Characterization of plasma triiodophenol binding proteins in vertebrates and tissue distribution of triiodophenol in Rana catesbeiana tadpoles

We investigated the interaction of 2,4,6-triiodophenol (TIP), a potent thyroid hormone disrupting chemical, with serum proteins from rainbow trout (Onchorhynchus mykiss), bullfrog (Rana catesbeiana), chicken (Gallus gallus), pig (Sus scrofa domesticus), and rat (Rattus norvegicus) using a [(125)I]TIP binding assay, gel filtration chromatography, and native polyacrylamide gel electrophoresis. [(125)I]TIP bound non-specifically to proteins in trout serum, specifically but weakly to proteins in bullfrog serum, and specifically and strongly to proteins in chicken, pig, and rat serum samples. Candidate TIP-binding proteins included lipoproteins (220-320kDa) in trout, albumin in bullfrog, albumin and transthyretin (TTR) in chicken and pig, and TTR in rat. TTR in the chicken, pig, and rat serum samples was responsible for the high-affinity, low-capacity binding sites for TIP (dissociation constant 2.2-3.5×10(-10)M). In contrast, a weak interaction of [(125)I]TIP with tadpole serum proteins accelerated [(125)I]TIP cellular uptake in vitro. Intraperitoneal injection of [(125)I]TIP in tadpoles revealed that the radioactivity was predominantly accumulated in the gallbladder and the kidney. The differences in the molecular and binding properties of TIP binding proteins among vertebrates would affect in part the cellular availability, tissue distribution and clearance of TIP.     

ACh对人垂体腺瘤细胞内蛋白激酶C、胞内游离钙及cAMP/cGMP的影响

我们曾发现ACh可明显地抑制垂体腺瘤细胞的增殖代谢,为深入探讨ACh抑制垂体腺瘤细胞增殖作用的机制,观察了ACh作用后垂体腺瘤细胞内蛋白激酶C(PKC)、[Ca^2 ]i及cAMP／cGMP的变化。结果发现：(1)与空白处理组相比,使用PKC的激动剂PMA处理培养的人垂体腺瘤细胞时可使胞浆、胞膜和细胞总PKC活性浓度均升高,但ACh(10μmol/L)作用15min后,胞浆、胞膜和细胞总PKC活性均下降,且此作用可被阿托品阻断;(2)ACh(10μmol/L)作用于单个人垂体腺瘤细胞后,立即使垂体腺瘤细胞[Ca^2 ]i相对水平降低,但此作用可被阿托品阻断;(3)ACh作用于人垂体腺瘤细胞15min后,胞内cAMP水平均明显升高,而cGMP没有改变。该结果为探讨ACh抑制垂体腺瘤细胞增殖的分子机制提供了重要线索,同时提示,ACh对垂体瘤细胞增殖分化的调控作用是细胞内多信息系统相互整合的结果。     

Partial purification,subunit structure and thermal stability of the photochemical reaction center of the thermophilic green bacterium Chloroflexus aurantiacus

Spectrally pure reaction center preparations from Chloroflexus aurantiacus have been obtained in a stable form; however, the product contained several contaminating polypeptides. The reaction center pigment molecules (probably three bacteriochlorophyll a and three bacteriopheophytin a molecules) are associated with two polypeptides (M_r = 30000 and 28000) in a reaction center complex of M_r = 52000. No carotenoid is present in the complex. These data together with previous spectral data suggest that the Chloroflexus reaction center represents a more primitive evolutionary form of the purple bacterial reaction center, and that it has little if any relationship to the green bacterial component. A reaction center preparation from Rhodopseudomonas sphaeroides R26 was fully denatured at 50°C while the Chloroflexus reaction center required higher temperatures (70–75°C) for complete denaturation. Thus, an intrinsic membrane protein of a photosynthetic thermophile has been demonstrated to have greater thermal stability than the equivalent component of a mesophile.     

Botulinum toxin's axonal transport from periphery to the spinal cord

Axonal transport of enzymatically active botulinum toxin A (BTX-A) from periphery to the CNS has been described in facial and trigeminal nerve, leading to cleavage of synaptosomal-associated protein 25 (SNAP-25) in central nuclei. Aim of present study was to examine the existence of axonal transport of peripherally applied BTX-A to spinal cord via sciatic nerve. We employed BTX-A-cleaved SNAP-25 immunohistochemistry of lumbar spinal cord after intramuscular and subcutaneous hind limb injections, and intraneural BTX-A sciatic nerve injections. Truncated SNAP-25 in ipsilateral spinal cord ventral horns and dorsal horns appeared after single peripheral BTX-A administrations, even at low intramuscular dose applied (5 U/kg). Cleaved SNAP-25 appearance in the spinal cord after BTX-A injection into the sciatic nerve was prevented by proximal intrasciatic injection of colchicine (5 mM, 2 μl). Cleaved SNAP-25 in ventral horn, using choline-acetyltransferase (ChAT) double labeling, was localized within cholinergic neurons. These results extend the recent findings on BTX-A retrograde axonal transport in facial and trigeminal nerve. Appearance of truncated SNAP-25 in spinal cord following low-dose peripheral BTX-A suggest that the axonal transport of BTX-A occurs commonly following peripheral application.     

Fungal communities living within leaves of native Hawaiian dicots are structured by landscape‐scale variables as well as by host plants

A phylogenetically diverse array of fungi live within healthy leaf tissue of dicotyledonous plants. Many studies have examined these endophytes within a single plant species and/or at small spatial scales, but landscape‐scale variables that determine their community composition are not well understood, either across geographic space, across climatic conditions, or in the context of host plant phylogeny. Here, we evaluate the contributions of these variables to endophyte beta diversity using a survey of foliar endophytic fungi in native Hawaiian dicots sampled across the Hawaiian archipelago. We used Illumina technology to sequence fungal ITS1 amplicons to characterize foliar endophyte communities across five islands and 80 host plant genera. We found that communities of foliar endophytic fungi showed strong geographic structuring between distances of 7 and 36 km. Endophyte community structure was most strongly associated with host plant phylogeny and evapotranspiration, and was also significantly associated with NDVI, elevation and solar radiation. Additionally, our bipartite network analysis revealed that the five islands we sampled each harboured significantly specialized endophyte communities. These results demonstrate how the interaction of factors at large and small spatial and phylogenetic scales shapes fungal symbiont communities.     

硒对NO诱导的内皮细胞内游离钙离子浓度变化的影响

用Fura-2显微荧光测钙技术,研究了用外源性一氧化氮（NO）供体S-亚硝基谷胱甘肽（GSNO）诱导的,人脐静脉内皮细胞系ECV-304细胞胞内游离钙离子浓度（［Ca²⁺］_i ）升高以及硒的抑制效应.结果表明,GSNO作用于ECV-304细胞,短时间内即可导致其胞内游离钙离子浓度升高.胞外液换为无钙液或向胞外液中加入CdCl₂（1 mmol/L）对GSNO引起的［Ca²⁺］_i升高无影响.提示,GSNO刺激主要引起胞内钙库释放.而且,一氧化氮清除剂血红蛋白（Hb）对这一过程有抑制作用,说明GSNO引起的胞内钙库释放由NO介导.经亚硒酸钠（1 μmol/L）处理的细胞,其NO引起的［Ca²⁺］_i升高幅度明显被抑制,说明NO的这种作用可能与细胞的氧化还原状态有关.     

Molecular determination of kleptoplast origins from the sea slug Plakobranchus ocellatus (Sacoglossa,Gastropoda) reveals cryptic bryopsidalean (Chlorophyta) diversity in the Hawaiian Islands

The sacoglossan sea slug species complex Plakobranchus ocellatus is a common algivore throughout the tropical Pacific, including the Hawaiian Islands. Plakobranchus ocellatus is kleptoplastic—it sequesters and retains algal chloroplasts—a characteristic that can be exploited to molecularly characterize diminutive bryopsidalean algae that are typically difficult to locate, collect, and identify. Previous DNA barcode analyses of both P. ocellatus and its kleptoplasts have been conducted primarily in the western Pacific and have only minimally sampled the most eastern populations in the Hawaiian Islands. Using two chloroplast markers, rbcL and tufA, kleptoplast samples from an Oahu population of P. ocellatus were amplified and cloned to identify their algal sources. Plakobranchus ocellatus sequester chloroplasts from up to 11 bryopsidalean algal species, all but one being diminutive in thallus size. Notably, eight of the detected algal species were new records to the Hawaiian Islands. A sequestration preference study demonstrated that the O‘ahu population of P. ocellatus preferentially sequesters chloroplasts from diminutive, epilithic taxa. Using coxI barcoding of P. ocellatus, we showed the O‘ahu population to be part of a clade that includes sequences from the neighboring island Maui, Australia, and the Philippines. The use of P. ocellatus as a novel sampling tool allows the exploration of the green algal community diversity and composition at a fine scale.     

Cofilin-1 is involved in regulation of actin reorganization during influenza A virus assembly and budding

Influenza A virus (IAV) assembly and budding on host cell surface plasma membrane requires actin cytoskeleton reorganization. The underlying molecular mechanism involving actin reorganization remains unclarified. In this study, we found that the natural antiviral compound petagalloyl glucose (PGG) inhibits F-actin reorganization in the host cell membrane during the late stage of IAV infection, which are associated with the suppression of total cofilin-1 level and its phosphorylation. Knock-down of cofilin-1 reduces viral yields. These findings provide the first evidence that cofilin-1 plays an important role in regulating actin reorganization during IAV assembly and budding.     

Immune response to human influenza virus hemagglutinin expressed in Escherichia coli

Cloned DNA fragments coding for parts of strain WSN (H1N1) influenza virus hemagglutinin (HA) were fused to a bacterial leader DNA derived from the Escherichia coli trp operon. Fusion proteins produced consisted of 190 amino acids of trpLE' protein at the amino terminus, and HA amino acids, either 1-308, 1-396, or 1-548 (complete HA), at the carboxyl terminus. These proteins were expressed at high levels (10-20% of total protein) in E. coli starved for tryptophan. A CNBr fragment (HA1-211) was derived from HA-308. Each of the proteins was purified and used for immunizing mice and rabbits. The antibody produced was shown to bind to (i) the HA fusion proteins, (ii) detergent-treated viral HA, (iii) HA, on intact virions, and (iv) the HA on the surface of cells infected with influenza virus. This shows that the HA fusion proteins expressed in bacteria can elicit antibodies that recognize at least some determinants of the native viral HA, and probably could lead to development of an anti-influenza vaccine.