Influence of the propagation strategy for obtaining robust Saccharomyces cerevisiae cells that efficiently co-ferment xylose and glucose in lignocellulosic hydrolysates

Development of xylose-fermenting yeast strains that are tolerant to the inhibitors present in lignocellulosic hydrolysates is crucial to achieve efficient bioethanol production processes. In this study, the importance of the propagation strategy for obtaining robust cells was studied. Addition of hydrolysate during propagation of the cells adapted them to the inhibitors, resulting in more tolerant cells with shorter lag phases and higher specific growth rates in minimal medium containing acetic acid and vanillin than unadapted cells. Addition of hydrolysate during propagation also resulted in cells with better fermentation capabilities. Cells propagated without hydrolysate were unable to consume xylose in wheat straw hydrolysate fermentations, whereas 40.3% and 97.7% of the xylose was consumed when 12% and 23% (v/v) hydrolysate, respectively, was added during propagation. Quantitative polymerase chain reaction revealed changes in gene expression, depending on the concentration of hydrolysate added during propagation. This study highlights the importance of using an appropriate propagation strategy for the optimum performance of yeast in fermentation of lignocellulosic hydrolysates.     

Role of ERK1/2 in platelet lysate‐driven endothelial cell repair

Mechanisms of endothelial repair induced by a platelet lysate (PL) were studied on human (HuVEC, HMVEC‐c) and non‐human (PAOEC, bEnd5) endothelial cells. A first set of analyses on these cells showed that 20% (v/v) PL promotes scratch wound healing, with a maximum effect on HuVEC. Further analyses made on HuVEC showed that the ERK inhibitor PD98059 maximally inhibited the PL‐induced endothelial repair, followed in order of importance by the calcium chelator BAPTA‐AM, the PI3K inhibitor wortmannin and the p38 inhibitor SB203580. The PL exerted a chemotactic effect on HuVEC, which was abolished by all the above inhibitors, and induced a PD98059‐sensitive increase of cell proliferation rate. Confocal calcium imaging of fluo‐3‐loaded HuVEC showed that PL was able to induce cytosolic free Ca²⁺ oscillations, visible also in Ca²⁺‐free medium, suggesting an involvement of Ins3P‐dependent Ca²⁺ release. Western blot analysis on scratch wounded HuVEC showed that PL induced no activation of p38, a transient activation of AKT, and a sustained activation of ERK1/2. The complex of data indicates that, although different signalling pathways are involved in PL‐promoted endothelial repair, the process is chiefly under the control of ERK1/2. J. Cell. Biochem. 110: 783–793, 2010. © 2010 Wiley‐Liss, Inc.     

Seasonal variability of detoxificant response and heavy metal accumulation in tissues of both sexes in <Emphasis Type="Italic">Tinca tinca</Emphasis> (L.) from Lake Trasimeno

Tinca tinca were sampled seasonally from Lake Trasimeno in order to investigate the influence of heavy metal accumulation and changing environmental parameters on the antioxidant responses in this species. Liver, gills and kidney of both sexes were analyzed for total glutathione, catalase, glutathione peroxidase, glutathione reductase, glutathione S-transferase, glyoxalase I, glyoxalase II, while muscle was analyzed for arsenic, lead, cadmium and mercury. The MRLs (maximum residue limits) set by actual regulation were not exceeded for heavy metals, but a seasonal and sex-linked variability of antioxidant parameters was observed, and major variations occurred especially in summer and autumn. In this shallow lake, heavy metal accumulation did not affect the biochemical variations, which are rather due to the decreased water level inducing hyperoxia and hypoxia within 24 h, high temperature and loss of mating and refuge areas.     

Arabidopsis natural variation in insect egg-induced cell death reveals a role for LECTIN RECEPTOR KINASE-I.1

In Arabidopsis (Arabidopsis thaliana), a hypersensitive-like response (HR-like response) is triggered underneath the eggs of the large white butterfly Pieris brassicae (P. brassicae), and this response is dependent on salicylic acid (SA) accumulation and signaling. Previous reports indicate that the clade I L-type LECTIN RECEPTOR KINASE-I.8 (LecRK-I.8) is involved in early steps of egg recognition. A genome-wide association study was used to better characterize the genetic structure of the HR-like response and discover loci that contribute to this response. We report here the identification of LecRK-I.1, a close homolog of LecRK-I.8, and show that two main haplotypes that explain part of the variation in HR-like response segregate among natural Arabidopsis accessions. Besides, signatures of balancing selection at this locus suggest that it may be ecologically important. Disruption of LecRK-I.1 results in decreased HR-like response and SA signaling, indicating that this protein is important for the observed responses. Furthermore, we provide evidence that LecRK-I.1 functions in the same signaling pathway as LecRK-I.8. Altogether, our results show that the response to eggs of P. brassicae is controlled by multiple LecRKs.

A cell-surface receptor controls natural variation of egg-induced cell death in Arabidopsis.     

Microtubule assembly in Friend erythroleukemia cells spread on fibronectin- and lectin-coated substrates

A comparison was carried out between parental Friend Erythroleukemia cells (FLC, 745 A clone) and highly fibronectin (FN)-sensitive clones of FLC for their ability to adhere, spread and organize microtubular (MT) apparatus, when seeded on FN- or lectin-coated plastic substrates. While FN was able to induce the spreading only in the FN-sensitive FLC clones (further referred to as FF clones) and not in the parental 745 A cells, the lectins Concanavalin A (ConA) and Leukoagglutinin (LeuA) promoted the spreading of both 745 A and FF cells, with no differences between the two cell lines. Wheat germ agglutinin (WGA), instead, is almost ineffective in triggering cell spreading in both cell clones. The spreading of FLC, either 745 A or FF, on any of the ligands tested, is always accompanied by a massive reorganization of the MT apparatus of the cell. Possible mechanisms involved in the selective spreading effect, exerted by FN, are discussed.     

Interaction between Notch signalling and Lunatic fringe during somite boundary formation in the mouse.

BACKGROUND: The process of somitogenesis can be divided into three major events: the prepatterning of the mesoderm; the formation of boundaries between the prospective somites; and the cellular differentiation of the somites. Expression and functional studies have demonstrated the involvement of the murine Notch pathway in somitogenesis, although its precise role in this process is not yet well understood. We examined the effect of mutations in the Notch pathway elements Delta like 1 (Dll1), Notch1 and RBPJkappa on genes expressed in the presomitic mesoderm (PSM) and have defined the spatial relationships of Notch pathway gene expression in this region. RESULTS: We have shown that expression of Notch pathway genes in the PSM overlaps in the region where the boundary between the posterior and anterior halves of two consecutive somites will form. The Dll1, Notch1 and RBPJkappa mutations disrupt the expression of Lunatic fringe (L-fng), Jagged1, Mesp1, Mesp2 and Hes5 in the PSM. Furthermore, expression of EphA4, mCer 1 and uncx4.1, markers for the anterior-posterior subdivisions of the somites, is down-regulated to different extents in Notch pathway mutants, indicating a global alteration of pattern in the PSM. CONCLUSIONS: We propose a model for the mechanism of somite border formation in which the activity of Notch in the PSM is restricted by L-fng to a boundary-forming territory in the posterior half of the prospective somite. In this region, Notch function activates a set of genes that are involved in boundary formation and anterior-posterior somite identity.     

Scratch wound closure of C2C12 mouse myoblasts is enhanced by human platelet lysate

The effect of a platelet lysate (PL) on muscle wound healing, based on in vitro scratch wound of C2C12 mouse myoblasts, has been investigated. Cell viability assays show that PL induced an increase in cell proliferation at concentrations of 1-20%, but was slightly cytotoxic at 100%. PL promoted wound closure after scratch wounding of cell monolayers. The p38 inhibitor SB203580 and the PI3K inhibitor, wortmannin, decreased the PL effect, whereas the ERK inhibitor, PD98059, did not. Transwell migration of cells was also increased by PL, and although SB203580 abrogated this effect, wortmannin reduced it, whereas PD98059 was ineffective. Western blot analyses of scratch wounded cells showed activation of AKT and p38, while in the presence of PL there was a faster and sustained activation of AKT and p38 (up to 6 h), and a transient activation of ERK1/2. Taken together, the data show that PL promotes C2C12 wound healing by enhancing cell proliferation and motility.