HBXIP functions as a cofactor of survivin in apoptosis suppression

Survivin is an anti-apoptotic protein that is overexpressed in most human cancers. We show that survivin forms complexes with a cellular protein, hepatitis B X-interacting protein (HBXIP), which was originally recognized for its association with the X protein of hepatitis B virus (HBX). Survivin-HBXIP complexes, but neither survivin nor HBXIP individually, bind pro-caspase-9, preventing its recruitment to Apaf1, and thereby selectively suppressing apoptosis initiated via the mitochondria/cytochrome c pathway. Viral HBX protein also interacts with the survivin- HBXIP complex and suppresses caspase activation in a survivin-dependent manner. Thus, HBXIP functions as a cofactor for survivin, and serves as a link between the cellular apoptosis machinery and a viral pathogen involved in hepatocellular carcinogenesis.     

Inactivation of Numb and Numblike in embryonic dorsal forebrain impairs neurogenesis and disrupts cortical morphogenesis

Numb and Numblike, conserved homologs of Drosophila Numb, have been implicated in cortical neurogenesis; however, analysis of their involvement in later stages of cortical development has been hampered by early lethality of double mutants in previous studies. Using Emx1(IREScre) to induce more restricted inactivation of Numb in the dorsal forebrain of numblike null mice beginning at E9.5, we have generated viable double mutants that displayed striking brain defects. It was thus possible to examine neurogenesis during the later peak phase (E12.5-E16.5). Loss of Numb and Numblike in dorsal forebrain resulted in neural progenitor hyperproliferation, delayed cell cycle exit, impaired neuronal differentiation, and concomitant defects in cortical morphogenesis. These findings reveal novel and essential function of Numb and Numblike during the peak period of cortical neurogenesis. Further, these double mutant mice provide an unprecedented viable animal model for severe brain malformations due to defects in neural progenitor cells.     

Importance of topography and soil texture in the spatial distribution of two sympatric dipterocarp trees in a Bornean rainforest

Relationships between spatial distributions and site conditions, namely topography and soil texture, were analyzed for two congeneric emergent trees, Dryobalanops aromatica and Dryobalanops lanceolata (Dipterocarpaceae), in a tropical rainforest in Sarawak, East Malaysia. A 52-ha permanent plot was divided into 1300 quadrats measuring 20m×20m; for each Dryobalanops species, the number and total basal area of trees 1cm in d.b.h. were compared among groups of quadrats with different site conditions. Because spatial distributions of both Dryobalanops and site-condition variables were aggregated, Monte-Carlo permutation tests were applied to analyze the relationships. Both single and multifactor statistical tests showed that the density and basal area distributions of the two species were significantly non-random in relation to soil texture and topographic variables. D.aromatica was significantly more abundant at higher elevations, in sandy soils, and on convex and steep slopes. In contrast, D.lanceolata preferred lower elevations and less sandy soils. In the study plot, there were very few sites (3 of 1150 quadrats tested) where the models of Hayashis method predicted the co-occurrence of the two species. These results suggest that between-species differences in habitat preferences are so large that they alone explain the spatially segregated distributions of these two species within the 52-ha study plot.     

Depolarization-induced Ca2+ release in ischemic spinal cord white matter involves L-type Ca2+ channel activation of ryanodine receptors

The mechanisms of Ca(2+) release from intracellular stores in CNS white matter remain undefined. In rat dorsal columns, electrophysiological recordings showed that in vitro ischemia caused severe injury, which persisted after removal of extracellular Ca(2+); Ca(2+) imaging confirmed that an axoplasmic Ca(2+) rise persisted in Ca(2+)-free perfusate. However, depletion of Ca(2+) stores or reduction of ischemic depolarization (low Na(+), TTX) were protective, but only in Ca(2+)-free bath. Ryanodine or blockers of L-type Ca(2+) channel voltage sensors (nimodipine, diltiazem, but not Cd(2+)) were also protective in zero Ca(2+), but their effects were not additive with ryanodine. Immunoprecipitation revealed an association between L-type Ca(2+) channels and RyRs, and immunohistochemistry confirmed colocalization of Ca(2+) channels and RyR clusters on axons. Similar to "excitation-contraction coupling" in skeletal muscle, these results indicate a functional coupling whereby depolarization sensed by L-type Ca(2+) channels activates RyRs, thus releasing damaging amounts of Ca(2+) under pathological conditions in white matter.     

Purification, crystallization and preliminary X-ray studies of a p-nitrophenylphosphatase from Bacillus stearothermophilus

Thermostable p-nitrophenylphosphatase from Bacillus stearothermophilus has been expressed in Escherichia coli, purified and crystallized. The crystals belong to space group C(2), with unit-cell parameters a = 67.17 A, b = 57.84 A, c = 62.49 A and alpha = 90.0 degrees, beta = 95.4 degrees, gamma = 90.0 degrees. Diffraction data were collected to 1.40 A resolution with a completeness of 94.7% (96.6% for the last shell), an R(fac) value of 0.074 (0.341) and an I/sigma (I) value of 30.1 (2.67).     

The Cause of the Difference in Leaf Net Photosynthetic Rate between Two Soybean Cultivars

To explore the cause of difference in photosynthetic performance between different cultivars of crops, leaf net photosynt rate (P _N) and photosystem 2 (PS2) photochemical efficiency (F_v/F_m), apparent quantum yield of carbon assimilation (_c), electron transport rate, photophosphorylation activity, etc. were measured in two soybean cultivars, Heinong 42 and Heinong 37. At pod setting and filling, significant differences in P _N between them were observed. The former with a higher P _N (from 7 to 38 %) had a significantly higher leaf thickness, leaf dry mass/area (LMA), chlorophyll content, soluble protein content, apparent quantum yield of electron transport through PS2 (_e), carboxylation efficiency (CE), and ribulose-1,5-bisphosphate carboxylase (RuBPC) activity. The significantly higher P _N of Heinong 42 is mainly due to its higher content and activity of RuBPC.     

Characterizing rice lesion mimic mutants and identifying a mutant with broad-spectrum resistance to rice blast and bacterial blight

Many plant mutants develop spontaneous lesions that resemble disease symptoms in the absence of pathogen attack. In several pathosystems, lesion mimic mutations have been shown to be involved in programmed cell death, which in some instances leads to enhanced disease resistance to multiple pathogens. We investigated the relationship between spontaneous cell death and disease resistance in rice with nine mutants with a range of lesion mimic phenotypes. All nine mutations are controlled by recessive genes and some of these mutants have stunted growth and other abnormal characteristics. The lesion mimics that appeared on the leaves of these mutants were caused by cell death as measured by trypan blue staining. Activation of six defense-related genes was observed in most of the mutants when the mimic lesions developed. Four mutants exhibited significant enhanced resistance to rice blast. One of the mutants, spl11, confers non-race-specific resistance not only to blast but also to bacterial blight. The level of resistance in the spl11 mutant to the two pathogens correlates with the defense-related gene expression and lesion development on the leaves. The results suggest that some lesion mimic mutations in rice may be involved in disease resistance, and cloning of these genes may provide a clue to developing broad-spectrum resistance to diverse pathogens.     

The quarternary molecular architecture of TetA, a secondary tetracycline transporter from Escherichia coli

TetA, a tetracycline cation/proton antiporter, was expressed in Escherichia coli with a C-terminal tag of six histidines, solubilized in dodecyl maltoside and purified in a single step using Ni2+ affinity chromatography. Two-dimensional crystals were obtained after reconstitution of purified protein with lipids. Electron microscopy of negatively stained crystals revealed a trigonal symmetry, from which we infer that this secondary transporter has a trimeric structure. An overall molecular envelope can be described by a triangle of side approximately 100 A enclosing a central stain-filled depression. These dimensions are consistent with those obtained from projection views of single, isolated TetA particles that also display a trimeric architecture, confirming that the threefold symmetry is not simply a consequence of crystal-packing interactions. These data represent the first direct view of the quarternary arrangement of any antibiotic efflux pump. They are fully consistent with biochemical data on TetA, which indicate that it functions as a multimer and that the monomer consists of two domains, one of which plays the major part in oligomerization interactions.     

Distinct topologies of mono- and decavanadate binding and photo-oxidative cleavage in the sarcoplasmic reticulum ATPase

UV irradiation of the sarcoplasmic reticulum (SR) ATPase in the presence of vanadate cleaves the enzyme at either of two different sites. Under conditions favoring the presence of monovanadate, and in the presence of Ca(2+), ADP, and Mg(2+), cleavage results in two fragments of 71- and 38-kDa electrophoretic mobility. On the other hand, under conditions permitting formation of decavanadate, and in the absence of Ca(2+) and ADP, cleavage results in two fragments of 88- and 21-kDa electrophoretic mobility. The amino terminus resulting from cleavage is blocked and resistant to Edman degradation. However, the initial photo-oxidation product can be reduced with NaB(3)H(4,) resulting in incorporation of radioactive (3)H label. Extensive digestion of the labeled protein with trypsin then yields labeled peptides that are specific for the each of the photo-oxidation conditions, and can be sequenced after purification. Collection of the Edman reaction fractional products reveals the radioactive label and demonstrates that Thr(353) is the residue oxidized by monovanadate at the phosphorylation site (i.e. Asp(351)). Correct positioning of monovanadate at the phosphorylation site requires binding of Mg(2+) and ADP to the Ca(2+)-dependent conformation of the enzyme. Subsequent hydrolytic cleavage is likely assisted by the neighboring Asp(601), and yields the 71- and 38-kDa fragments. On the other hand, Ser(186) (and possibly the following three residues: Val(187), Ile(188), and Lys(189)) is the residue that is photo-oxidized by decavanadate in the absence of ADP. Hydrolytic cleavage of the oxidized product at this site is likely assisted by neighboring acidic residues, and yields the 88- and 21-kDa fragments. The bound decavanadate, which we find to produce steric interference with TNP-AMP binding, must therefore extend to the A domain (i.e. small cytosolic loop) in order to oxidize Ser(186). This protein conformation is only obtained in the absence of Ca(2+).