DIFFERENTIAL LIFE HISTORY PHASE EXPRESSION IN TWO COEXISTING SPECIES OF SCYTOSIPHON (PHAEOPHYCEAE) IN NORTHERN CHILE1

The identity of two phaeophycean taxa that monopolized the middle‐lower rocky intertidal zone of a coastal area chronically exposed to copper mine wastes in northern Chile was unraveled. One of them was preliminarily identified as the gametophytic stage of Scytosiphon lomentaria (Lyngbye) Link. The other, a dark crust, resembled the alternate stage of some Scytosiphon species. Comparative analysis of morphology, life history, and DNA sequences strongly suggests that crusts corresponded to sporophytic S. tenellus Kogame and confirm that erect thalli belonged to S. lomentaria. A clear segregation of erect and crustose thalli was found using internal transcribed spacer region 1 and RUBISCO spacer sequences. Furthermore, whereas crusts always grouped with S. tenellus, erect thalli always grouped with S. lomentaria. Life history studies failed to connect the two entities. First, field‐collected S. tenellus produced progeny that either recycled the crust, which reproduced by unilocular zoidangia, or developed into erect thalli. The latter, unlike typical gametophytic S. lomentaria, developed patchy sori of plurilocular zoidangia. Second, S. lomentaria displayed a direct‐type life cycle, in which progeny from erect individuals only developed into erect thalli and produced only plurilocular zoidangia. This constitutes the first experimental study on Scytosiphon from the Pacific coast of South America and the first report of S. tenellus on this coast. It is also the first report of the crustose stage of Scytosiphon appearing as a perennial and dominant algal species in a temperate rocky intertidal system.     

Activities of de-N-glycosylation are ubiquitously found in tomato plant

Activities of two de-N-glycosylation enzymes, PNGase (peptide N ⁴(N-acetyl-glucosaminyl)asparagine amidase) and ENGase (endo N-acetyl-β-D-glucosaminidase), involved in the release of N-glycans from N-glycoproteins, were monitored in several organs of tomato plants (Lycopersicon esculentum, Mill., cv. Dombito) with a fluorescence-HPLC procedure using a resofurin-labelled N-glycopeptide substrate. PNGase and ENGase activities were detected in every organ assayed but with quantitative differences. The highest activities were found in the youngest parts of the plant, i.e. apical buds, flowers and leaf blades. PNGase activities were consistently higher than ENGase activities (three-fold in average). Both de-N-glycosylation activities were associated with high levels of proteins and protease activities. During fruit growth and ripening, these three parameters decreased notably. The ubiquitous detection of these enzyme activities in the different organs is probably associated with the previously characterized unconjugated N-glycans in tomato. The possible role of PNGase and ENGase degradation products (i.e. unconjugated N-glycans) are discussed in relation with their biological functions in plant development.     

Higher reproductive success for chimeras than solitary individuals in the kelp <Emphasis Type="Italic">Lessonia spicata</Emphasis> but no benefit for individual genotypes

Chimerism is a peculiar, yet widespread, type of group living in which genetically heterogeneous entities are created through fusion between conspecifics. Here we tested whether chimerism provides direct benefits to the kelp Lessonia spicata, by analyzing its consequences on reproductive investment and success, at both the genotype and thallus levels. In addition, we quantified the frequency of chimerism in two natural populations, tested if group members were close kin, and evaluated the effects of relatedness and the number of genotypes per thallus on reproduction. Chimeric thalli were frequent (>60 %) in natural populations of L. spicata. In most cases, average intragroup relatedness was not significantly different from the background population. Reproductive investment was not significantly affected by the type of thallus (chimeric versus non-chimeric), by the number of genotypes per thallus or the average relatedness within thallus. Chimerism did not result in net benefits or costs in terms of genotypic reproductive success or probability of reproducing at the genotypic level. Yet, at the thallus level, chimerism increased reproductive success and the probability of reproducing, since more than one genotype reproduced in chimeric thallus. At the population level, chimerism affects L. spicata reproductive success by allowing the coexistence of a higher density of potential reproducers and mates compared to a scenario with only non-chimeric thallus. Chimerism may then have an important effect on the effective population size and possibly in reducing selfing rates.     

Masc2, a gene from Ascobolus encoding a protein with a DNA-methyltransferase activity in vitro, is dispensable for in vivo methylation

We have shown previously that masc1 , a gene encoding a putative C5-DNA-methyltransferase (MTase), was necessary for the de novo 'Methylation Induced Premeiotically' (MIP) process and sexual reproduction in Ascobolus , whereas it was dispensable for maintenance methylation. A second MTase gene from Ascobolus , masc2 , encodes a protein, Masc2, which possesses the large amino-terminal part characteristic of eukaryotic maintenance MTases. In vitro assays have shown that Masc2 displays a methylation activity, suggesting that it might be the MTase responsible for maintenance methylation. To check its function in vivo , we engineered a disruption of the masc2 gene. The resulting mutant strains did not exhibit any particular phenotype during either vegetative growth or sexual reproduction. Neither the masc2 mutation nor the double masc1 masc2 mutation had any detectable effect upon the maintenance of the pre-existing methylation of single gene copies previously subjected to MIP, natural retroelement-like repeats and tandemly repeated rDNA. The masc2 mutation did not alter either MIP or the other de novo methylation process that operates in vegetative cells. Nor did it impair the meiotic process of methylation transfer. These results suggest that at least a third MTase gene responsible for maintenance and vegetative de novo methylation is present in Ascobolus .     

Experimental transplants of the large kelp Lessonia nigrescens (Phaeophyceae) in high-energy wave exposed rocky intertidal habitats of northern Chile: Experimental, restoration and management applications

Potential for addressing ecological and physiological issues becomes severely limited when the organisms required to experimentally test specific hypotheses are absent from the study areas. This report describes a simple and inexpensive device for re-planting kelps into the lower intertidal zone of wave-swept rocky habitats, using Lessonia nigrescens Bory as a model organism. The device allows a wide range of plant sizes to be anchored by the holdfast. Transplanted kelps regenerated, re-attached to the substratum and overgrew the transplanting device. These results confirm the feasibility of using this technique to tackle a variety of highly relevant questions involving ecological, physiological, conservation, restoration and management issues.     

Stable allele replacement and unstable non-homologous integration events during transformation of Ascobolus immersus

An homologous transformation system for the filamentous fungus Ascobolus immersus has been developed, based on the complementation of a met2 mutation by the wild-type (wt) allele gene encoding homoserine O-transacetylase. Transformation of A. immersus met2 mutants occurs with moderate frequencies (about 50 transformants per microgram input DNA). Analysis of the DNA of the met2+ transformants showed that transformation resulted either in a single integration of the donor DNA into the genome by many different nonhomologous recombination events or in the substitution of the endogenous met2 mutation by the wt transforming allele. The relative frequencies of both events depended on the vector sequences carrying the cloned met2 gene. Whereas the substitution event led, as expected, to genetically stable transformants, the non-homologous integration was always associated with a strong instability when transformants were crossed and underwent meiosis.     

Self-fertilization is the main sexual reproduction mechanism in native wine yeast populations

Saccharomyces cerevisiae is a model eukaryotic organism for classical genetics and genomics, and yet its ecology is still largely unknown. In this work, a population genetic analysis was performed on five yeast populations isolated from wine-making areas with different enological practices using simple sequence repeats and restriction fragment length polymorphism of mitochondrial DNA as molecular markers on 292 strains. In accordance with other studies, genome size estimation suggests that native S. cerevisiae strains are mainly homothallic and diploids. Analysis of mtDNA data showed that yeast populations from nonindustrial areas have 40% higher genetic diversity than populations isolated from industrial areas, demonstrating that industrial enological practices are likely to affect native yeast populations negatively by reducing its biodiversity. On the other hand, genetic differentiation analysis based on their microsatellite showed no correlation between genetic and geographic distance and a nonsignificant value when a Mantel test was applied. Finally, in the five populations studied, positive inbreeding ( F _is) values from 0.4 to 0.75, a low but significant level of linkage disequilibrium and a high number of multilocus genotypes were detected. These results strongly advocate that sexual reproduction is frequent enough to erase clonal signature in natural populations and that self-fertilization is the main mating system.     

Species replacement along a linear coastal habitat: phylogeography and speciation in the red alga Mazzaella laminarioides along the south east pacific

ABSTRACT: BACKGROUND: The Chilean shoreline, a nearly strait line of coast expanding across 35 latitudinal degrees, represents an interesting region to assess historical processes using phylogeographic analyses. Stretching along the temperate section of the East Pacific margin, the region is characterized by intense geologic activity and has experienced drastic geomorphological transformations linked to eustatic and isostatic changes during the Quaternary. In this study, we used two molecular markers to evaluate the existence of phylogeographic discontinuities and detect the genetic footprints of Pleistocene glaciations among Patagonian populations of Mazzaella laminarioides, a low-dispersal benthic intertidal red seaweed that inhabits along ~3,700 km of the Chilean coastal rocky shore. RESULTS: Three main genetic lineages were found within M. laminarioides. They are distributed along the Chilean coast in strict parapatry. The deep divergence among lineages suggests that they could be considered putative genetic sibling species. Unexpectedly, genetic breaks were not strictly concordant with the biogeographic breaks described in the region. A Northern lineage was restricted to a broad transition zone located between 30degreesS and 33degreesS and showed signals of a recent bottleneck. The reduction of population size could be related to warm events linked to El Nino Southern Oscillation, which is known to cause massive seaweed mortality in this region. To the south, we propose that transient habitat discontinuities driven by episodic tectonic uplifting of the shoreline around the Arauco region (37degreesS-38degreesS); one of the most active forearc-basins in the South East Pacific; could be at the origin of the Central/South genetic break. The large beaches, located around 38degreesS, are likely to contribute to the lineages' integrity by limiting present gene flow. Finally, the Southern lineage, occupies an area affected by ice-cover during the last glaciations. Phylogeny suggested it is a derived clade and demographic analyses showed the lineage has a typical signature of postglacial recolonization from a northern glacial refugium area. CONCLUSIONS: Even if environmental adaptation could have strengthened divergence among lineages in M. laminarioides, low dispersal capacity and small population size are sufficient to generate phylogeographic discontinuities determined by genetic drift alone. Interestingly, our results confirm that seaweed population connectivity over large geographic scales does not rely only on dispersal capacity but also seem to depend highly on substratum availability and population density of the receiving locality.     

Response of cultured tomato cells subjected to excess zinc: role of cell wall in zinc compartmentation

The aim of this preliminary study was to evaluate the role of the cell wall in Zn accumulation and tolerance by tomato suspension-cultured cells. Growth parameters, Zn distribution and accumulation by tomato cells were determined in function of zinc concentration. A particular attention was paid to the variations of the total cell wall material (cell wall carbohydrates, proteins, and exopolymers) in relation to extracellular levels of Zn. Cells treated with 0.5–5 mM Zn showed typical symptoms of heavy metal toxicity as testified by various growth parameters. Fresh and dry weights as well as total cell volume per vial decreased with increasing Zn concentration in the culture medium. Concurrently, the cell wall biomass increased, as well as the Zn amount retained by cell wall polymers. Cell wall appeared to assume important roles in Zn fixation and could therefore limit Zn influx into the cell. Our results also suggested that zinc fixation by cell wall was not only due to an increase in cell wall biomass but also to an improvement of its binding capacity.