Upholding science in health, safety and environmental risk assessments and regulations.

A public appeal has been advanced by a large group of scientists, concerned that science has been misused in attempting to quantify and regulate unmeasurable hazards and risks.1 The appeal recalls that science is unable to evaluate hazards that cannot be measured, and that science in such cases should not be invoked to justify risk assessments in health, safety and environmental regulations. The appeal also notes that most national and international statutes delineating the discretion of regulators are ambiguous about what rules of evidence ought to apply. Those statutes should be revised to ensure that the evidence for regulatory action is grounded on the standards of the scientific method, whenever feasible. When independent scientific evidence is not possible, policies and regulations should be informed by publicly debated trade-offs between socially desirable uses and social perceptions of affordable precaution. This article explores the premises, implications and actions supporting the appeal and its objectives.

Contrasted microcolinearity and gene evolution within a homoeologous region of wheat and barley species.

We study here the evolution of genes located in the same physical locus using the recently sequenced Ha locus in seven wheat genomes in diploid, tetraploid, and hexaploid species and compared them with barley and rice orthologous regions. We investigated both the conservation of microcolinearity and the molecular evolution of genes, including coding and noncoding sequences. Microcolinearity is restricted to two groups of genes (Unknown gene-2, VAMP, BGGP, Gsp-1, and Unknown gene-8 surrounded by several copies of ATPase), almost conserved in rice and barley, but in a different relative position. Highly conserved genes between wheat and rice run along with genes harboring different copy numbers and highly variable sequences between close wheat genomes. The coding sequence evolution appeared to be submitted to heterogeneous selective pressure and intronic sequences analysis revealed that the molecular clock hypothesis is violated in most cases.

The promoter of the wheat puroindoline-a gene (PinA) exhibits a more complex pattern of activity than that of the PinB gene and is induced by wounding and pathogen attack in rice.

Puroindolines form the molecular basis of wheat grain hardness. However, little is known about puroindoline gene regulation. We previously reported that the Triticum aestivum puroindoline-b gene (PinB) promoter directs beta-glucuronidase gene (uidA) seed-specific expression in transgenic rice. In this study, we isolated a puroindoline-a gene (PinA), analyzed PinA promoter activity by 5' deletions and compared PinA and PinB promoters in transgenic rice. Seeds of PinA-1214 and PinB-1063 transgenic plants strongly expressed uidA in endosperm, in the aleurone layer and in epidermis cells in a developmentally regulated manner. The GUS activity was also observed in PinA-1214 embryos. Whereas the PinB promoter is seed specific, the PinA promoter also directed, but to a lower level, uidA expression in roots of seedlings and in the vascular tissues of palea and pollen grains of dehiscent anthers during flower development. In addition, the PinA promoter was induced by wounding and by Magnaporthe grisea. By deletion analysis, we showed that the "390-bp" PinA promoter drives the same expression pattern as the "1214-bp" promoter. Moreover, the "214-bp" PinA promoter drives uidA expression solely in pollen grains of dehiscent anthers. The presence of putative cis-regulatory elements that may be related to PinA expression is discussed from an evolutionary point of view. By electrophoretic mobility shift assay, we showed that putative cis-elements (WUN-box, TCA motifs and as-1-like binding sites) whose presence in the PinA promoter may be related to wounding and/or the pathogen response form complexes with nuclear extracts isolated from wounded wheat leaves.

The Triticum aestivum non-specific lipid transfer protein (TaLtp) gene family: comparative promoter activity of six TaLtp genes in transgenic rice.

Plant non-specific lipid transfer proteins (nsLTPs) are encoded by a multigene family and support physiological functions, which remain unclear. We adapted an efficient ligation-mediated polymerase chain reaction (LM-PCR) procedure that enabled isolation of 22 novel Triticum aestivum nsLtp (TaLtp) genes encoding types 1 and 2 nsLTPs. A phylogenetic tree clustered the wheat nsLTPs into ten subfamilies comprising 1-7 members. We also studied the activity of four type 1 and two type 2 TaLtp gene promoters in transgenic rice using the 1-Glucuronidase reporter gene. The activities of the six promoters displayed both overlapping and distinct features in rice. In vegetative organs, these promoters were active in leaves and root vascular tissues while no beta-Glucuronidase (GUS) activity was detected in stems. In flowers, the GUS activity driven by the TaLtp7.2a, TaLtp9.1a, TaLtp9.2d, and TaLtp9.3e gene promoters was associated with vascular tissues in glumes and in the extremities of anther filaments whereas only the TaLtp9.4a gene promoter was active in anther epidermal cells. In developing grains, GUS activity and GUS immunolocalization data evidenced complex patterns of activity of the TaLtp7.1a, TaLtp9.2d, and TaLtp9.4a gene promoters in embryo scutellum and in the grain epicarp cell layer. In contrast, GUS activity driven by TaLtp7.2a, TaLtp9.1a, and TaLtp9.3e promoters was restricted to the vascular bundle of the embryo scutellum. This diversity of TaLtp gene promoter activity supports the hypothesis that the encoded TaLTPs possess distinct functions in planta.

Wheat non-specific lipid transfer protein genes display a complex pattern of expression in developing seeds.

Nine cDNA clones encoding non-specific lipid transfer proteins (nsLTPs) were isolated from Triticum aestivum and Triticum durum cDNA libraries and characterized. One cDNA is predicted to encode a type 2 nsLTP (7 kDa) while others encode type 1 nsLTPs (9 kDa). All encoded proteins contain an N-terminal signal sequence and possess the characteristic features of nsLTPs. The genomic structures of the wheat nsLtp genes show that type 2 TaLtp7.1a, TaLtp7.2a and type 1 TaLtp9.2b genes lack introns while the other type 1 genes consist of one intron. Construction of a phylogenic tree of Poaceae nsLTPs shows that wheat nsLTPs can be divided into eleven distinct groups and are closely related to barley sequences. Using reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, the expression patterns of nine nsLtp genes were studied during wheat seed development and germination. We identified three different profiles of nsLtp gene transcript accumulation. Whereas TdLtp7.1a, TdLtp9.4a and TdLtp9.7a transcripts were detected during all maturation stages, TdLtp7.2a, TdLtp9.2a, TdLtp9.3a, TdLtp9.5a and TdLtp9.6a transcripts were only present in the first and TdLtp9.1a in the last stages of seed development. Moreover, these nine wheat nsLtp genes are not seed-specific and are also expressed in the coleoptile of young seedlings. The present study revealed the complexity of the wheat nsLtp gene family and showed that the expression of nsLtp genes is developmentally regulated in the seeds, suggesting a specific function for each of the corresponding proteins.

Comparative expression of five Lea Genes during wheat seed development and in response to abiotic stresses by real-time quantitative RT-PCR.

Gene expression profiles of group 2 (dehydrins) and group 4 Late embryogenesis abundant (Lea) genes in developing seeds of Triticum durum and T. aestivum and in coleoptiles and coleorhizae of T. durum seedlings were monitored by real-time quantitative RT-PCR. The five genes exhibited clear differences in their accumulation pattern in wheat seed and in response to dehydration, low temperature, salinity and ABA. Td29b, Td16 and Td27e gene transcripts accumulate late in embryogenesis as expected for Lea genes, Td11 gene transcripts were present throughout seed development whereas no Td25a gene transcripts were detected in seeds. Drastic changes in the relative levels of Td29b, Td16, Td27e and Td11 transcripts occurred at the shift between the cell expansion and desiccation phases. All genes except the Td11 gene are more highly induced by dehydration in coleorhizae than in coleoptiles. In contrast, response to low temperature, salinity or ABA is higher in coleoptiles than in coleorhizae. Depending on both the gene and on the type of stress, a wide range of induction levels (8- to 100,000-fold) was observed.

Molecular basis of evolutionary events that shaped the hardness locus in diploid and polyploid wheat species (Triticum and Aegilops).

The Hardness (Ha) locus controls grain hardness in hexaploid wheat (Triticum aestivum) and its relatives (Triticum and Aegilops species) and represents a classical example of a trait whose variation arose from gene loss after polyploidization. In this study, we investigated the molecular basis of the evolutionary events observed at this locus by comparing corresponding sequences of diploid, tertraploid, and hexaploid wheat species (Triticum and Aegilops). Genomic rearrangements, such as transposable element insertions, genomic deletions, duplications, and inversions, were shown to constitute the major differences when the same genomes (i.e., the A, B, or D genomes) were compared between species of different ploidy levels. The comparative analysis allowed us to determine the extent and sequences of the rearranged regions as well as rearrangement breakpoints and sequence motifs at their boundaries, which suggest rearrangement by illegitimate recombination. Among these genomic rearrangements, the previously reported Pina and Pinb genes loss from the Ha locus of polyploid wheat species was caused by a large genomic deletion that probably occurred independently in the A and B genomes. Moreover, the Ha locus in the D genome of hexaploid wheat (T. aestivum) is 29 kb smaller than in the D genome of its diploid progenitor Ae. tauschii, principally because of transposable element insertions and two large deletions caused by illegitimate recombination. Our data suggest that illegitimate DNA recombination, leading to various genomic rearrangements, constitutes one of the major evolutionary mechanisms in wheat species.