A genome-wide BAC end-sequence survey of sugarcane elucidates genome composition, and identifies BACs covering much of the euchromatin

BAC-end sequences (BESs) of hybrid sugarcane cultivar R570 are presented. A total of 66,990 informative BESs were obtained from 43,874 BAC clones. Similarity search using a variety of public databases revealed that 13.5 and 42.8 % of BESs match known gene-coding and repeat regions, respectively. That 11.7 % of BESs are still unmatched to any nucleotide sequences in the current public databases despite the fact that a close relative, sorghum, is fully sequenced, indicates that there may be many sugarcane-specific or lineage-specific sequences. We found 1,742 simple sequence repeat motifs in 1,585 BESs, spanning 27,383 bp in length. As simple sequence repeat markers derived from BESs have some advantages over randomly generated markers, these may be particularly useful for comparing BAC-based physical maps with genetic maps. BES and overgo hybridization information was used for anchoring sugarcane BAC clones to the sorghum genome sequence. While sorghum and sugarcane have extensive similarity in terms of genomic structure, only 2,789 BACs (6.4 %) could be confidently anchored to the sorghum genome at the stringent threshold of having both-end information (BESs or overgos) within 300 Kb. This relatively low rate of anchoring may have been caused in part by small- or large-scale genomic rearrangements in the Saccharum genus after two rounds of whole genome duplication since its divergence from the sorghum lineage about 7.8 million years ago. Limiting consideration to only low-copy matches, 1,245 BACs were placed to 1,503 locations, covering ~198 Mb of the sorghum genome or about 78 % of the estimated 252 Mb of euchromatin. BESs and their analyses presented here may provide an early profile of the sugarcane genome as well as a basis for BAC-by-BAC sequencing of much of the basic gene set of sugarcane.     

Lack of genetic differentiation between monarch butterflies with divergent migration destinations

Monarch butterflies are best known for their spectacular annual migration from eastern North America to Mexico. Monarchs also occur in the North American states west of the Rocky Mountains, from where they fly shorter distances to the California Coast. Whether eastern and western North American monarchs form one genetic population or are genetically differentiated remains hotly debated, and resolution of this debate is essential to understand monarch migration patterns and to protect this iconic insect species. We studied the genetic structure of North American migratory monarch populations, as well as nonmigratory populations in Hawaii and New Zealand. Our results show that eastern and western migratory monarchs form one admixed population and that monarchs from Hawaii and New Zealand have genetically diverged from North American butterflies. These findings suggest that eastern and western monarch butterflies maintain their divergent migrations despite genetic mixing. The finding that eastern and western monarchs form one genetic population also suggests that the conservation of overwintering sites in Mexico is crucial for the protection of monarchs in both eastern and western North America.     

A multi-study analysis of gut microbiome data from the blue mussel (Mytilus edulis) emphasises the impact of depuration on biological interpretation

The blue mussel (Mytilus edulis) is a suspension feeder which has been used in gut-microbiome surveys. Although raw 16S sequence data are often publicly available, unifying secondary analyses are lacking. The present work analysed raw data from seven projects conducted by one group over 7 years. Although each project had different motivations, experimental designs and conclusions, all selected samples were from the guts of M. edulis collected from a single location in Long Island Sound. The goal of this analysis was to determine which independent factors (e.g., collection date, depuration status) were responsible for governing composition and diversity in the gut microbiomes. Results indicated that whether mussels had undergone depuration, defined here as voidance of faeces in a controlled, no-food period, was the primary factor that governed gut microbiome composition. Gut microbiomes from non-depurated mussels were mixtures of resident and transient communities and were influenced by temporal factors. Resident communities from depurated mussels were influenced by the final food source and length of time host mussels were held under laboratory conditions. These findings reinforce the paradigm that gut microbiota are divided into resident and transient components and suggest that depuration status should be taken into consideration when designing and interpreting future experiments.     

Ultrastructure of the heart of the marine mussel,Geukensia demissa

The structure of the heart of Geukensia demissa, a common object of physiological and biochemical investigation, is described by scanning, transmission and freeze-fracture electron microscopy. A single-cell epithelial layer covers the ventricle, but an endothelium is lacking. Myofibers are small (6–7 μm diam.), mononucleate, and tapered. Glycogen is concentrated peripherally. Mitochondria are particularly concentrated under the sarcolemma, near the ends of the nucleus, and in rows between bundles of myofilaments. The myofilaments (6–8nm thin, 30–35 nm thick filament diam.) are loosely arranged into sarcomeres (2–4 μm) by Z bodies. Many of these Z bodies interconnect, and some anchor to the sarcolemma forming attachment plaques. Cells are joined by intercalated discs consisting of fascia adherentes, spot desmosomes, and gap junctions. The gap junctions include intramembrane particles. T tubules are absent. The sarcolemma is coupled to the junctional sarcoplasmic reticulum (JSR) over 357ndash;40% of the cell surface. Tubules extend from the JSR deep into and throughout the cell as an irregularly dispersed network. The SR occupies 1% of the cell volume. A few, small (0.1–1.0 μm) unmyelinated nerves are present, but no neuromuscular junctions were seen. The auricles have fewer and smaller myocytes than the ventricle. The auricles also contain podocytes with pedicels having 20–35 nm slits and containing sieve-like projections. The morphology of the Geukensia heart is similar to that of other bivalves.     

Inhibition of cell metabolism by a smokeless tobacco extract: tissue and species specificity.

Smokeless tobacco contains a nonnicotine inhibitor of posttranslational modification of collagen (hydroxylation of [3H]proline) by cultured chick embryo tibias and osteoblasts. This study was undertaken to determine whether a methanol extract of smokeless tobacco (STE) containing the inhibitor has similar effects on collagen-producing cells and tissues other than bone. Its effects on DNA synthesis and cell proliferation (incorporation of [3H]thymidine) were also determined. Frontal bone, aorta, and cartilage were incubated for 2 days in medium containing STE. Glycolysis (lactate production) was stimulated by 80% in cartilage, but was not affected in the other tissues; medium alkaline phosphatase activity was unaffected. In frontal bone and cartilage, [3H] hydroxyproline content was decreased 88% and 57%, respectively, and [3H]proline content was decreased 68% and 37%, respectively; neither was affected in the aorta. Confluent cultures of collagen-producing mouse fibroblasts or primary osteoblasts obtained from chick embryo calvarias were incubated for 2 days in medium containing increasing concentrations of STE. Glycolysis and DNA synthesis were not affected. Cell proliferation was unaffected in fibroblasts, but was inhibited (34%) at the highest STE concentration in osteoblasts. AIPase activity was not detectable in fibroblast medium, but was decreased up to 72% in osteoblast medium. Inhibition of collagen synthesis by STE was concentration related in both cell types. At the highest concentration, [3H] hydroxyproline and [3H]proline contents in the cell layers were decreased to the following respective values: fibroblasts 56% and 45% and osteoblasts 50% and 29%, respectively. When incubation with STE was discontinued for 1 day, recovery did not occur. These findings suggest that inhibition of collagen synthesis by STE is not specific for bone, that collagen-producing cells are directly affected, and that recovery is not immediate. This inhibitor could contribute to the periodontal disease often seen in users of smokeless tobacco. Its identification and removal would produce a safer product.     

Integrin-dependent neuroblastoma cell adhesion and migration on laminin is regulated by expression levels of two enzymes in the O-mannosyl-linked glycosylation pathway, PomGnT1 and GnT-Vb

O-mannosyl-linked glycans constitute a third of all brain O-linked glycoproteins, and yet very little is understood about their functions. Several congenital muscular dystrophies with central nervous system defects are caused by genetic disruptions in glycosyltransferases responsible for the synthesis of O-mannosyl glycans. The glycosyltransferase GnT-Vb, also known as GnT-IX, is expressed abundantly in the brain and testis and is proposed to be the enzyme that branches O-mannosyl-linked glycans. In this study, we show in a human neuronal model that GnT-Vb expression enhances neurite outgrowth on laminin. GnT-Vb has been shown to perform both N-linked and O-mannosyl-linked glycosylation. To determine if the effect on neurite outgrowth was due to N-linked or O-mannosyl-linked glycosylation by GnT-Vb we suppressed the expression of glycosyltransferases important for the elongation of both N-linked and O-mannosyl-linked glycans using RNA interference. Our results suggest that GnT-Vb and PomGnT1, enzymes involved in the O-mannosyl glycosylation pathway, play an active role in modulating integrin and laminin-dependent adhesion and migration of human neuronal cells.     

Local intrahepatic CD8+ T cell activation by a non-self-antigen results in full functional differentiation

The response of T cells to liver Ags sometimes results in immune tolerance. This has been proposed to result from local, intrahepatic priming, while the expression of the same Ag in liver-draining lymph nodes is believed to result in effective immunity. We tested this model, using an exogenous model Ag expressed only in hepatocytes, due to infection with an adeno-associated virus vector. T cell activation was exclusively intrahepatic, yet in contrast to the predictions of the current model, this resulted in clonal expansion, IFN-gamma synthesis, and cytotoxic effector function. Local activation of naive CD8(+) T cells can therefore cause full CD8(+) T cell activation, and hepatocellular presentation cannot be used to explain the failure of CTL effector function against some liver pathogens such as hepatitis C.