Sequence of three members and expression of a new major subfamily of glutelin genes from rice

Three members have been isolated of an additional glutelin gene subfamily, named subfamily B, consisting of about five members per haploid rice genome. Restriction fragment length polymorphism analysis showed major differences between Japonica and Indica lines, indicating the divergence of the subfamily since the split between the two varieties. While corresponding exons of the subfamily B showed 80 to 88% nucleotide sequence homology, those exons were only 60–65% homologous to those of the glutelin A subfamily [15, 19, 24], distinguishing them from the subfamily A. Intron position and derived polypeptide structure, in addition to the nucleotide sequence, confirm the subfamily B members as glutelins. Analysis of RNA from seeds of different stages of development showed that the subfamily B members were expressed at the same time as those of subfamily A, demonstrating coordinated regulation of the two subfamilies.     

Role of a novel coiled-coil domain-containing protein CCDC69 in regulating central spindle assembly

The formation of the central spindle (or the spindle midzone) is essential for cytokinesis in animal cells. In this study, we report that coiled-coil domain-containing protein 69 (CCDC69) is implicated in controlling the assembly of central spindles and the recruitment of midzone components. Exogenous expression of CCDC69 in HeLa cells interfered with microtubule polymerization and disrupted the formation of bipolar mitotic spindles. Endogenous CCDC69 proteins were localized to the central spindle during anaphase. RNA interference (RNAi)-mediated knockdown of CCDC69 led to the formation of aberrant central spindles and disrupted the localization of midzone components such as aurora B kinase, protein regulator of cytokinesis 1 (PRC1), MgcRacGAP/HsCYK-4, and polo-like kinase 1 (Plk1) at the central spindle. Aurora B kinase was found to bind to CCDC69 and this binding depended on the coiled-coil domains at the C-terminus of CCDC69. Further, disruption of aurora B function in HeLa cells by treatment with a small chemical inhibitor led to the mislocalization of CCDC69 at the central spindle. Our results indicate that CCDC69 acts as a scaffold to regulate the recruitment of midzone components and the assembly of central spindles.Key words: CCDC69, aurora B, Plk1, central spindles, midzone components, cytokinesis     

The levels of main urinary metabolite of prostaglandin F1α and F2α in human subjects measured by radioimmunoassay

Radioimmunoassay technique for measuring 5α,7α-dihydroxy-11-keto-tetranorprosta-1,16-dioic acid, the main urinary metabolite of PGF₁α and PGF₂α (PGF₂α-MUM), was further improved.It was postulated based on some experimental data that the PGF₂α-MUM exists in the urine mostly as dioic acid form, not as δ-lactone formThe daily excretion of PGF₂α-MUM in men ranged from 14.43 μg to 36.14 μg and in women from 5.21 μg to 14.25 μg.     

Substrate Recognition by the Multifunctional Cytochrome P450 MycG in Mycinamicin Hydroxylation and Epoxidation Reactions

The majority of characterized cytochrome P450 enzymes in actinomycete secondary metabolic pathways are strictly substrate-, regio-, and stereo-specific. Examples of multifunctional biosynthetic cytochromes P450 with broader substrate and regio-specificity are growing in number and are of particular interest for biosynthetic and chemoenzymatic applications. MycG is among the first P450 monooxygenases characterized that catalyzes both hydroxylation and epoxidation reactions in the final biosynthetic steps, leading to oxidative tailoring of the 16-membered ring macrolide antibiotic mycinamicin II in the actinomycete Micromonospora griseorubida. The ordering of steps to complete the biosynthetic process involves a complex substrate recognition pattern by the enzyme and interplay between three tailoring modifications as follows: glycosylation, methylation, and oxidation. To understand the catalytic properties of MycG, we structurally characterized the ligand-free enzyme and its complexes with three native metabolites. These include substrates mycinamicin IV and V and their biosynthetic precursor mycinamicin III, which carries the monomethoxy sugar javose instead of the dimethoxylated sugar mycinose. The two methoxy groups of mycinose serve as sensors that mediate initial recognition to discriminate between closely related substrates in the post-polyketide oxidative tailoring of mycinamicin metabolites. Because x-ray structures alone did not explain the mechanisms of macrolide hydroxylation and epoxidation, paramagnetic NMR relaxation measurements were conducted. Molecular modeling based on these data indicates that in solution substrate may penetrate the active site sufficiently to place the abstracted hydrogen atom of mycinamicin IV within 6 Å of the heme iron and ∼4 Å of the oxygen of iron-ligated water.     

Deodorizing effects of tea catechins on amines and ammonia

Deodorizing effects of tea catechins on amines were examined under alkaline conditions to eliminate the neutralization reaction. They showed deodorizing activity on ethylamine, but none on dimethylamine or trimethylamine. Deodorizing activity on ethylamine was found to be in the order of (-)-epigallocatechin gallate > gallic acid > (-)-epigallocatechin (EGC) > (-)-epicatechin gallate > ethyl gallate > (+)-catechin = (-)-epicatechin. Further, reaction products of EGC with methylamine, ethylamine, and ammonia were detected by HPLC, indicating that a deodorizing reaction other than neutralization occurs. From structural analysis of the reaction product with the methylamine isolated as a peracetylated derivative, the product was presumed to be methylamine substituted EGC, in which the hydroxyl group of EGC at the 4' position is replaced by the methylamino group. The same replacement reaction took place in the case of ethylamine and ammonia.     

Distinct chromatin environment associated with phosphorylated H3S10 histone during pollen mitosis I in orchids

Pollen developmental pathway in plants involving synchronized transferal of cellular divisions from meiosis (microsporogenesis) to mitosis (pollen mitosis I/II) eventually offers a unique “meiosis-mitosis shift” at pollen mitosis I. Since the cell type (haploid microspore) and fate of pollen mitosis I differ from typical mitosis (in meristem cells), it is immensely important to analyze the chromosomal distribution of phosphorylated H3S10 histone during atypical pollen mitosis I to comprehend the role of histone phosphorylation in pollen development. We investigated the chromosomal phosphorylation of H3S10 histone during pollen mitosis I in orchids using immunostaining technique. The chromosomal distribution of H3S10ph during pollen mitosis I revealed differential pattern than that of typical mitosis in plants, however, eventually following the similar trends of mitosis in animals where H3S10 phosphorylation begins in the pericentromeric regions first, later extending to the whole chromosomes, and finally declining at anaphase/early cytokinesis (differentiation of vegetative and generative cells). The study suggests that the chromosomal distribution of H3S10ph during cell division is not universal and can be altered between different cell types encoded for diverse cellular processes. During pollen development, phosphorylation of histone might play a critical role in chromosome condensation events throughout pollen mitosis I in plants.     

Antiviral Effect of Theaflavins on Tobacco Mosaic Virus

Theaflavins, the oxidized products formed from tea leaf catechins during black tea fermentation, showed an antiviral activity on TMV. From the survey of the interactions of theaflavins with RNA and its related substances, it was presumed that theaflavins disturbed the replication cycle of TMV through binding to TMV-RNA.