Transmission ratio distortion of molecular markers in a doubled haploid population originated from a natural hybrid between Osmunda japonica and O. lancea

In ferns, intra-gametophytic selfing occurs as a mode of reproduction where two gametes from the same gametophyte form a completely homozygous sporophyte. Intra-gametophytic selfing is considered to be prevented by lethal or deleterious recessive genes in several diploid species. In order to investigate the modes and tempo of selection acting different developmental stages, doubled haploids obtained from intra-gametophytic selfing within isolated gametophytes of a putative F1 hybrid between Osmunda japonica and O. lancea were analyzed with EST_derived molecular markers, and the distribution pattern of transmission ratio distortion (TRD) along linkage map was clarified. As the results, the markers with skewness were clustered in two linkage groups. For the two highly distorted regions, gametophytes and F2 population were also examined. The markers skewed towards O. japonica on a linkage group (LG_2) showed skewness also in gametophytes, and the TRD was generated in the process of spore formation or growth of gametophytes. Also, selection appeared to be operating in the gametophytic stage. The markers on other linkage group (LG_11) showed highest skewness towards O. lancea in doubled haploids, and it was suggested that the segregation of LG_11 were influenced by zygotic lethality or genotypic evaluation and that some deleterious recessive genes exist in LG_11 and reduce the viability of homozygotes with O. japonica alleles. It is very likely that a region of LG_11were responsible for the low frequencies of intra-gametophytic selfing in O. japonica.     

Rampant gene exchange across a strong reproductive barrier between the annual sunflowers, Helianthus annuus and H. petiolaris

Plant species may remain morphologically distinct despite gene exchange with congeners, yet little is known about the genomewide pattern of introgression among species. Here we analyze the effects of persistent gene flow on genomic differentiation between the sympatric sunflower species Helianthus annuus and H. petiolaris. While the species are strongly isolated in testcrosses, genetic distances at 108 microsatellite loci and 14 sequenced genes are highly variable and much lower (on average) than for more closely related but historically allopatric congeners. Our analyses failed to detect a positive association between levels of genetic differentiation and chromosomal rearrangements (as reported in a prior publication) or proximity to QTL for morphological differences or hybrid sterility. However, a significant increase in differentiation was observed for markers within 5 cM of chromosomal breakpoints. Together, these results suggest that islands of differentiation between these two species are small, except in areas of low recombination. Furthermore, only microsatellites associated with ESTs were identified as outlier loci in tests for selection, which might indicate that the ESTs themselves are the targets of selection rather than linked genes (or that coding regions are not randomly distributed). In general, these results indicate that even strong and genetically complex reproductive barriers cannot prevent widespread introgression.     

Evolution of Mhc–DRB Introns: Implications for the Origin of Primates

Introns are generally believed to evolve too rapidly and too erratically to be of much use in phylogenetic reconstructions. Few phylogenetically informative intron sequences are available, however, to ascertain the validity of this supposition. In the present study the supposition was tested on the example of the mammalian class II major histocompatibility complex (Mhc) genes of the DRB family. Since the Mhc genes evolve under balancing selection and are believed to recombine or rearrange frequently, the evolution of their introns could be expected to be particularly rapid and subject to scrambling. Sequences of intron 4 and 5 DRB genes were obtained from polymerase chain reaction-amplified fragments of genomic DNA from representatives of six eutherian orders—Primates, Scandentia, Chiroptera, Dermoptera, Lagomorpha, and Insectivora. Although short stretches of the introns have indeed proved to be unalignable, the bulk of the intron sequences from all six orders, spanning >85 million years (my) of evolution, could be aligned and used in a study of the tempo and mode of intron evolution. The analysis has revealed the Mhc introns to evolve at a rate similar to that of other genes and of synonymous sites of non-Mhc genes. No evidence of homogenization or large-scale scrambling of the intron sequences could be found. The Mhc introns apparently evolve largely by point mutations and insertions/deletions. The phylogenetic signals contained in the intron sequences could be used to identify Scandentia as the sister group of Primates, to support the existence of the Archonta superorder, and to confirm the monophyly of the Chiroptera. Received: 26 October 1998 / Accepted: 21 December 1998     

Dissociation of double-headed cytoplasmic dynein into single-headed species and its motile properties

Cytoplasmic dynein is a minus-end directed microtubule motor and plays important roles in the transport of various intracellular cargoes. Cytoplasmic dynein comprises two identical heavy chains and forms a dimer (double-headed dynein); the total molecular weight of the cytoplasmic dynein complex is about 1.5 million. The dynein motor domain is structurally very different from those of kinesin and myosin, and our understanding of the mechanisms of dynein energy transduction is limited mainly because of the difficulty in obtaining a sufficient quantity of purified and active cytoplasmic dynein. We purified cytoplasmic dynein, which was free from dynactin and other dynein-associated proteins. The purified cytoplasmic dynein was active in an in vitro motility assay. The controlled dialysis of the purified dynein against 4 M urea resulted in its complete dissociation into monomeric species (single-headed dynein). The separation of the dynein heads by the treatment was reversible. The MgATPase activities of the single-headed and reconstituted double-headed dynein were comparable to that of intact dynein. The double-headed dynein bundled microtubules in the absence of ATP; the single-headed dynein did not. The single-headed dynein produced in vitro microtubule-gliding motility at velocities very similar to those of double-headed dynein at various ATP concentrations. These results indicate that a single cytoplasmic dynein heavy chain is sufficient to produce robust microtubule motility. Application of the double- and single-headed dynein molecules in various assay systems will elucidate the mechanism of action of the cytoplasmic dynein.     

Properties of the full-length heavy chains of Tetrahymena ciliary outer arm dynein separated by urea treatment

An important challenge is to understand the functional specialization of dynein heavy chains. The ciliary outer arm dynein from Tetrahymena thermophila is a heterotrimer of three heavy chains, called alpha, beta and gamma. In order to dissect the contributions of the individual heavy chains, we used controlled urea treatment to dissociate Tetrahymena outer arm dynein into a 19S beta/gamma dimer and a 14S alpha heavy chain. The three heavy chains remained full-length and retained MgATPase activity. The beta/gamma dimer bound microtubules in an ATP-sensitive fashion. The isolated alpha heavy chain also bound microtubules, but this binding was not reversed by ATP. The 19S beta/gamma dimer and the 14S alpha heavy chain could be reconstituted into 22S dynein. The intact 22S dynein, the 19S beta/gamma dimer, and the reconstituted dynein all produced microtubule gliding motility. In contrast, the separated alpha heavy chain did not produce movement under a variety of conditions. The intact 22S dynein produced movement that was discontinuous and slower than the movement produced by the 19S dimer. We conclude that the three heavy chains of Tetrahymena outer arm dynein are functionally specialized. The alpha heavy chain may be responsible for the structural binding of dynein to the outer doublet A-tubule and/or the positioning of the beta/gamma motor domains near the surface of the microtubule track.     

Prooxidative activities of tea catechins in the presence of Cu2+

The ability of various tea catechins to generate H2O2 and the hydroxyl radical in the presence of the Cu2+ ion was investigated and compared with the effect of iron ions. The presence of Cu2+ accelerated the generation of H2O2 by EGC, while EGCg with Cu2+ generated a little H2O2. The presence of iron ions inhibited the generation of H2O2 by EGC. EGC and EC with Cu2+ generated the hydroxyl radical, while EGCg and ECg with Cu2+ did not. The fact that EGCg showed less prooxidative activity than EGC can be explained by the chelating ability of catechin gallates to metal ions under the experimental conditions.     

Does seed production of spring ephemerals decrease when spring comes early?

To predict the effect of global warming on plant reproductive success, seed-sets of spring ephemerals were compared between a year of extremely warm spring (2002) and normal years at cool-temperate deciduous forests in northern Japan. The spring of 2002 was the warmest in the last 40years and most spring-ephemeral plants bloomed 7–17days earlier than usual. The seed-set of bumblebee-pollinated Corydalis ambigua drastically decreased in 2002 in every population. The small bee-pollinated Gagea lutea also significantly decreased in 2002. However, the seed-sets of two fly pollinated species, Adonis ramosa and Anemone flaccida, were not influenced by early flowering. These results indicat that the effect of global warming on seed production of spring ephemerals differs between species depending on the type of pollinators, and that bee-pollinated species can have serious impacts on reproductive success as a result of climate change.     

Crystallization and preliminary X-ray analysis of plant class I chitinase from rice

We report here on crystallization and preliminary X-ray analysis of plant class I chitinase from rice (OsChia1b). Similar single crystals of full-length OsChia1b were obtained under two independent conditions. The crystals grown under these conditions diffracted up to 2.1 and 2.5 angstroms resolution, respectively, at a synchrotron beamline, and were found to belong to the tetragonal space group P4(3)2(1)2.