Alkenone and alkenoic acid compositions of the membrane fractions of Emiliania huxleyi

The lipid classes and unsaturation ratios of long-chain alkenones (nC37-C39), related alkyl alkenoate compounds (nC37-C38) and alkenoic acids (nC14-C22) were determined in isolated membrane and organelle fractions of Emiliania huxleyi. The percentage distribution of these compounds was predominantly high in the endoplasmic reticulum (ER) and coccolith-producing compartment (CPC)-rich membrane fraction, although alkenones and alkenoates could be detected in all membrane fractions. In particular, the alkenones were mainly located in CPC, since their distribution was closely correlated with that of uronic acids which are markers of CPC. In contrast, the alkenoic acids seemed to be mainly located in chloroplast (thylakoid)-rich fractions. The alkenone unsaturation ratio and the ratio of alkenoates to alkenones were similar in all fractions, while the unsaturation ratio of alkenoic acids in the thylakoid-rich and plasma membrane (PM)/Golgi body-rich fractions was overwhelmingly higher than that in the ER/CPC-rich fractions. Thus, alkenoic acids seemed to be typical membrane-bound lipids, and could be closely related to photosynthesis and involved in regulating membrane fluidity and rigidity in E. huxleyi. It is presumed from these results that the alkenones and alkenoates were membrane-unbound lipids that might be associated with the function of CPC.     

Predicting subcellular localization of proteins for Gram-negative bacteria by support vector machines based on n-peptide compositions

Gram-negative bacteria have five major subcellular localization sites: the cytoplasm, the periplasm, the inner membrane, the outer membrane, and the extracellular space. The subcellular location of a protein can provide valuable information about its function. With the rapid increase of sequenced genomic data, the need for an automated and accurate tool to predict subcellular localization becomes increasingly important. We present an approach to predict subcellular localization for Gram-negative bacteria. This method uses the support vector machines trained by multiple feature vectors based on n-peptide compositions. For a standard data set comprising 1443 proteins, the overall prediction accuracy reaches 89%, which, to the best of our knowledge, is the highest prediction rate ever reported. Our prediction is 14% higher than that of the recently developed multimodular PSORT-B. Because of its simplicity, this approach can be easily extended to other organisms and should be a useful tool for the high-throughput and large-scale analysis of proteomic and genomic data.     

Localization of beta-D-glucosidase activity and glucovanillin in vanilla bean (Vanilla planifolia Andrews)

The morphology, anatomy and histology of mature green vanilla beans were examined by light and transmission electron microscopy. Beans have a triangular cross-section with a central cavity containing seeds. Each angle is lined with tubular cells, or papillae, while the cavity sides consist of placental laminae. The epicarp and endocarp are formed by one or two layers of very small cells, while the mesocarp contains large, highly vacuolarized cells, the cytoplasm being restricted to a thin layer along the cell walls. The radial distributions of glucovanillin and beta-glucosidase activity, measured on p-nitrophenyl-beta-glucopyranoside and glucovanillin, are superimposable and show how beta-glucosidase activity increases from the epicarp towards the placental zone, whereas glucovanillin is exclusively located in the placentae and papillae. Subcellular localization of beta-glucosidase activity was achieved by incubating sections of vanilla beans in a buffer containing 5-bromo-4-chloro-3-indolyl-beta-d-glucopyranoside as a substrate. Activity was observed in the cytoplasm (and/or the periplasm) of mesocarp and endocarp cells, with a more diffuse pattern observed in the papillae. A possible mechanism for the hydrolysis of glucovanillin and release of the aromatic aglycon vanillin involves the decompartmentation of cytoplasmic (and/or periplasmic) beta-glucosidase and vacuolar glucovanillin.     

Nickel Toxicity and Distribution in Maize Roots

A new histochemical method for Ni determination has been developed and employed to study the pattern of Ni distribution in plant tissues. Two-day-old seedlings of maize (Zea mays L.) were transferred onto 15, 20, 25, and 35 M Ni(NO₃)₂ solutions in the presence of 3 mM Ca(NO₃)₂, and Ni localization in shoot and root tissues was investigated at days 2 and 7 of the incubation. Following two days of incubation, Ni was found in all root tissues, and its content increased with the period of exposure and from the tip to the root base. Independent of root region and tissue, Ni content in the protoplasts exceeded that in the cell walls. Ni penetrated the endodermal barrier and accumulated in the endodermis and pericycle to the highest concentration. Ni accumulation in the pericycle restricted root branching. Ni did not affect the final cell length, and the inhibition of root growth resulted from suppressed cell division. In the shoots, Ni content was below the level discerned by the dimethylglyoximine method; we therefore conclude that maize belongs to excluder plants, with their root systems functioning as a barrier limiting heavy metal intake by aboveground organs. The pattern of Ni transport differs from that of Cd and Pb; this difference stands for specific toxic effects of Ni, including an arrest of root branching.     

Mitochondrial localization of Smad5 in a human chondrogenic cell line

Bone morphogenetic proteins (BMPs) are members of the transforming growth factor-beta (TGF-beta) superfamily and regulate the formation of cartilage and bone tissues as well as other key events during development. TGF-beta superfamily signaling is mediated intracellularly by Smad proteins, some of which can translocate into the cell nucleus and influence gene expression. Although much progress has been made in understanding how TGF-beta superfamily signaling regulates expression of target genes, little formal proof has been presented regarding the intracellular distribution of the Smad proteins before their entry into the nucleus. In the literature, non-nuclear Smad proteins are generally referred to as cytoplasmic. Using confocal microscopy, we here show for the first time that immunofluorescent labeling of Smad5, one of the Smad proteins associated with BMP signaling, colocalizes with the mitochondrion-specific probe MitoTracker, demonstrating a mitochondrial distribution of Smad5 in non-stimulated chondroprogenitor cells.