Pyrrospirones A and B, apoptosis inducers in HL-60 cells, from an endophytic fungus, Neonectria ramulariae Wollenw KS-246

Pyrrospirones A and B have been isolated from unpolished rice cultures of the endophytic fungus Neonectria ramulariae Wollenw KS-246. Their absolute stereostructures (1 and 2) were elucidated through spectroscopic methods using 1D and 2D NMR techniques and chemical transformations, including the modified Mosher's method. The compounds exhibited cytotoxicity and induced apoptosis in human promyelocytic leukemia HL-60 cells.     

Unfolding and aggregation of transthyretin by the truncation of 50 N-terminal amino acids

Senile systemic amyloidosis (SSA) is caused by amyloid deposits of wild-type transthyretin in various organs. Amyloid deposits from SSA contain large amounts of the C-terminal fragments starting near amino acid residue 50 as well as full-length transthyretin. Although a number of previous studies suggest the importance of the C-terminal fragments in the pathogenesis of SSA, little is known about the structure and aggregation properties of the C-terminal fragments of transthyretin. To understand the role of C-terminal fragments in SSA, we examined the effects of the truncation of the N-terminal portions on the structure and aggregation properties of wild-type transthyretin. The deletion mutant lacking 50 N-terminal residues was largely unfolded in terms of secondary and tertiary structure, leading to self-assembly into spherical aggregations under nearly physiological conditions. By contrast, the deletion mutant lacking 37 N-terminal residues did not have a strong tendency to aggregate, although it also adopted a largely unfolded conformation. These results suggest that global unfolding of transthyretin by proteolysis near amino acid residue 50 is an important step of self-assembly into aggregations in SSA.     

Synthesis,SAR, and X-ray structure of tricyclic compounds as potent FBPase inhibitors

With the aim of discovering a novel class of fructose-1,6-bisphosphatase (FBPase) inhibitors, a series of compounds based on tricyclic scaffolds was synthesized. Extensive SAR studies led to the finding of 8l with an IC₅₀ value of 0.013 μM against human FBPase. An X-ray crystallographic study revealed that 8l bound at AMP binding sites of human liver FBPase with hydrogen bonding interactions similar to AMP.     

Constant association of ophiostomatoid fungi with the bark beetle <Emphasis Type="Italic">Ips subelongatus</Emphasis> invading Japanese larch logs

Ceratocystiopsis, Ceratocystis, Grosmannia, and Ophiostoma species were isolated from Ips subelongatus and beetle-infested Japanese larch logs collected at several areas in central and northern Honshu Island, Japan, to determine constant associates of I. subelongatus. Ceratocystiopsis minuta, two species of Ceratocystis, three species of Grosmannia, and four species of Ophiostoma were isolated. Of the fungi isolated in the present study, G. laricis, O. brunneociliatum, and O. piceae were constant associates of the beetles. Ceratocystis fujiensis, Ceratocystiopsis minuta, and Ophiostoma sp. F were occasionally isolated with high frequencies of occurrence but were not consistent associates. Ceratocystis fujiensis was most often isolated as the leading fungal invasion in the sapwood of Japanese larch logs invaded by I. subelongatus, confirming that the fungus acts as a primary invader of sapwood in beetle-attacked logs. Contribution No. 221, Laboratory of Plant Parasitic Mycology, Graduate School of Life and Environmental Sciences, University of Tsukuba     

Temperature Dependence of Single Molecule Rotation of the Escherichia coli ATP Synthase F1 Sector Reveals the Importance of ��-�� Subunit Interactions in the Catalytic Dwell

The temperature-dependent rotation of F₁-ATPase γ subunit was observed in V_max conditions at low viscous drag using a 60-nm gold bead (Nakanishi-Matsui, M., Kashiwagi, S., Hosokawa, H., Cipriano, D. J., Dunn, S. D., Wada, Y., and Futai, M. (2006) J. Biol. Chem. 281, 4126–4131). The Arrhenius slopes of the speed of the individual 120° steps and reciprocal of the pause length between rotation steps were very similar, indicating a flat energy pathway followed by the rotationally coupled catalytic cycle. In contrast, the Arrhenius slope of the reciprocal pause length of the γM23K mutant F₁ was significantly increased, whereas that of the rotation rate was similar to wild type. The effects of the rotor γM23K substitution and the counteracting effects of βE381D mutation in the interacting stator subunits demonstrate that the rotor-stator interactions play critical roles in the utilization of stored elastic energy. The γM23K enzyme must overcome an abrupt activation energy barrier, forcing it onto a less favored pathway that results in uncoupling catalysis from rotation.F-ATPase (F_oF₁), consisting of the catalytic sector F₁ (α₃β₃γδϵ) and the transmembrane proton transport sector F_o (ab₂c₁₀), synthesizes or hydrolyzes ATP coupled with proton transport (for reviews, see Ref. 1–6). As Abrahams et al. (7) discovered in the first high resolution x-ray structure, a critical feature of the F₁-ATPase is the inherent asymmetry of the three β subunits in different conformations, β_TP, β_DP, and β_E, referring to the nucleotide bound in each catalytic site, ATP, ADP, or empty, respectively. A rotational mechanism has been firmly established mostly based on direct observation in single molecule experiments of the behavior of the rotor complex ϵγc₁₀, relative to the stator complex α₃β₃δab₂ (reviewed in Ref. 1). ATP hydrolysis-dependent rotation of the γ and ϵ subunits in purified bacterial F₁ (8, 9), the ϵγc₁₀ complex in detergent solubilized F_oF₁ (10–13), and the ϵγc₁₀ complexin F_oF₁ in lipid bilayers (14) were shown experimentally by single molecule observations using fluorescent actin filament as a probe. Relative rotation of the single copy F_o a subunit was also shown in F₀F₁, which was immobilized through the ring of ∼10 c subunits, suggesting that the rotor and stator are interchangeable mechanical units (14). ATP synthesis by F-ATPase is believed to follow the reverse mechanism of ATP hydrolysis because mechanically induced rotation of the γ subunit in immobilized F₁ in the presence of ADP and P_i results in net ATP synthesis (15, 16). There remain many questions about the mechanism of coupling between catalysis and transport via mechanical rotation. In particular, the mechanism of coupling H⁺ transport to rotation of the subunit c₁₀ ring is still not well understood (4).In contrast, there is considerably more information on the mechanism of coupling catalysis to γ and ϵ subunit rotation. Observations of γ subunit rotation in the catalytic F₁ sector are consistent with Boyer''s binding change model (17); thus coupling between the chemistry and rotation can be assessed by studies of the soluble F₁, and these findings relate to the mechanism of the entire ATP synthase complex. The γ subunit rotates relative to the α₃β₃ hexamer in distinct 120° steps. A 120° rotation step consisting of pause and rotation substeps appears to correspond to the hydrolysis of one ATP, assuming that three ATP molecules are hydrolyzed per 360° revolution (18). Additional pauses observed at low ATP concentrations are attributed to the “ATP waiting” dwell (19). Yasuda et al. (19) and Shimabukuro et al. (20) further resolved that each 120° step occurred in two substeps: an 80° substep whose onset was dependent upon the Mg·ATP concentration, and a 40° substep, which was not affected by substrate concentration (19). The pause before the 80° substep, the ATP waiting dwell became shorter with increasing [Mg·ATP]. In contrast, the pause duration before the 40° rotation step was modulated by the slow hydrolysis rate of ATPγS² or by the catalytic site mutant βE190D (in the Bacillus PS3 F₁), which was found to significantly increase the length of the catalytic dwell (20). These data together indicate that the dwell before the 40° step is the “catalytic dwell” (20) and defines the order of the substeps during the 120° rotation step observed in high Mg·ATP concentrations (21).In this paper, we address the question of when the rate-limiting step of steady state catalysis occurs, with respect to the rotational behavior. Pre-steady state analysis of the burst kinetics of ATP hydrolysis at nearly V_max conditions demonstrated that the rate-limiting transition state occurs after the reversible hydrolysis/synthesis step and before release of phosphate (P_i) (22, 23). The rate-limiting step is likely associated with a rotation step because a γ-β cross-linked enzyme is still able to undergo the initial ATP hydrolysis, but the rotation-impeded enzyme is unable to release P_i (23). Significantly, the kinetics of steady state hydrolysis can only be assessed when the Mg·ATP concentration is high enough to fill all three catalytic sites. The only model consistent with these data is one that involves all three catalytic sites. During each 120° catalytic cycle, one site binds ATP, a different site carries out reversible hydrolysis/synthesis, and the third site releases product P_i and ADP (22, 23).Steady state analyses, which take advantage of a particular γ subunit mutation γM23K (24), are consistent with this model. Replacement of the conserved γMet-23 with lysine causes an uncoupling between catalysis and γ subunit rotation caused by altered interactions between γ and β subunits (25). Importantly, Al-Shawi and Nakamoto (26) and Al-Shawi et al. (25, 27) found that the γM23K mutation strongly affected the rate-limiting transition state of steady state ATP hydrolysis and ATP synthesis. The slope of the Arrhenius plots and thus the energy of activation were significantly increased in the mutant enzyme. Several second site suppressor mutations, mostly in the γ subunit (28, 29) but also in the β subunits (30, 31), were genetically identified because they restored coupled ATP synthesis. Significantly, all were in the γ-β interface. Thermodynamic analyses found that the second site suppressors generally compensated for the primary γM23K mutations by reducing the increased activation energy (25, 27, 31). Although most of the second site mutations were found distant from the γM23K site, the x-ray crystal structures (7) suggested that γM23K may directly interact with conserved βGlu-381. As expected, replacement of βGlu-381 with aspartate also suppressed the uncoupling effects of γM23K (31).To identify the rate-limiting transition state step in the rotational behavior, we analyzed the temperature dependence of the γM23K mutant in V_max conditions observed in single molecule experiments. Interestingly, direct observation of this mutant using the micron-length actin filaments did not detect differences in the rotation behavior at room temperature (9). In contrast, we find in the data presented here that there is dramatic effect of the mutation on the temperature dependence of the length of the catalytic dwell or pause between the 120° rotation steps. This is likely because of two factors: first, we used a bead small enough not to invoke a drag on the rotation (32), and second, the temperature dependence of the rate of the rotation steps is critical for the analyses of the mechanism.     

Review of Indo-Pacific dwarf lionfishes (Scorpaenidae: Pteroinae) in the <Emphasis Type="Italic">Dendrochirus brachypterus</Emphasis> complex,with description of a new species from the western Indian Ocean

A taxonomic review of the Dendrochirus brachypterus complex resulted in the recognition of five species, including Dendrochirus barberi (Steindachner 1900), Dendrochirus bellus (Jordan and Hubbs 1925), Dendrochirus brachypterus (Cuvier in Cuvier and Valenciennes 1829), Dendrochirus hemprichi sp. nov. and Dendrochirus tuamotuensis Matsunuma and Motomura 2013. The complex is defined as having usually 9 dorsal-fin soft rays, usually 5 anal-fin soft rays, 17–20 (rarely 20) pectoral-fin rays, no ocellated spots on the soft-rayed portion of the dorsal fin and usually 2 (sometimes none) barbels on the snout tip. Dendrochirus barberi, known from the Hawaiian Islands and Johnston Atoll, is characterized by usually 18 pectoral-fin rays, a relatively high number of scale rows in the longitudinal series (modally 51 vs. 39–49 in other species) and mottled markings on the pectoral fin in large specimens. Dendrochirus bellus, restricted to the northwestern Pacific Ocean from the South China Sea north to southern Japan, is characterized by usually 17 pectoral-fin rays, a relatively low number of scale rows in the longitudinal series (modally 38 vs. 44–51 in other species), and the absence of skin flaps on the orbit surface and uppermost preopercular spine base. Dendrochirus tuamotuensis, recorded only from the Tuamotu Archipelago, is characterized by 19 pectoral-fin rays, the posterior margin of the pectoral fin strongly notched, and a relatively shallow and narrow head and body. Dendrochirus hemprichi sp. nov. is distributed in the western Indian Ocean, including the Red Sea. Although previously confused with a closely related congener (D. brachypterus, known from the northern and eastern Indian Ocean and western Pacific), D. hemprichi can be distinguished from the former by having fewer scale rows between the last dorsal-fin spine base and lateral line, and between the sixth dorsal-fin spine base and lateral line [4–7 (5) in D. hemprichi vs. 5–7 (6) in D. brachypterus, in both cases], a slightly greater interorbital width at the mid-orbit [5.5–10.7 (mean 7.8) % SL vs. 4.5–8.9 (6.8) % of SL] and at preocular spine base [4.4–9.1 (6.6) % SL vs. 3.5–7.8 (5.7) % of SL], and slightly shorter posteriormost (usually 13th) dorsal-fin spine length [11.8–19.9 (15.3) % SL vs. 13.3–21.3 (17.2) % of SL]. Moreover, D. hemprichi tends to have relatively more spinous points on the head spines and ridges, compared with D. brachypterus. Notwithstanding the morphological similarity between the two species, an obvious genetic difference was observed between D. hemprichi and D. brachypterus. Dendrochirus chloreus Jenkins 1903 and Dendrochirus hudsoni Jordan and Evermann 1903 were synonymized under Pterois barberi, as in some previous studies. Scorpaena koenigii Bloch 1789 was regarded as conspecific with D. brachypterus, which it predated. However, the former name should be suppressed under Reversal of Precedence.     

Review of the genus <Emphasis Type="Italic">Banjos</Emphasis> (Perciformes: Banjosidae) with descriptions of two new species and a new subspecies

A taxonomic review of the genus Banjos (Perciformes: Banjosidae), previously restricted to a single species, Banjos banjos (Richardson 1846), recorded from the northwestern Pacific Ocean from the South China Sea north to Japan, as well as Lombok (Indonesia), New Caledonia and Australia, resulted in the recognition of three species, including B. banjos (northwestern Pacific Ocean, Indonesia and western Australia), Banjos aculeatus sp. nov. (eastern Australia) and Banjos peregrinus sp. nov. [northern Australia (Timor Sea)]. Records of B. banjos from New Caledonia probably also represent B. aculeatus, which is clearly distinct from other congeners in having a relatively long, strongly serrated spine at the posteroventral angle of the preopercle and an entirely dusky membrane on the spinous dorsal fin in juveniles < ca. 70 mm SL, in addition to slightly longer first and second dorsal-fin spines. Banjos peregrinus is characterized by a relatively greater head length, orbit diameter, postorbital length and pre-pelvic-fin length, as well as poorly developed serration of the exposed margin of the cleithrum. Within B. banjos, a population from the southeastern Indian Ocean, including Indonesia and western Australia, is regarded as a distinct subspecies (Banjos banjos brevispinis ssp. nov.), distinguishable from B. b. banjos from the northwestern Pacific Ocean by a relatively narrow least interorbital width, and shorter second and eighth dorsal-fin spines. Ontogenetic morphological changes within the genus and the status of the holotype of Anoplus banjos Richardson 1846 are discussed in detail.     

A screening system for artificial small RNAs that inhibit the growth of Escherichia coli

We have developed a screening system for artificial small RNAs (sRNAs) that inhibit the growth of Escherichia coli. In this system, we used a plasmid library to express artificial sRNAs (approximately 200 bases long) containing 60 bases of random nucleotide sequence. The induced expression of the known rydB sRNA in the system reduced the amount of its possible target mRNA, rpoS, supporting the reliability of the method. To isolate clones of sRNAs that inhibited the growth of E. coli, we used two successive screening steps: (i) colony size selection on plates and (ii) monitoring E. coli growth in a 96-well plate format. As a result, 83 artificial sRNAs were identified that showed a range of inhibitory effects on bacterial growth. We also introduced nucleotide replacements into one of the highly inhibitory sRNA clones, H12, which partially abolished the inhibition of bacterial growth, suggesting that bacterial growth was inhibited in a sequence-specific manner.     

Self-Organization on Social Media: Endo-Exo Bursts and Baseline Fluctuations

A salient dynamic property of social media is bursting behavior. In this paper, we study bursting behavior in terms of the temporal relation between a preceding baseline fluctuation and the successive burst response using a frequency time series of 3,000 keywords on Twitter. We found that there is a fluctuation threshold up to which the burst size increases as the fluctuation increases and that above the threshold, there appears a variety of burst sizes. We call this threshold the critical threshold. Investigating this threshold in relation to endogenous bursts and exogenous bursts based on peak ratio and burst size reveals that the bursts below this threshold are endogenously caused and above this threshold, exogenous bursts emerge. Analysis of the 3,000 keywords shows that all the nouns have both endogenous and exogenous origins of bursts and that each keyword has a critical threshold in the baseline fluctuation value to distinguish between the two. Having a threshold for an input value for activating the system implies that Twitter is an excitable medium. These findings are useful for characterizing how excitable a keyword is on Twitter and could be used, for example, to predict the response to particular information on social media.