Phosphorus and carbon availability regulate structural composition and complexity of AM fungal mycelium

The regulation of the structural composition and complexity of the mycelium of arbuscular mycorrhizal (AM) fungi is not well understood due to their obligate biotrophic nature. The aim of this study was to investigate the structure of extraradical mycelium at high and low availability of carbon (C) to the roots and phosphorus (P) to the fungus. We used monoxenic cultures of the AM fungus Rhizophagus irregularis (formerly Glomus intraradices) with transformed carrot roots as the host in a cultivation system including a root-free compartment into which the extraradical mycelium could grow. We found that high C availability increased hyphal length and spore production and anastomosis formation within individual mycelia. High P availability increased the formation of branched absorbing structures and reduced spore production and the overall length of runner hyphae. The complexity of the mycelium, as indicated by its fractal dimensions, increased with both high C and P availability. The results indicate that low P availability induces a growth pattern that reflects foraging for both P and C. Low C availability to AM roots could still support the explorative development of the mycelium when P availability was low. These findings help us to better understand the development of AM fungi in ecosystems with high P input and/or when plants are subjected to shading, grazing or any management practice that reduces the photosynthetic ability of the plant.     

A new concept of olfactory biosensor based on interdigitated microelectrodes and immobilized yeasts expressing the human receptor OR17-40

This work shows the feasibility of an olfactory biosensor based on the immobilization of Saccharomyces cerevisiae yeast cells genetically modified to express the human olfactory receptor OR17-40 onto interdigitated microconductometric electrodes. This olfactory biosensor has been applied to the detection of its specific odorant (helional) with a high sensitivity (threshold 10⁻¹⁴ M). In contrast, no significant response was observed using a non-specific odorant (heptanal), which suggests a good selectivity. Thus, this work may represent a first step towards a new kind of bioelectronic noses based on whole yeast cells and allowing a real time monitoring of olfactory receptor activation. Presented at the joint biannual meeting of the SFB-GEIMM-GRIP, Anglet, France, 14–19 October, 2006.     

RasGAP Shields Akt from Deactivating Phosphatases in Fibroblast Growth Factor Signaling but Loses This Ability Once Cleaved by Caspase-3

Fibroblast growth factor receptors (FGFRs) are involved in proliferative and differentiation physiological responses. Deregulation of FGFR-mediated signaling involving the Ras/PI3K/Akt and the Ras/Raf/ERK MAPK pathways is causally involved in the development of several cancers. The caspase-3/p120 RasGAP module is a stress sensor switch. Under mild stress conditions, RasGAP is cleaved by caspase-3 at position 455. The resulting N-terminal fragment, called fragment N, stimulates anti-death signaling. When caspase-3 activity further increases, fragment N is cleaved at position 157. This generates a fragment, called N2, that no longer protects cells. Here, we investigated in Xenopus oocytes the impact of RasGAP and its fragments on FGF1-mediated signaling during G₂/M cell cycle transition. RasGAP used its N-terminal Src homology 2 domain to bind FGFR once stimulated by FGF1, and this was necessary for the recruitment of Akt to the FGFR complex. Fragment N, which did not associate with the FGFR complex, favored FGF1-induced ERK stimulation, leading to accelerated G₂/M transition. In contrast, fragment N2 bound the FGFR, and this inhibited mTORC2-dependent Akt Ser-473 phosphorylation and ERK2 phosphorylation but not phosphorylation of Akt on Thr-308. This also blocked cell cycle progression. Inhibition of Akt Ser-473 phosphorylation and entry into G₂/M was relieved by PHLPP phosphatase inhibition. Hence, full-length RasGAP favors Akt activity by shielding it from deactivating phosphatases. This shielding was abrogated by fragment N2. These results highlight the role played by RasGAP in FGFR signaling and how graded stress intensities, by generating different RasGAP fragments, can positively or negatively impact this signaling.     

Age-Related Disparity in Immediate Prognosis of Patients with Triple-Negative Breast Cancer: A Population-Based Study from SEER Cancer Registries

Background

Triple-negative breast cancer (TNBC) has been demonstrated to carry poor prognosis, but whether or not there exists any age-related variation in TNBC outcomes has yet to be elucidated. The current population-based study investigated the early survival pattern of elderly women with TNBC and identified outcome-correlated factors.

Patients and Methods

We searched the Surveillance, Epidemiology, and End Results (SEER) database and enrolled female primary non-metastatic TNBC cases. The patients were subdivided into elderly (≥70 years) and young groups (<70 years). The survival status of elderly patients was compared to that of the younger women. The primary and secondary endpoints were cancer-specific survival (CSS) and overall survival (OS) respectively.

Results

9908 female TNBC patients diagnosed from 2010 to 2011 were included in the current study (20.4% elderly). Elderly patients with relatively advanced diseases exhibited distinctly worse cancer-specific (log-rank, p<0.001) and overall survival (log-rank, p<0.001) than their young counterparts. Advanced age at diagnosis (≥70 years) was significantly predictive of poor outcome in terms of CSS (hazard ratio (HR), 2.125; 95% confidence interval (CI), 1.664 to 2.713; p<0.001) and OS (HR, 3.042; 95%CI, 2.474 to 3.740; p<0.001). Underuse of curative treatment especially radiotherapy was more prevalent in elderly women with stage II or III diseases than in younger patients.

Conclusion

Elderly patients with TNBC displayed elevated early mortality within the first two years of diagnosis compared to the younger individuals. The observed lower rate of loco-regional treatment might be associated with worse cancer-specific outcome for these patients.     

TLR2 and TLR4 Mediate Differential Responses to Limb Ischemia through MyD88-Dependent and Independent Pathways

Introduction

The danger signal HMGB1 is released from ischemic myocytes, and mediates angiogenesis in the setting of hindlimb ischemia. HMGB1 is a ligand for innate immune receptors TLR2 and TLR4. While both TLR2 and TLR4 signal through myeloid differentiation factor 88 (MyD88), TLR4 also uniquely signals through TIR-domain-containing adapter-inducing interferon-β (TRIF). We hypothesize that TLR2 and TLR4 mediate ischemic myocyte regeneration and angiogenesis in a manner that is dependent on MyD88 signaling.

Methods

Mice deficient in TLR2, TLR4, MyD88 and TRIF underwent femoral artery ligation in the right hindlimb. Laser Doppler perfusion imaging was used to assess the initial degree of ischemia and the extent of perfusion recovery. Muscle regeneration, necrosis and fat replacement at 2 weeks post-ligation were assessed histologically and vascular density was quantified by immunostaining. In vitro, endothelial tube formation was evaluated in matrigel in the setting of TLR2 and TLR4 antagonism.

Results

While control and TLR4 KO mice demonstrated prominent muscle regeneration, both TLR2 KO and TRIF KO mice exhibited marked necrosis with significant inflammatory cell infiltrate. However, MyD88 KO mice had a minimal response to the ischemic insult with little evidence of injury. This observation could not be explained by differences in perfusion recovery which was similar at two weeks in all the strains of mice. TLR2 KO mice demonstrated abnormal vessel morphology compared to other strains and impaired tube formation in vitro.

Discussion

TLR2 and TRIF signaling are necessary for muscle regeneration after ischemia while MyD88 may instead mediate muscle injury. The absence of TLR4 did not affect muscle responses to ischemia. TLR4 may mediate inflammatory responses through MyD88 that are exaggerated in the absence of TLR2. Additionally, the actions of TLR4 through TRIF may promote regenerative responses that are required for recovery from muscle ischemia.     

The Implication of Early Chromatin Changes in X Chromosome Inactivation

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Progressive defects observed in mouse sperm during the course of three generations of selenium deficiency

Three successive generations of mice were fed a Torula yeast based Se-deficient diet with or without 0.1 ppm Se in the drinking water. The Se-deficient mice, in the course of three generations, showed a decrease in body weight, testis weight, epididymal weight, and sperm production. The percentage of morphologically abnormal sperm increased in successive generations. The majority of sperm defects were found in the midpiece region of the tail. Many of these aberrant sperm were motile. A progressive decrease in fertility was noted during the first two generations of Se deficiency. This system confirms the essential role of Se in spermatogenesis and provides a model for the evaluation of the primary effect of Se deprivation on the structural development of sperm.     

Synergistic activation of ENaC by three membrane-bound channel-activating serine proteases (mCAP1, mCAP2, and mCAP3) and serum- and glucocorticoid-regulated kinase (Sgk1) in Xenopus Oocytes

Sodium balance is maintained by the precise regulation of the activity of the epithelial sodium channel (ENaC) in the kidney. We have recently reported an extracellular activation of ENaC-mediated sodium transport (I(Na)) by a GPI-anchored serine protease (mouse channel-activating protein, mCAP1) that was isolated from a cortical collecting duct cell line derived from mouse kidney. In the present study, we have identified two additional membrane-bound serine proteases (mCAP2 and mCAP3) that are expressed in the same cell line. We show that each of these proteases is able to increase I(Na) 6-10-fold in the Xenopus oocyte expression system. I(Na) and the number (N) of channels expressed at the cell surface (measured by binding of a FLAG monoclonal I(125)-radioiodinated antibody) were measured in the same oocyte. Using this assay, we show that mCAP1 increases I(Na) 10-fold (P < 0.001) but N remained unchanged (P = 0.9), indicating that mCAP1 regulates ENaC activity by increasing its average open probability of the whole cell (wcP(o)). The serum- and glucocorticoid-regulated kinase (Sgk1) involved in the aldosterone-dependent signaling cascade enhances I(Na) by 2.5-fold (P < 0.001) and N by 1.6-fold (P < 0.001), indicating a dual effect on N and wcP(o). Compared with Sgk1 alone, coexpression of Sgk1 with mCAP1 leads to a ninefold increase in I(Na) (P < 0.001) and 1.3-fold in N (P < 0.02). Similar results were observed for mCAP2 and mCAP3. The synergism between CAPs and Sgk1 on I(Na) was always more than additive, indicating a true potentiation. The synergistic effect of the two activation pathways allows a large dynamic range for ENaC-mediated sodium regulation crucial for a tight control of sodium homeostasis.     

Reactive oxygen species in the elongation zone of maize leaves are necessary for leaf extension

The production and role of reactive oxygen species (ROS) in the expanding zone of maize (Zea mays) leaf blades were investigated. ROS release along the leaf blade was evaluated by embedding intact seedlings in 2',7'-dichlorofluorescein-containing agar and examining the distribution of 2',7'-dichlorofluorescein fluorescence along leaf 4, which was exposed by removing the outer leaves before embedding the seedling. Fluorescence was high in the expanding region, becoming practically non-detectable beyond 65 mm from the ligule, indicating high ROS production in the expansion zone. Segments obtained from the elongation zone of leaf 4 were used to assess the role of ROS in leaf elongation. The distribution of cerium perhydroxide deposits in electron micrographs indicated hydrogen peroxide (H(2)O(2)) presence in the apoplast. 2',7'-Dichlorofluorescein fluorescence and apoplastic H(2)O(2) accumulation were inhibited with diphenyleneiodonium (DPI), which also inhibited O*(2)(-) generation, suggesting a flavin-containing enzyme activity such as NADPH oxidase was involved in ROS production. Segments from the elongation zone incubated in water grew 8% in 2 h. KI treatments, which scavenged H(2)O(2) but did not inhibit O*(2)(-) production, did not modify growth. DPI significantly inhibited segment elongation, and the addition of H(2)O(2) (50 or 500 microM) to the incubation medium partially reverted the inhibition caused by DPI. These results indicate that a certain concentration of H(2)O(2) is necessary for leaf elongation, but it could not be distinguished whether H(2)O(2), or other ROS, are the actual active agents.