Differentiation in Male and Female Speech Styles

From the moment of their birth, a person's life is determined by their sex. Ms. Goroshko wants to know why this difference is so striking, why society is so concerned to sustain it, and how it is able to persist even when certain national or behavioural stereotypes are erased between people. She is convinced of the existence of not only social, but biological differences between men and women, and set herself the task, in a manuscript totalling 126 pages, written in Ukrainian and including extensive illustrations, of analysing these distinctions as they are manifested in language. She points out that, even before 1900, certain stylistic differences between the ways that men and women speak had been noted. Since then it has become possible, for instance in the case of Japanese, to point to examples of male and female sub-languages. In general, one can single out the following characteristics. Males tend to write with less fluency, to refer to events in a verb-phrase, to be time-oriented, to involve themselves more in their references to events, to locate events in their personal sphere of activity, and to refer less to others. Therefore, concludes Ms Goroshko, the male is shown to be more active, more ego-involved in what he does, and less concerned about others. Women, in contrast, were more fluent, referred to events in a noun-phrase, were less time-oriented, tended to be less involved in their event-references, locate events within their interactive community and refer more to others. They spent much more time discussing personal and domestic subjects, relationship problems, family, health and reproductive matters, weight, food and clothing, men, and other women. As regards discourse strategies, Ms Goroshko notes the following. Men more often begin a conversation, they make more utterances, these utterances are longer, they make more assertions, speak less carefully, generally determine the topic of conversation, speak more impersonally, use more vulgar expressions, and use fewer diminutives and more imperatives. Women's speech strategies, apart from being the opposite of those enumerated above, also contain more euphemisms, polite forms, apologies, laughter and crying. All of the above leads Ms. Goroshko to conclude that the differences between male and female speech forms are more striking than the similarities. Furthermore she is convinced that the biological divergence between the sexes is what generates the verbal divergence, and that social factors can only intensify or diminish the differentiation in verbal behaviour established by the sex of a person. Bearing all this in mind, Ms Goroshko set out to construct a grammar of male and female styles of speaking within Russian. One of her most important research tools was a certain type of free association test. She took a list comprising twelve stimuli (to love, to have, to speak, to fuck, a man, a woman, a child, the sky, a prayer, green, beautiful) and gave it to a group of participants specially selected, according to a preliminary psychological testing, for the high levels of masculinity or femininity they displayed. Preliminary responses revealed that the female reactions were more diverse than the male ones, there were more sentences and word combinations in the female reactions, men gave more negative responses to the stimulus and sometimes didn't want to react at all, women reacted more to adjectives and men to nouns, and that, surprisingly, women coloured more negatively their reactions to the words man, to love and a child (Ms. Goroshko is inclined to attribute this to the present economic situation in Russia). Another test performed by Ms. Goroshko was the so-called "defective text" developed by A.A. Brudny. All participants were distributed with packets of complete sentences, which had been taken from a text and then mixed at random. The task was to reconstruct the original text. There were three types of test, the first descriptive, the second narrative, and the third logical. Ms. Goroshko created computer programmes to analyse the results. She found that none of the reconstructed texts was coincident with the original, differing both from the original text and amongst themselves and that there were many more disparities in the male than the female texts. In the descriptive and logical texts the differences manifested themselves more clearly in the male texts, and in the narrative texts in the female texts. The widest dispersal of values was observed at the outset, while the female text ending was practically coincident with the original (in contrast to the male ending). The greatest differences in text reconstruction for both males and females were registered in the middle of the texts. Women, Ms. Goroshko claims, were more sensitive to the semantic structure of the texts, since they assembled the narrative text much more accurately than the other two, while the men assembled more accurately the logical text. Texts written by women were assembled more accurately by women and texts by men by men. On the basis of computer analysis, Ms. Goroshko found that female speech was substantially more emotional. It was expressed by various means, hyperbole, metaphor, comparisons, epithets, ways of enumeration, and with the aid of interjections, rhetorical questions, exclamations. The level of literacy was higher for female speech, and there were fewer mistakes in grammar and spelling in female texts. The last stage of Ms Goroshko's research concerned the social stereotypes of beliefs about men and women in Russian society today. A large number of respondents were asked questions such as "What merits must a woman possess?", "What are male vices and virtues?", etc. After statistical manipulation, an image of modern man and woman, as it exists in the minds of modern Russian men and women, emerged. Ms. Goroshko believes that her findings are significant not only within the field of linguistics. She has already successfully worked on anonymous texts and been able to decide on the sex of the author and consequently believes that in the future her research may even be of benefit to forensic science.

Nutrient-responsive mTOR signalling grows on Sterile ground

The control of cell growth, that is cell size, is largely controlled by mTOR (the mammalian target of rapamycin), a large serine/threonine protein kinase that regulates ribosome biogenesis and protein translation. mTOR activity is regulated both by the availability of growth factors, such as insulin/IGF-1 (insulin-like growth factor 1), and by nutrients, notably the supply of certain key amino acids. The last few years have seen a remarkable increase in our understanding of the canonical, growth factor-regulated pathway for mTOR activation, which is mediated by the class I PI3Ks (phosphoinositide 3-kinases), PKB (protein kinase B), TSC1/2 (the tuberous sclerosis complex) and the small GTPase, Rheb. However, the nutrient-responsive input into mTOR is important in its own right and is also required for maximal activation of mTOR signalling by growth factors. Despite this, the details of the nutrient-responsive signalling pathway(s) controlling mTOR have remained elusive, although recent studies have suggested a role for the class III PI3K hVps34. In this issue of the Biochemical Journal, Findlay et al. demonstrate that the protein kinase MAP4K3 [mitogen-activated protein kinase kinase kinase kinase-3, a Ste20 family protein kinase also known as GLK (germinal centre-like kinase)] is a new component of the nutrient-responsive pathway. MAP4K3 activity is stimulated by administration of amino acids, but not growth factors, and this is insensitive to rapamycin, most likely placing MAP4K3 upstream of mTOR. Indeed, MAP4K3 is required for phosphorylation of known mTOR targets such as S6K1 (S6 kinase 1), and overexpression of MAP4K3 promotes the rapamycin-sensitive phosphorylation of these same targets. Finally, knockdown of MAP4K3 levels causes a decrease in cell size. The results suggest that MAP4K3 is a new component in the nutrient-responsive pathway for mTOR activation and reveal a completely new function for MAP4K3 in promoting cell growth. Given that mTOR activity is frequently deregulated in cancer, there is much interest in new strategies for inhibition of this pathway. In this context, MAP4K3 looks like an attractive drug target since inhibitors of this enzyme should switch off mTOR, thereby inhibiting cell growth and proliferation, and promoting apoptosis.

Modulation of human CYP19A1 activity by mutant NADPH P450 oxidoreductase

Mutations in NADPH P450 oxidoreductase (POR) cause a broad spectrum of human disease with abnormalities in steroidogenesis. We have studied the impact of P450 reductase mutations on the activity of CYP19A1. POR supported CYP19A1 activity with a calculated Km of 126 nm for androstenedione and a Vmax of 1.7 pmol/min. Mutations R457H and V492E located in the FAD domain of POR that disrupt electron transfer caused a complete loss of CYP19A1 activity. The A287P mutation of POR decreased the activities of CYP17A1 by 60-80% but had normal CYP19A1 activity. Molecular modeling and protein docking studies suggested that A287P is involved in the interaction of POR:CYP17A1 but not in the POR:CYP19A1 interaction. Mutations C569Y and V608F in the NADPH binding domain of POR had 49 and 28% of activity of CYP19A1 compared with normal reductase and were more sensitive to the amount of NADPH available for supporting CYP19A1 activity. Substitution of NADH for NADPH had a higher impact on C569Y and V608F mutants of POR. Similar effects were obtained at low/high (5.5/8.5) pH, but using octanol to limit the flux of electrons from POR to CYP19A1 inhibited activity supported by all variants. High molar ratios of KCl also reduced the CYP19A1 supporting activities of C569Y and V608F mutants of POR to a greater extent compared to normal POR and A287P mutant. Because POR supports many P450s involved in steroidogenesis, bone formation, and drug metabolism, variations in the effects of POR mutations on specific enzyme activities may explain the broad clinical spectrum of POR deficiency.

Activities of ertapenem, a new long-acting carbapenem, against penicillin-sensitive or -resistant pneumococci in experimental meningitis

The penetration of ertapenem, a new carbapenem with a long half-life, reached 7.1 and 2.4% into inflamed and noninflamed meninges, respectively. Ertapenem had excellent antibacterial activity in the treatment of experimental meningitis due to penicillin-sensitive and -resistant pneumococci, leading to a decrease of 0.69 +/- 0.17 and 0.59 +/- 0.22 log(10) CFU/ml x h, respectively, in the viable cell counts in the cerebrospinal fluid. The efficacy of ertapenem was comparable to that of standard regimens (ceftriaxone monotherapy against the penicillin-sensitive strain and ceftriaxone combined with vancomycin against the penicillin-resistant strain). In vitro, ertapenem in concentrations above the MIC was highly bactericidal against both strains. Even against a penicillin- and quinolone-resistant mutant, ertapenem had similar bactericidal activity in vitro.

Evaluating Integration. Research contributions for a gender sensitive migration policy: Research contributions for a gender sensitive migration policy

The integration of academic and non-academic knowledge is a key concern for researchers who aim at bridging the gap between research and policy. Researchers involved in the sustainability-oriented NCCR North-South programme have made the experience that linking different types of knowledge requires time and effort, and that methodologies are still lacking. One programme component was created at the inception of this transdisciplinary research programme to support exchange between researchers, development practitioners and policymakers. After 8 years of research, the programme is assessing whether research has indeed enabled a continuous communication across and beyond academic boundaries and has effected changes in the public policies of poor countries. In a first review of the data, we selected two case studies explicitly addressing the lives of women. In both cases – one in Pakistan, the other in Nepal – the dialogue between researchers and development practitioners contributed to important policy changes for female migration. In both countries, outmigration has become an increasingly important livelihood strategy. National migration policies are gendered, limiting the international migration of women. In Nepal, women were not allowed to migrate to specific countries such as the Gulf States or Malaysia. This was done in the name of positive discrimination, to protect women from potential exploitation and harassment in domestic work. However, women continued to migrate in many other and often illegal and more risky ways, increasing their vulnerability. In Pakistan, female labour migration was not allowed at all and male migration increased the vulnerability of the families remaining back home. Researchers and development practitioners in Nepal and Pakistan brought women’s shared experience of and exposure to the mechanisms of male domination into the public debate, and addressed the discriminating laws. Now, for the first time in Pakistan, the new draft policy currently under discussion would enable broadly-based female labour migration. What can we learn from the two case studies with regard to ways of relating experience- and research-based knowledge? The paper offers insights into the sequence of interactions between researchers, local people, development practitioners, and policy-makers, which eventually contributed to the formulation of a rights-based migration policy. The reflection aims at exploring the gendered dimension of ways to co-produce and share knowledge for development across boundaries. Above all, it should help researchers to better tighten the links between the spheres of research and policy in future.

Diagnostic performance of high-sensitive troponin T in patients with renal insufficiency

In the present study, we wanted to (1) evaluate whether high-sensitive troponin T levels correlate with the grade of renal insufficiency and (2) test the accuracy of high-sensitive troponin T determination in patients with renal insufficiency for diagnosis of acute myocardial infarction (AMI). In this cross-sectional analysis, all patients who received serial measurements of high-sensitive troponin T from August 1, 2010, to October 31, 2012, at the Department of Emergency Medicine were included. We analyzed data on baseline characteristics, reason for referral, medication, cardiovascular risk factors, and outcome in terms of presence of AMI along with laboratory data (high-sensitive troponin T, creatinine). A total of 1,514 patients (67% male, aged 65 ± 16 years) were included, of which 382 patients (25%) had moderate to severe renal insufficiency and significantly higher levels of high-sensitive troponin T on admission (0.028 vs 0.009, p <0.0001). In patients without AMI, high-sensitive troponin T correlated inversely with the estimated glomerular filtration rate (R = -0.12, p <0.0001). Overall, sensitivity of an elevated high-sensitive troponin for diagnosis of AMI was 0.64 (0.56 to 0.71) and the specificity was 0.48 (0.45 to 0.51). The area under the curve of the receiver operating characteristic for all patients was 0.613 (standard error [SE] 0.023), whereas it was 0.741 (SE 0.029) for patients with a Modification of Diet in Renal Disease estimated glomerular filtration rate >60 ml/min presenting with acute chest pain or dyspnea and 0.535 (SE 0.056) for patients with moderate to severe renal insufficiency presenting with acute chest pain or dyspnea. In conclusion, the diagnostic accuracy for presence of AMI of a baseline measurement of high-sensitive troponin in patients with renal insufficiency was poor and resembles tossing a coin.

High-Sensitive Troponin Measurement in Emergency Department Patients Presenting with Syncope: A Retrospective Analysis

OBJECTIVE To study the relevance of high-sensitive troponin measurements in the acute workup in patients admitted to the emergency department of a large university hospital due to syncope. METHODS In this retrospective study all patients admitted to the emergency department because of syncope of the Inselspital, University Hospital Bern between 01 August 2010 and 31 October 2012, with serial determination of high-sensitive troponin (baseline and three hours control) were included. Of all identified patients we obtained data on demographics, laboratory data, ECG as well as on outcome. A change in high-sensitive troponin in the three hours control of +/-30% compared to baseline was considered significant. RESULTS A total of 121 patients with a mean age of 67 years (SD 16) were included in the study. 79 patients (65%) were male and 42 (35%) were female. There was no significant difference in the median high sensitive-troponin level at baseline and in the three hours control (0.01 mcg/L [0.003 to 0.022] versus 0.011 mcg/L [0.003 to 0.022], p = 0.47). Median percent change in high-sensitive troponin level between baseline and control was 0% (-9.1 to 5). 51 patients (42%) had elevated high-sensitive troponin levels at baseline with 7 patients (6%) showing a dynamic of +/-30% change from the baseline measurement in the 3 hours control. 3 of these patients received coronary angiography due to the dynamic in high-sensitive troponin, none of whom needed intervention for coronary revascularization. CONCLUSIONS On basis of the current study, where no single patient took benefit from determination of high-sensitive troponin, measurement of cardiac troponins should be reserved for patients with syncope presenting with symptoms suggestive for the presence of an acute cardiac syndrome.

Different dark conformations function in color-sensitive photosignaling by the sensory rhodopsin I-HtrI complex.

The haloarchaeal phototaxis receptor sensory rhodopsin I (SRI) in complex with its transducer HtrI delivers an attractant signal from excitation with an orange photon and a repellent signal from a second near-UV photon excitation. Using a proteoliposome system with purified SRI in complex with its transducer HtrI, we identified by site-directed fluorescence labeling a site (Ser(155)) on SRI that is conformationally active in signal relay to HtrI. Using site-directed spin labeling of Ser(155)Cys with a nitroxide side chain, we detected a change in conformation following one-photon excitation such that the spin probe exhibits a splitting of the outer hyperfine extrema (2A'(zz)) significantly smaller than that of the electron paramagnetic resonance spectrum in the dark state. The dark conformations of five mutant complexes that do not discriminate between orange and near-UV excitation show shifts to lower or higher 2A'(zz) values correlated with the alterations in their motility behavior to one- and two-photon stimuli. These data are interpreted in terms of a model in which the dark complex is populated by two conformers in the wild type, one that inhibits the CheA kinase (A) and the other that activates it (R), shifted in the dark by mutations and shifted in the wild-type SRI-HtrI complex in opposite directions by one-photon and two-photon reactions.

Construction of improved temperature-sensitive and mobilizable vectors and their use for constructing mutations in the adhesin-encoding acm gene of poorly transformable clinical Enterococcus faecium strains.

Inactivation by allelic exchange in clinical isolates of the emerging nosocomial pathogen Enterococcus faecium has been hindered by lack of efficient tools, and, in this study, transformation of clinical isolates was found to be particularly problematic. For this reason, a vector for allelic replacement (pTEX5500ts) was constructed that includes (i) the pWV01-based gram-positive repAts replication region, which is known to confer a high degree of temperature intolerance, (ii) Escherichia coli oriR from pUC18, (iii) two extended multiple-cloning sites located upstream and downstream of one of the marker genes for efficient cloning of flanking regions for double-crossover mutagenesis, (iv) transcriptional terminator sites to terminate undesired readthrough, and (v) a synthetic extended promoter region containing the cat gene for allelic exchange and a high-level gentamicin resistance gene, aph(2'')-Id, to distinguish double-crossover recombination, both of which are functional in gram-positive and gram-negative backgrounds. To demonstrate the functionality of this vector, the vector was used to construct an acm (encoding an adhesin to collagen from E. faecium) deletion mutant of a poorly transformable multidrug-resistant E. faecium endocarditis isolate, TX0082. The acm-deleted strain, TX6051 (TX0082Deltaacm), was shown to lack Acm on its surface, which resulted in the abolishment of the collagen adherence phenotype observed in TX0082. A mobilizable derivative (pTEX5501ts) that contains oriT of Tn916 to facilitate conjugative transfer from the transformable E. faecalis strain JH2Sm::Tn916 to E. faecium was also constructed. Using this vector, the acm gene of a nonelectroporable E. faecium wound isolate was successfully interrupted. Thus, pTEX5500ts and its mobilizable derivative demonstrated their roles as important tools by helping to create the first reported allelic replacement in E. faecium; the constructed this acm deletion mutant will be useful for assessing the role of acm in E. faecium pathogenesis using animal models.

Phenotypic and genotypic analysis of mouse hepatitis virus (JHM) complementation group A temperature-sensitive mutants

Temperature sensitive (ts) mutant viruses have helped elucidate replication processes in many viral systems. Several panels of replication-defective ts mutants in which viral RNA synthesis is abolished at the nonpermissive temperature (RNA$\sp{-})$ have been isolated for Mouse Hepatitis Virus, MHV (Robb et al., 1979; Koolen et al., 1983; Martin et al., 1988; Schaad et al., 1990). However, no one had investigated genetic or phenotypic relationships between these different mutant panels. In order to determine how the panel of MHV-JHM RNA$\sp{-}$ ts mutants (Robb et al., 1979) were genetically related to other described MHV RNA$\sp{-}$ ts mutants, the MHV-JHM mutants were tested for complementation with representatives from two different sets of MHV-A59 ts mutants (Koolen et al., 1983; Schaad et al., 1990). The three ts mutant panels together were found to comprise eight genetically distinct complementation groups. Of these eight complementation groups, three complementation classes are unique to their particular mutant panel; genetically equivalent mutants were not observed within the other two mutant panels. Two complementation groups were common to all three mutant panels. The three remaining complementation groups overlapped two of the three mutant sets. Mutants MHV-JHM tsA204 and MHV-A59 ts261 were shown to be within one of these overlapping complementation groups. The phenotype of the MHV-JHM mutants within this complementation class has been previously characterized (Leibowitz et al., 1982; Leibowitz et al, 1990). When these mutants were grown at the permissive temperature, then shifted up to the nonpermissive temperature at the start of RNA synthesis, genome-length RNA and leader RNA fragments accumulated, but no subgenomic mRNA was synthesized. MHV-A59 ts261 produced leader RNA fragments identical to those observed with MHV-JHM tsA204. Thus, these two MHV RNA$\sp{-}$ ts mutants that were genetically equivalent by complementation testing were phenotypically similar as well. Recombination frequencies obtained from crosses of MHV-A59 ts261 with several of the gene 1 MHV-A59 mutants indicated that the causal mutation(s) of MHV-A59 ts261 was located near the overlapping junction of ORF1a and ORF1b, in the 3$\sp\prime$ end of ORF1a, or the 5$\sp\prime$ end of ORF1b. Sequence analysis of this junction and 1400 nucleotides into the 5$\sp\prime$ end of ORF1b of MHV-A59 ts261 revealed one nucleotide change from the wildtype MHV-A59. This substitution at nucleotide 13,598 (A to G) was a silent mutation in the ORF1a reading frame, but resulted in an amino acid change in ORF1b gene product (I to V). This amino acid change would be expressed only in the readthrough translation product produced upon successful ribosome frameshifting. A revertant of MHV-A59 ts261 (R2) also retained this guanidine residue, but had a second substitution at nucleotide 14,475 in ORF1b. This mutation results in the substitution of valine for an isoleucine.^ The data presented here suggest that the mutation in MHV-A59 ts261 (nucleotide 13,598) would be responsible for the MHV-JHM complementation group A phenotype. A second-site reversion at nucleotide 14,475 may correct this defect in the revertant. Sequencing of gene 1 immediately upstream of nucleotide 13,296 and downstream of nucleotide 15,010 must be conducted to test this hypothesis. ^

18F-RB390: innovative ligand for imaging the T877A androgen receptor mutant in prostate cancer via positron emission tomography (PET).

BACKGROUND Detecting prostate cancer before spreading or predicting a favorable therapy are challenging issues for impacting patient's survival. Presently, 2-[(18) F]-fluoro-2-deoxy-D-glucose ((18) F-FDG) and/or (18) F-fluorocholine ((18) F-FCH) are the generally used PET-tracers in oncology yet do not emphasize the T877A androgen receptor (AR) mutation being exclusively present in cancerous tissue and escaping androgen deprivation treatment. METHODS We designed and synthesized fluorinated 5α-dihydrotestosterone (DHT) derivatives to target T877A-AR. We performed binding assays to select suitable candidates using COS-7 cells transfected with wild-type or T877A AR (WT-AR, T877A-AR) expressing plasmids and investigated cellular uptake of candidate (18) F-RB390. Stability, biodistribution analyses and PET-Imaging were assessed by injecting (18) F-RB390 (10MBq), with and without co-injection of an excess of unlabeled DHT in C4-2 and PC-3 tumor bearing male SCID mice (n = 12). RESULTS RB390 presented a higher relative binding affinity (RBA) (28.1%, IC50  = 32 nM) for T877A-AR than for WT-AR (1.7%, IC50  = 357 nM) related to DHT (RBA = 100%). A small fraction of (18) F-RB390 was metabolized when incubated with murine liver homogenate or human blood for 3 hr. The metabolite of RB390, 3-hydroxysteroid RB448, presented similar binding characteristics as RB390. (18) F-RB390 but not (18) F-FDG or (18) F-FCH accumulated 2.5× more in COS-7 cells transfected with pSG5AR-T877A than with control plasmid. Accumulation was reduced with an excess of DHT. PET/CT imaging and biodistribution studies revealed a significantly higher uptake of (18) F-RB390 in T877A mutation positive xenografts compared to PC-3 control tumors. This effect was blunted with DHT. CONCLUSION Given the differential binding capacity and the favorable radioactivity pattern, (18) F-RB390 represents the portrayal of the first imaging ligand with predictive potential for mutant T877A-AR in prostate cancer for guiding therapy. Prostate 75:348-359, 2015. © 2014 Wiley Periodicals, Inc.

Map positions of third chromosomal female sterile and lethal mutations of Drosophila melanogaster

Chromosomal mutations induced by ethyl methanesulfonate (EMS) treatment can cause female sterility or maternal-effect lethality in Drosophila. EMS is particularly useful to researchers because it creates mutations independent of position effects. However, because researchers have little control over the chromosomal site of mutation, post-mutagenic genetic mapping is required to determine the cytological location of the mutation. To make a valuable set of mutants more useful to the research community, we have mapped the uncharacterized part of the female-sterile – maternal-effect lethal Tübingen collection. We mapped 49 female-sterile – maternal-effect lethal alleles and 72 lethal alleles to individual deficiency intervals on the third chromosome. In addition, we analyzed the phenotype of ovaries resulting from female sterile mutations. The observed phenotypes range from tumorous ovaries and early blocks in oogenesis, to later blocks, slow growth, blocks in stage 10, to apparently full development of the ovary. The mapping and phenotypic characterization of these 121 mutations provide the necessary information for the researcher to consider a specific mutant as a candidate for their gene of interest.Key words: Drosophila melanogaster, oogenesis, female sterile, maternal-effect lethal, EMS-induced mutations.

Heterotrimeric G protein -mediated signal transduction in Neurospora crassa

Heterotrimeric G protein-mediated signal transduction is one of numerous means that cells utilize to respond to external stimuli. G proteins consist of α, β andγ subunits. Extracellular ligands bind to seven-transmembrane helix receptors, triggering conformational changes. This is followed by activation of coupled G proteins through the exchange of GDP for GTP on the Gα subunit. Once activated, Gα-GTP dissociates from the βγ dimer. Both of these two moieties can interact with downstream effectors, such as adenylyl cyclase, phospholipase C, phosphodiesterases, or ion channels, leading to a series of changes in cellular metabolism and physiology. ^ Neurospora crassa is a eukaryotic multicellular filamentous fungus, with asexual/vegetative and sexual phases to its life cycle. Three Gα (GNA-1, GNA-2, GNA-3) and one Gβ (GNB-1) proteins have been identified in this organism. This dissertation investigates GNA-1 and GNB-1 mediated signaling pathways in N. crassa. ^ GNA-1 was the first identified microbial Gα that belongs to a mammalian superfamily (Gαi). Deletion of GNA-1 leads to multiple defects in N. crassa. During the asexual cycle, Δgna-1 strains display a slower growth rate and delayed conidiation on solid medium. In the sexual cycle, the Δgna-1 mutant is male-fertile but female-sterile. Biochemical studies have shown that Δ gna-1 strains have lower adenosine 3′–5 ′ cyclic monophosphate (cAMP) levels than wild type under conditions where phenotypic defects are observed. In this thesis work, strains containing one of two GTPase-deficient gna-1 alleles (gna-1 R178C, gna-1Q204L) leading to constitutive activation of GNA-1 have been constructed and characterized. Activation of GNA-1 causes uncontrolled aerial hyphae proliferation, elevated sensitivity to heat and oxidative stresses, and lower carotenoid synthesis. To further study the function of GNA-1, constructs to enable expression of mammalian Gαi superfamily members were transformed into a Δ gna-1 strain, and complementation of Δgna-1 defects investigated. Gαs, which is not a member of Gα i superfamily was used as a control. These mammalian Gα genes were able to rescue the vegetative growth rate defect of the Δ gna-1 strain in the following order: Gαz > Gα o > Gαs > Gαt > Gαi. In contrast, only Gαo was able to complement the sexual defect of a Δgna-1 strain. With regard to the thermotolerance phenotype, none of the mammalian Gα genes restored the sensitivity to a wild type level. These results suggest that GNA-1 regulates two independent pathways during the vegetative and sexual cycles in N. crassa. ^ GNB-1, a G protein β subunit from N. crassa, was identified and its functions investigated in this thesis work. The sequence of the gnb-1 gene predicts a polypeptide of 358 residues with a molecular mass of 39.7 kDa. GNB-1 exhibits 91% identity to Cryphonectria parasitica CPGB-1, and also displays significant homology with human and Dictyostelium Gβ genes (∼66%). A Δ gnb-1 strain was constructed and shown to exhibit defects in asexual spore germination, vacuole number and size, mass accumulation and female fertility. A novel role for GNB-1 in regulation of GNA-1 and GNA-2 protein levels was also demonstrated. ^

The effect of elevated CO2 and increased temperature on in vitro fertilization success and initial embryonic development of single male:female crosses of broad-cast spawning corals at mid- and high-latitude locations

The impact of global climate change on coral reefs is expected to be most profound at the sea surface, where fertilization and embryonic development of broadcast-spawning corals takes place. We examined the effect of increased temperature and elevated CO2 levels on the in vitro fertilization success and initial embryonic development of broadcast-spawning corals using a single male:female cross of three different species from mid- and high-latitude locations: Lyudao, Taiwan (22° N) and Kochi, Japan (32° N). Eggs were fertilized under ambient conditions (27 °C and 500 µatm CO2) and under conditions predicted for 2100 (IPCC worst case scenario, 31 °C and 1000 µatm CO2). Fertilization success, abnormal development and early developmental success were determined for each sample. Increased temperature had a more profound influence than elevated CO2. In most cases, near-future warming caused a significant drop in early developmental success as a result of decreased fertilization success and/or increased abnormal development. The embryonic development of the male:female cross of A. hyacinthus from the high-latitude location was more sensitive to the increased temperature (+4 °C) than the male:female cross of A. hyacinthus from the mid-latitude location. The response to the elevated CO2 level was small and highly variable, ranging from positive to negative responses. These results suggest that global warming is a more significant and universal stressor than ocean acidification on the early embryonic development of corals from mid- and high-latitude locations.

(Table 1) Breeding parameters for 60 male Adélie penguins (Pygoscelis adeliae) at station Dumont d'Urville (Antarctica) depending on treatment (control vs. CORT)

Corticosterone, the main stress hormone in birds, mediates resource allocation, allowing animals to adjust their physiology and behaviour to changes in the environment. Incubation is a time and energy-consuming phase of the avian reproductive cycle. It may be terminated prematurely, when the parents' energy stores are depleted or when environmental conditions are severe. In this study, the effects of experimentally elevated baseline corticosterone levels on the parental investment of incubating male Adelie penguins were investigated. Incubation duration and reproductive success of 60 penguins were recorded. The clutches of some birds were replaced by dummy eggs, which recorded egg temperatures and rotation rates, enabling a detailed investigation of incubation behaviour. Corticosterone levels of treated birds were 2.4-fold higher than those of controls 18 days post treatment. Exogenous corticosterone triggered nest desertion in 61% of the treated birds; consequently reducing reproductive success, indicating that corticosterone can reduce or disrupt parental investment. Regarding egg temperatures, hypothermic events became more frequent and more pronounced in treated birds, before these birds eventually abandoned their nest. The treatment also significantly decreased incubation temperatures by 1.3 °C and lengthened the incubation period by 2.1 days. However, the number of chicks at hatching was similar among successful nests, regardless of treatment. Weather conditions appeared to be particularly important in determining the extent to which corticosterone levels affected the behaviour of penguins, as treated penguins were more sensitive to severe weather conditions. This underlines the importance of considering the interactions of organisms with their environment in studies of animal behaviour and ecophysiology.