor must be using pre-equilibrium binding information for its operation.This observation led us to study the consequences of fast and slow ligand-receptor dynamics on the ability of cells to sense extracellular cues. In biology, the rates of ligand binding and unbinding to membrane receptors span a large range, including many cases with dynamics similar to, or even slower than, that of mating pheromone (e.g., rates for EGF, insulin, glucagon, IFN-α1a, and IL-2 in

Antepartum Screening for Group B Streptococcus by Three FDA-Cleared Molecular Tests and Effect of Shortened Enrichment Culture on Molecular Detection Rates

Neonatal Streptococcus agalactiae infections cause significant morbidity and mortality, and antenatal screening is recommended. We compared three U.S. Food and Drug Administration (FDA)-cleared nucleic acid amplification tests (NAATs) to culture using 314 vaginal/rectal swabs after 18 to 24 h (recommended period) and 4 to 8 h (shortened period) of broth enrichment. Agreement of the NAATs with each other was high (97.1% to 98.4%), but culture was less sensitive than all NAATs (67% to 73%). A shortened period of broth culture enrichment resulted in 1 false-negative result in 68 (1.5%). The NAATs performed comparably and were more sensitive than culture.     

Sex Differences in Hierarchical Stability in a Formation of a Mixed-sex Group of Rhesus Macaques

Forming groups of captive rhesus macaques (Macaca mulatta) is a common management practice. New formations of unfamiliar macaques can be costly, with high levels of trauma, particularly as intense aggression is used to establish a dominance hierarchy. Combining previous subgroups into one new group may be beneficial, as some individuals already have established dominance relationships. We tested this hypothesis by forming a new mixed-sex group of rhesus macaques that combined an established group of females with an established group of males. Prior to the mixed-sex group formation, both the female and male hierarchies had been stable for 3 y; after mixed-sex group formation these hierarchies were maintained by the females and were initially maintained by the males for 3 wks. However, the temporary hospitalization (due to a laceration caused by aggression) of the alpha male destabilized the male hierarchy. Age and weight then predicted male rank. Temporary hospitalizations resulted in rank changes for the males, evidenced by reversals in subordination signals. This study indicates that using established groups of familiar individuals may maintain female hierarchical stability in a mixed-sex group formation, but further research is needed to understand how to maintain and predict male hierarchical stability to reduce trauma. Improved knowledge of hierarchical stability would be invaluable to managers of large rhesus macaque groups and would help improve the welfare of captive rhesus macaques.

Social group formations of captive nonhuman primates (NHPs) occur frequently for a variety of reasons (for example, breeding, research purposes, permanent housing arrangements). Rhesus macaques are the most used NHP in biomedical research.¹⁵ Classified as highly despotic, rhesus macaques rely on a heavy use of aggression to maintain and reinforce their dominance hierarchy,⁴⁷ which ultimately governs access to resources and mates.⁵⁰ Extensive aggression can result in significant traumas and even death.^12,51 Reducing trauma is therefore an important goal both to maintain the animals’ welfare and to minimize the associated cost of care. Thus, this research is aimed at improving the success of forming groups with minimal trauma.One strategy to potentially improve the success of group formations and decrease aggression is to mix familiar individuals with an already established dominance relationship.^18,53 However, dominance ranks are not static and can change if aspects of the social environment change,^1,11,17 such as the loss of keystone individuals³⁴ or addition of new allies.¹⁷ As many NHPs are housed in pairs or small social groups, substantial research has been aimed at understanding pairing success,^14,31,38 but less is known about factors that contribute to successful formations of large, naturalistic social groups. Because high levels of aggression and trauma can occur even in well-established groups,^8,45 gauging social stability in large group formations of rhesus macaques is difficult, as aggression does not necessarily equate to incompatibility or trauma.^8,31,38 Understanding whether individuals maintain their previous hierarchies is critical information for behavioral managers as they form new social groups.Wild female rhesus macaques remain in their natal groups and acquire adjacent ranks to their mothers through coalitionary support.¹⁰ Males also retain their ranks near their mothers while in their natal groups,¹⁶ but after dispersal at sexual maturity and entry into a new breeding group, males mainly enter the group at the bottom of the hierarchy and move up after dispersals or deaths (see²⁶ for general background information and an unusual case of rank acquisition). Because males emigrate from multiple natal groups, many of them will be unfamiliar to one another. Whether male dominance relationships in their natal groups affect their rank in subsequent groups as adults is uncertain, although some research suggests that postnatal nepotism occurs in long-tailed macaques (Macaca fascicularis), with males maintaining a high dominance rank longer in non-natal groups when other male kin are present.²⁵ In many groups in which males originate from multiple natal groups, tenure in the new social group often dictates male dominance rank.²⁶The current research sought to determine whether female and male rhesus macaques would maintain their established same-sex hierarchies during a group formation in which an established group of females was introduced to an established group of males. If previous relationships are maintained, this could indicate that using established groups of familiar same-sex individuals could lessen the trauma and associated costs of a group formation, as individuals would not need to use high levels of aggression to sort out their hierarchy. We predicted that previous social rank, rather than individual attributes, would predict social rank in the new mixed-sex social group.     

Complete response to therapy: why do primary central nervous system lymphoma patients not return to work?

Journal of Neuro-Oncology - Evidence supporting routine postoperative antiepileptic drug (AED) prophylaxis following oncologic neurosurgery is limited, and actual practice patterns are largely...     

Prevalence and predictors of low sexual assertiveness

Background: Personal beliefs about sexual attitudes and rights likely affect reproductive health behaviors. We examined the prevalence and predictors of low sexual assertiveness (SA) among sexually active females across three age groups (14-17 yo, 18-21 yo, and 22-26 yo).Methods: 904 white, black, and Mexican-American females presenting at community-based family planning clinics in Southeast Texas between April 1997 and February 1998 completed an anonymous self-report questionnaire that assessed demographic and reproductive characteristics, dating behaviors, mental health measures and occurrence of physical and sexual assault. Subjects also completed a sexual assertiveness scale that assessed one's right to make various reproductive health decisions and sexual communications to partners. A total SA score was obtained by summing the 13 items; scores were then dichotomized so those scoring in the lowest quartile could be compared to the others. Data were analyzed using chi-square and logistic regression stratified by age group (244 aged 14-17; 392 aged 18-21; and 268 aged 22-26).Results: 27% of subjects aged 14-17, 22% aged 18-21, and 17% aged 22-26 were identified as having low SA. For 14-17 years olds, logistic regression analyses revealed that those with low SA as compared to adequate SA were significantly more likely to be Mexican American (OR = 4.8) or black (OR = 3.1) than white, to have hand /= 1 (OR = 1.8). Among 22-26 years olds, low SA was associated with gravidity >/= 1 (OR = 2.8), being a high school dropout (OR = 2.1), and inconsistent birth control use in the last 12 months (OR = 1.9).Conclusions: These data suggest that 1 in 4 sexually active women report low SA and for two age groups these attitudes are associated with inconsistent contraception. Among 18-21 years olds, low SA is also associated with forced sexual contact and depressive symptoms. Thus, efforts to prevent unintended pregnancy and unwanted sexual contact may be more effective if individual levels of SA are taken into account.     

Carbonic anhydrase inhibitor sulthiame reduces intracellular pH and epileptiform activity of hippocampal CA3 neurons

PURPOSE: Sulthiame is a carbonic anhydrase (CA) inhibitor with an anticonvulsant effect in the treatment of benign and symptomatic focal epilepsy in children. The aim of the study was to elucidate the mode of action of sulthiame with respect to possible changes of intracellular pH (pHi) that might develop along with sulthiame's anticonvulsant properties. METHODS: The effects of sulthiame (a) on pHi of 2',7-bis(2-carboxyethyl)-5(6)-carboxyfluorescein-acetoxymetyl ester (BCECF-AM) loaded CA3 neurones as well as (b) on epileptiform activity (induced by 50 microM 4-aminopyridine) were compared with those of the CA inhibitors acetazolamide and benzolamide. RESULTS: In the majority of neurons, sulthiame (1.0-1.5 mM; n = 8) as well as the membrane permeant acetazolamide (0.5-1.0 mM; n = 6) reversibly decreased pHi by 0.18 +/- 0.05 (SD) and 0.17 +/- 0.10 (SD) pH units, respectively, within 10 min. The poor membrane permeant benzolamide (1.0-2.0 mM) had no influence on pHi (n = 8). Sulthiame (1.0-2.5 mM) and acetazolamide (1.0-2.0 mM) reversibly reduced the frequency of action potentials and epileptiform bursts after 10-15 min (n = 9, n = 7), whereas benzolamide (1.0-2.0 mM) had no effect (n = 6). CONCLUSIONS: The results suggest that sulthiame acts as a membrane-permeant CA inhibitor whose beneficial effect on epileptiform activity results at least in part from a modest intracellular acidosis of central neurons.     

Soy protein isolates of varying isoflavone content exert minor effects on serum reproductive hormones in healthy young men

Inverse associations between soy and prostate cancer and the contribution of hormones to prostate cancer prompted the current study to determine whether soy protein could alter serum hormones in men. Thirty-five men consumed milk protein isolate (MPI), low-isoflavone soy protein isolate (SPI) (low-iso SPI; 1.64 +/- 0.19 mg isoflavones/d), and high-iso SPI (61.7 +/- 7.35 mg isoflavones/d) for 57 d each in a randomized crossover design. Twenty-four-hour urine samples indicated that urinary isoflavones were significantly increased by the high-iso SPI relative to the low-iso SPI and MPI. Serum collected on d 1, 29, and 57 of each treatment revealed that dihydrotestosterone (DHT) and DHT/testosterone were significantly decreased by the low-iso SPI [9.4% (P = 0.036) and 9.0% (P = 0.004), respectively] and the high-iso SPI [15% (P = 0.047) and 14% (P = 0.013), respectively], compared with the MPI at d 57. Other significant effects included a decrease in testosterone by the low-iso SPI relative to the MPI (P = 0.023) and high-iso SPI (P = 0.020) at d 29; an increase in dehydroepiandrosterone sulfate by the low-iso SPI relative to the MPI at d 29 (P = 0.001) and relative to the MPI (P = 0.0003) and high-iso SPI (P = 0.005) at d 57; and increases in estradiol and estrone by the low-iso SPI relative to the MPI at d 57 (P = 0.010 and P = 0.005, respectively). In conclusion, soy protein, regardless of isoflavone content, decreased DHT and DHT/testosterone with minor effects on other hormones, providing evidence for some effects of soy protein on hormones. The relevance of the magnitude of these effects to future prostate cancer risk requires further investigation.     

HSP27 in patients with ovarian carcinoma: still an independent prognostic indicator at 60 months follow-up

OBJECTIVE: Heat shock protein 27 (HSP27) is produced in response to pathophysiologic stress in animal cells. The authors have previously shown that HSP27 is an independent prognostic indicator in patients with ovarian carcinoma. The present study was performed to see whether HSP27 remained an independent prognostic indicator with longer follow-up. METHODS: One hundred and three consecutive patients with epithelial ovarian carcinoma were studied. Slides were prepared from fresh tissue. HPS27 staining was performed as previously described. Patient records were examined for FIGO stage, grade, histology, level of cytoreduction and survival. RESULTS: One hundred and three patients were followed for a mean of 60 months. Twenty patients had FIGO Stage I disease, four Stage II, 59 Stage III, and 20 Stage IV. Immunohistochemical (IHC) staining for HSP27 was not related to histologic grade, level of cytoreduction or histologic subtype. A statistically significant decrease in HSP27 staining was found to correlate with increased FIGO stage (p = 0.008). Using cox-regression analysis, HSP27 staining (p = 0.025), stage (p = 0.0012), and level of cytoreduction (p < 0.0001) were independent predictors of survival in these patients. CONCLUSION: Cox-regression analysis found HSP27 to be an independent indicator of prognosis and survival in patients with ovarian carcinoma who had longer follow-up. Decreased HSP27 staining was related to decreased survival. This study confirms the authors' earlier report on the importance of HSP27 as a prognostic indicator in ovarian carcinoma.