Identification of a protein kinase gene associated with pistillody,homeotic transformation of stamens into pistil-like structures,in alloplasmic wheat

Homeotic transformation of stamens into pistil-like structures (called pistillody) has been reported in cytoplasmic substitution (alloplasmic) lines of bread wheat (Triticum aestivum) having the cytoplasm of a wild relative species, Aegilops crassa. Our previous studies indicated that pistillody is caused by alterations of the class B MADS-box gene expression pattern associated with mitochondrial gene(s) in the Ae. crassa cytoplasm. To elucidate the nuclear gene involved in the cross-talk between pistillody-related mitochondrial gene(s) and nuclear homeotic genes, we performed cDNA subtraction analysis using cDNAs derived from young spikes of a pistillody line and a normal line. As a result, we identified a protein kinase gene, WPPK1 (wheat pistillody-related protein kinase 1), which is upregulated in the young spikes of the pistillody line. RT-PCR analysis indicated that WPPK1 is strongly expressed in pistils and pistil-like stamens in the pistillody line, suggesting that it is involved in the formation of pistil-like stamens as well as pistils. The full-length cDNA sequence for WPPK1 showed high similarity with a flowering plant PVPK-1 protein kinase, and phylogenetic analysis indicated that it is a member of AGC group protein kinases. Furthermore, a phosphorylation assay indicated that it has protein kinase activity. In situ hybridization analysis revealed that WPPK1 is expressed in developing pistils and pistil-like stamens as well as in their primordia. These indicate that in the alloplasmic line, WPPK1 plays a role in formation and development of pistil-like stamens.     

Purification and characterization of fibrinolytic alkaline protease from <Emphasis Type="Italic">Fusarium</Emphasis> sp. BLB

Fusarium sp. BLB, which produces a strongly fibrinolytic enzyme, was isolated from plant leaf (Hibiscus). Fibrinolytic alkaline protease was purified from a culture filtrate of Fusarium sp. BLB by precipitation with (NH₄)₂SO₄ and column chromatography with CM-Toyopearl 650M and Superdex 75. The purified enzyme was homogeneous on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight was 27,000 by SDS-PAGE. Maximum activity of protease was observed at pH 9.5 and 50°C. Purified protease was active between pH 2.5 and 11.5 and was found to be stable up to 50°C. The enzyme derived from Fusarium sp. BLB is useful for thrombolytic therapy because this enzyme showed pH resistance. The activity was inhibited by diisopropylfluorophosphate and phenylmethylsulfonyl fluoride. The N-terminal amino acid sequence of the enzyme showed a similarity to those of proteases from Fusarium sp., Streptomyces griseus, Bos taurus bovine, Katsuwo pelamis digestive tract, and Lumbricus rubellus.     

Effect of CO2 on Colony Development by Bifidobacterium Species

This report investigates the requirement for CO₂ for colony formation by Bifidobacterium species in both anoxic and oxic environments. All tested Bifidobacterium species exhibited difficulty in developing colonies in an atmosphere of 100% N₂ but developed well when 1% CO₂ was present. In the presence of CO₂, the oxygen tolerance of the tested species was not improved. In the absence of CO₂, only B. boum, a microaerophilic species, could develop colonies under an N₂-based 5% O₂ atmosphere, indicating that while CO₂ is not an essential factor for colony development, both CO₂ and O₂ have stimulatory effects on B. boum colony development.     

Efficient Extraction of Thioreodoxin from Saccharomyces cerevisiae by Ethanol

Thioredoxin, an antioxidant protein, is a promising molecule for development of functional foods because it protects the gastric mucosa and reduces the allergenicity of allergens. To establish a method for obtaining an ample amount of yeast thioredoxin, we found here that thioredoxin is released from Saccharomyces cerevisiae by treatment with 20% ethanol. We also found that Japanese sake contains a considerable amount of thioredoxin.     

Msn2p/Msn4p-activation is essential for the recovery from freezing stress in yeast

Recent data suggest that the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate receptor subtype plays a pivotal role in the pathogenesis of effective disorders and in the action of antidepressant drugs. After chronic treatment with the antidepressants desipramine or paroxetine, we measured by immunoprecipitation and Western blotting, the changes in the interaction of AMPA receptor subunits with proteins involved in trafficking and/or stabilization of the subunits into synaptic membranes of the hippocampus. Both antidepressants increased the interaction of GluR1 subunit with stargazin and of GluR2/3 with NSF. Paroxetine increased the interaction of GluR1 with Rab4A, and desipramine markedly increased the interaction of GluR1 with SAP97. Paroxetine, but not desipramine, also increased membrane levels of CaMKII, autophosphorylated CaMKII and GluR1 phosphorylated at the CaMKII site. Interactions of GluR1 and GluR2/3 with proteins implicated in AMPA receptor trafficking and with scaffolding proteins appear to account for the enhanced membrane expression of AMPA receptors in the hippocampus after antidepressant treatment.     

A straightforward stereoselective synthesis of meso-, (S,S)- and (R,R)-2,6-diaminopimelic acids from cis-1,4-diacetoxycyclohept-2-ene

A straightforward synthesis of meso-2,6-diaminopimelic acid (DAP) meso-1 was developed from 1,4-diacetoxycyclohept-2-ene (2) via an oxidative ring cleavage. Subsequently, an enantio-divergent synthesis of (S,S)- and (R,R)-1 was performed using a homochiral monoacetate 7 available from 2 by enzymatic desymmetrization.     

Genetic and epigenetic alteration among three homoeologous genes of a class E MADS box gene in hexaploid wheat

Bread wheat (Triticum aestivum) is a hexaploid species with A, B, and D ancestral genomes. Most bread wheat genes are present in the genome as triplicated homoeologous genes (homoeologs) derived from the ancestral species. Here, we report that both genetic and epigenetic alterations have occurred in the homoeologs of a wheat class E MADS box gene. Two class E genes are identified in wheat, wheat SEPALLATA (WSEP) and wheat LEAFY HULL STERILE1 (WLHS1), which are homologs of Os MADS45 and Os MADS1 in rice (Oryza sativa), respectively. The three wheat homoeologs of WSEP showed similar genomic structures and expression profiles. By contrast, the three homoeologs of WLHS1 showed genetic and epigenetic alterations. The A genome WLHS1 homoeolog (WLHS1-A) had a structural alteration that contained a large novel sequence in place of the K domain sequence. A yeast two-hybrid analysis and a transgenic experiment indicated that the WLHS1-A protein had no apparent function. The B and D genome homoeologs, WLHS1-B and WLHS1-D, respectively, had an intact MADS box gene structure, but WLHS1-B was predominantly silenced by cytosine methylation. Consequently, of the three WLHS1 homoeologs, only WLHS1-D functions in hexaploid wheat. This is a situation where three homoeologs are differentially regulated by genetic and epigenetic mechanisms.     

The chloroplast genome from a lycophyte (microphyllophyte), <Emphasis Type="Italic">Selaginella uncinata</Emphasis>, has a unique inversion,transpositions and many gene losses

We determined the complete nucleotide sequence of the chloroplast genome of Selaginella uncinata, a lycophyte belonging to the basal lineage of the vascular plants. The circular double-stranded DNA is 144,170 bp, with an inverted repeat of 25,578 bp separated by a large single copy region (LSC) of 77,706 bp and a small single copy region (SSC) of 40,886 bp. We assigned 81 protein-coding genes including four pseudogenes, four rRNA genes and only 12 tRNA genes. Four genes, rps15, rps16, rpl32 and ycf10, found in most chloroplast genomes in land plants were not present in S. uncinata. While gene order and arrangement of the chloroplast genome of another lycophyte, Hupertzia lucidula, are almost the same as those of bryophytes, those of S. uncinata differ considerably from the typical structure of bryophytes with respect to the presence of a unique 20 kb inversion within the LSC, transposition of two segments from the LSC to the SSC and many gene losses. Thus, the organization of the S. uncinata chloroplast genome provides a new insight into the evolution of lycophytes, which were separated from euphyllophytes approximately 400 million years ago. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.