Leaf Eh and pH: A Novel Indicator of Plant Stress. Spatial, Temporal and Genotypic Variability in Rice (Oryza sativa L.)

A wealth of knowledge has been published in the last decade on redox regulations in plants. However, these works remained largely at cellular and organelle levels. Simple indicators of oxidative stress at the plant level are still missing. We developed a method for direct measurement of leaf Eh and pH, which revealed spatial, temporal, and genotypic variations in rice. Eh (redox potential) and Eh@pH7 (redox potential corrected to pH 7) of the last fully expanded leaf decreased after sunrise. Leaf Eh was high in the youngest leaf and in the oldest leaves, and minimum for the last fully expanded leaf. Leaf pH decreased from youngest to oldest leaves. The same gradients in Eh-pH were measured for various varieties, hydric conditions, and cropping seasons. Rice varieties differed in Eh, pH, and/or Eh@pH7. Leaf Eh increases and leaf pH decreases with plant age. These patterns and dynamics in leaf Eh-pH are in accordance with the pattern and dynamics of disease infections. Leaf Eh-pH can bring new insight on redox processes at plant level and is proposed as a novel indicator of plant stress/health. It could be used by agronomists, breeders, and pathologists to accelerate the development of crop cultivation methods leading to agroecological crop protection

The SECURE project – Stem canker of oilseed rape: : molecular methods and mathematical modelling to deploy durable resistance

N Evans et al, "The SECURE Project - Stem Canker of oilseed rape: Molecular methods and mathematical modeling to deploy durable resistance", in Vol 4 of the Proceedings of the 12th International Rapeseed Congress : Sustainable Development in Cruciferous Oilseed Crops Production, Wuhan, China, March 26 - 30, 2007. The proceedings are available online at: http://gcirc.org/intranet/irc-proceedings/12th-irc-wuhan-china-2007-vol-4.htmlModelling done during the SECURE project has demonstrated the dynamic nature of the interaction between phoma stem canker (Leptosphaeria maculans), the oilseed rape host (Brassica napus) and the environment. Experiments done with near-isogenic lines of L. maculans to investigate pathogen fitness support field data that suggest a positive effect of the avirulence allele AvrLm4 on pathogen fitness, and that the loss of this allele renders isolates less competitive under field conditions on cultivars without the resistance gene Rlm4. The highlight of molecular work was the cloning of AvrLm1 and AvrLm6. L. maculans is now one of the few fungal species for which two avirulence loci have been cloned. Subsequent research focused on understanding the function of AvrLm1 and AvrLm6 and on the analysis of sequences of virulent isolates to understand molecular evolution towards virulence. Isolates of L. maculans transformed with GFP and/or DsRed were used to follow growth of the fungus in B. napus near-isogenic-lines (NIL) with or without MX (Rlm6) resistance under different temperature and wetness conditions. The results greatly enhanced our knowledge of the infection process and the rate and extent of in planta growth on different cultivars. Conclusions from work to model durability of resistance have been tested under field conditions through a series of experiments to compare durability of resistance conferred by the major resistance gene Rlm6 alone in a susceptible background (EurolMX) or in a resistant background (DarmorMX) under recurrent selection over 4 growing seasons. A major priority of the project was knowledge transfer of results and recommendations to target audiences such as plant breeding companies and extension services. CETIOM developed a “diversification scheme” that encourages French growers to make an informed choice about the cultivars that are grown within the rotation based on the resistance genes carried by the individual cultivars. Use of such schemes, in association with survey data on the population structure of L. maculans at both national and European scales will provide opportunities for breeders and the industry to manage available B. napus resistance more effectively.Non peer reviewe

Distinct tau prion strains propagate in cells and mice and define different tauopathies

Prion-like propagation of tau aggregation might underlie the stereotyped progression of neurodegenerative tauopathies. True prions stably maintain unique conformations (“strains”) in vivo that link structure to patterns of pathology. We now find that tau meets this criterion. Stably expressed tau repeat domain indefinitely propagates distinct amyloid conformations in a clonal fashion in culture. Reintroduction of tau from these lines into naive cells reestablishes identical clones. We produced two strains in vitro that induce distinct pathologies in vivo as determined by successive inoculations into three generations of transgenic mice. Immunopurified tau from these mice recreates the original strains in culture. We used the cell system to isolate tau strains from 29 patients with 5 different tauopathies, finding that different diseases are associated with different sets of strains. Tau thus demonstrates essential characteristics of a prion. This might explain the phenotypic diversity of tauopathies and could enable more effective diagnosis and therapy

Separation of DNA Replication from the Assembly of Break-Competent Meiotic Chromosomes

The meiotic cell division reduces the chromosome number from diploid to haploid to form gametes for sexual reproduction. Although much progress has been made in understanding meiotic recombination and the two meiotic divisions, the processes leading up to recombination, including the prolonged pre-meiotic S phase (meiS) and the assembly of meiotic chromosome axes, remain poorly defined. We have used genome-wide approaches in Saccharomyces cerevisiae to measure the kinetics of pre-meiotic DNA replication and to investigate the interdependencies between replication and axis formation. We found that replication initiation was delayed for a large number of origins in meiS compared to mitosis and that meiotic cells were far more sensitive to replication inhibition, most likely due to the starvation conditions required for meiotic induction. Moreover, replication initiation was delayed even in the absence of chromosome axes, indicating replication timing is independent of the process of axis assembly. Finally, we found that cells were able to install axis components and initiate recombination on unreplicated DNA. Thus, although pre-meiotic DNA replication and meiotic chromosome axis formation occur concurrently, they are not strictly coupled. The functional separation of these processes reveals a modular method of building meiotic chromosomes and predicts that any crosstalk between these modules must occur through superimposed regulatory mechanisms

Characterization of an Nmr Homolog That Modulates GATA Factor-Mediated Nitrogen Metabolite Repression in Cryptococcus neoformans

Nitrogen source utilization plays a critical role in fungal development, secondary metabolite production and pathogenesis. In both the Ascomycota and Basidiomycota, GATA transcription factors globally activate the expression of catabolic enzyme-encoding genes required to degrade complex nitrogenous compounds. However, in the presence of preferred nitrogen sources such as ammonium, GATA factor activity is inhibited in some species through interaction with co-repressor Nmr proteins. This regulatory phenomenon, nitrogen metabolite repression, enables preferential utilization of readily assimilated nitrogen sources. In the basidiomycete pathogen Cryptococcus neoformans, the GATA factor Gat1/Are1 has been co-opted into regulating multiple key virulence traits in addition to nitrogen catabolism. Here, we further characterize Gat1/Are1 function and investigate the regulatory role of the predicted Nmr homolog Tar1. While GAT1/ARE1 expression is induced during nitrogen limitation, TAR1 transcription is unaffected by nitrogen availability. Deletion of TAR1 leads to inappropriate derepression of non-preferred nitrogen catabolic pathways in the simultaneous presence of favoured sources. In addition to exhibiting its evolutionary conserved role of inhibiting GATA factor activity under repressing conditions, Tar1 also positively regulates GAT1/ARE1 transcription under non-repressing conditions. The molecular mechanism by which Tar1 modulates nitrogen metabolite repression, however, remains open to speculation. Interaction between Tar1 and Gat1/Are1 was undetectable in a yeast two-hybrid assay, consistent with Tar1 and Gat1/Are1 each lacking the conserved C-terminus regions present in ascomycete Nmr proteins and GATA factors that are known to interact with each other. Importantly, both Tar1 and Gat1/Are1 are suppressors of C. neoformans virulence, reiterating and highlighting the paradigm of nitrogen regulation of pathogenesis

[SWI+], the Prion Formed by the Chromatin Remodeling Factor Swi1, Is Highly Sensitive to Alterations in Hsp70 Chaperone System Activity

The yeast prion [SWI+], formed of heritable amyloid aggregates of the Swi1 protein, results in a partial loss of function of the SWI/SNF chromatin-remodeling complex, required for the regulation of a diverse set of genes. Our genetic analysis revealed that [SWI+] propagation is highly dependent upon the action of members of the Hsp70 molecular chaperone system, specifically the Hsp70 Ssa, two of its J-protein co-chaperones, Sis1 and Ydj1, and the nucleotide exchange factors of the Hsp110 family (Sse1/2). Notably, while all yeast prions tested thus far require Sis1, [SWI+] is the only one known to require the activity of Ydj1, the most abundant J-protein in yeast. The C-terminal region of Ydj1, which contains the client protein interaction domain, is required for [SWI+] propagation. However, Ydj1 is not unique in this regard, as another, closely related J-protein, Apj1, can substitute for it when expressed at a level approaching that of Ydj1. While dependent upon Ydj1 and Sis1 for propagation, [SWI+] is also highly sensitive to overexpression of both J-proteins. However, this increased prion-loss requires only the highly conserved 70 amino acid J-domain, which serves to stimulate the ATPase activity of Hsp70 and thus to stabilize its interaction with client protein. Overexpression of the J-domain from Sis1, Ydj1, or Apj1 is sufficient to destabilize [SWI+]. In addition, [SWI+] is lost upon overexpression of Sse nucleotide exchange factors, which act to destabilize Hsp70's interaction with client proteins. Given the plethora of genes affected by the activity of the SWI/SNF chromatin-remodeling complex, it is possible that this sensitivity of [SWI+] to the activity of Hsp70 chaperone machinery may serve a regulatory role, keeping this prion in an easily-lost, meta-stable state. Such sensitivity may provide a means to reach an optimal balance of phenotypic diversity within a cell population to better adapt to stressful environments

Function of SSA Subfamily of Hsp70 Within and Across Species Varies Widely in Complementing Saccharomyces cerevisiae Cell Growth and Prion Propagation

BACKGROUND:The cytosol of most eukaryotic cells contains multiple highly conserved Hsp70 orthologs that differ mainly by their spatio-temporal expression patterns. Hsp70s play essential roles in protein folding, transport or degradation, and are major players of cellular quality control processes. However, while several reports suggest that specialized functions of Hsp70 orthologs were selected through evolution, few studies addressed systematically this issue. METHODOLOGY/PRINCIPAL FINDINGS:We compared the ability of Ssa1p-Ssa4p from Saccharomyces cerevisiae and Ssa5p-Ssa8p from the evolutionary distant yeast Yarrowia lipolytica to perform Hsp70-dependent tasks when expressed as the sole Hsp70 for S. cerevisiae in vivo. We show that Hsp70 isoforms (i) supported yeast viability yet with markedly different growth rates, (ii) influenced the propagation and stability of the [PSI(+)] and [URE3] prions, but iii) did not significantly affect the proteasomal degradation rate of CFTR. Additionally, we show that individual Hsp70 orthologs did not induce the formation of different prion strains, but rather influenced the aggregation properties of Sup35 in vivo. Finally, we show that [URE3] curing by the overexpression of Ydj1p is Hsp70-isoform dependent. CONCLUSION/SIGNIFICANCE:Despite very high homology and overlapping functions, the different Hsp70 orthologs have evolved to possess distinct activities that are required to cope with different types of substrates or stress situations. Yeast prions provide a very sensitive model to uncover this functional specialization and to explore the intricate network of chaperone/co-chaperone/substrates interactions