Smooth muscle-endothelial cell communication activates Reelin signaling and regulates lymphatic vessel formation

Active lymph transport relies on smooth muscle cell (SMC) contractions around collecting lymphatic vessels, yet regulation of lymphatic vessel wall assembly and lymphatic pumping are poorly understood. Here, we identify Reelin, an extracellular matrix glycoprotein previously implicated in central nervous system development, as an important regulator of lymphatic vascular development. Reelin-deficient mice showed abnormal collecting lymphatic vessels, characterized by a reduced number of SMCs, abnormal expression of lymphatic capillary marker lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1), and impaired function. Furthermore, we show that SMC recruitment to lymphatic vessels stimulated release and proteolytic processing of endothelium-derived Reelin. Lymphatic endothelial cells in turn responded to Reelin by up-regulating monocyte chemotactic protein 1 (MCP1) expression, which suggests an autocrine mechanism for Reelin-mediated control of endothelial factor expression upstream of SMC recruitment. These results uncover a mechanism by which Reelin signaling is activated by communication between the two cell types of the collecting lymphatic vessels--smooth muscle and endothelial cells--and highlight a hitherto unrecognized and important function for SMCs in lymphatic vessel morphogenesis and function.     

X-ray diffraction study of a new crystal form of the nucleosome core showing higher resolution

Crystals have been grown of intact (unproteolysed) nucleosome cores from a variety of sources. The unit cells are all very similar, with one core particle per asymmetric unit. The X-ray diffraction patterns extend to about 5 Å in the direction perpendicular to the plane of the flat particle, and to somewhat less than this in other directions. The arrangement of particles in the unit cell has been deduced from Patterson projection maps, which also indicate the presence of a particle dyad. The data are consistent with the earlier proposed model for the core particle in which the 146 base-pairs of DNA are wound in about $\text{1}\text{3}\text{4}$ turns of superhelix about a histone octamer core.High angle diffuse X-ray scattering from the crystals shows that the DNA of the core particle is in the B form. The anisotropy of the diffuse scattering shows that the DNA is not firmly fixed to the histone core all along the superhelix path, but only over limited regions whose location correlates well with those in which the DNA is differentially protected against nuclease digestion.     

A Rapid Ribosome Profiling Method Elucidates Chloroplast Ribosome Behavior in Vivo

The profiling of ribosome footprints by deep sequencing has revolutionized the analysis of translation by mapping ribosomes with high resolution on a genome-wide scale. We present a variation on this approach that offers a rapid and cost-effective alternative for the genome-wide profiling of chloroplast ribosomes. Ribosome footprints from leaf tissue are hybridized to oligonucleotide tiling microarrays of the plastid ORFeome and report the abundance and translational status of every chloroplast mRNA. Each assay replaces several time-consuming traditional methods while also providing information that was previously inaccessible. To illustrate the utility of the approach, we show that it detects known defects in chloroplast gene expression in several nuclear mutants of maize (Zea mays) and that it reveals previously unsuspected defects. Furthermore, it provided firm answers to several lingering questions in chloroplast gene expression: (1) the overlapping atpB/atpE open reading frames, whose translation had been proposed to be coupled, are translated independently in vivo; (2) splicing is not a prerequisite for translation initiation on an intron-containing chloroplast RNA; and (3) a feedback control mechanism that links the synthesis of ATP synthase subunits in Chlamydomonas reinhardtii does not exist in maize. An analogous approach is likely to be useful for studies of mitochondrial gene expression.     

Inherent asymmetry of the structure of F1-ATPase from bovine heart mitochondria at 6.5 A resolution.

ATP synthase, the assembly which makes ATP in mitochondria, chloroplasts and bacteria, uses transmembrane proton gradients generated by respiration or photosynthesis to drive the phosphorylation of ADP. Its membrane domain is joined by a slender stalk to a peripheral catalytic domain, F1-ATPase. This domain is made of five subunits with stoichiometries of 3 alpha: 3 beta: 1 gamma: 1 delta: 1 epsilon, and in bovine mitochondria has a molecular mass of 371,000. We have determined the 3-dimensional structure of bovine mitochondrial F1-ATPase to 6.5 A resolution by X-ray crystallography. It is an approximately spherical globule 110 A in diameter, on a 40 A stem which contains two alpha-helices in a coiled-coil. This stem is presumed to be part of the stalk that connects F1 with the membrane domain in the intact ATP synthase. A pit next to the stem penetrates approximately 35 A into the F1 particle. The stem and the pit are two examples of the many asymmetric features of the structure. The central element in the asymmetry is the longer of the two alpha-helices in the stem, which extends for 90 A through the centre of the assembly and emerges on top into a dimple 15 A deep. Features with threefold and sixfold symmetry, presumed to be parts of homologous alpha and beta subunits, are arranged around the central rod and pit, but the overall structure is asymmetric. The central helix provides a possible mechanism for transmission of conformational changes induced by the proton gradient from the stalk to the catalytic sites of the enzyme.     

Phytoplankton dynamics and sedimentation processes during spring and summer in Balsfjord,Northern Norway

Summary Chlorophyll a, phytoplankton species composition and carbon (PPC) estimated from cell-counts, were monitored together with hydrographic parameters and nutrients in the upper 50 m of Balsfjord (ca. 70° N), northern Norway between 08 February and 29 June 1982. Sediment traps were placed at 10, 50, 100, and 170 m (10 m above bottom) for intervals of 5–20 days during the study period. Trap contents were analyzed for phytoplankton as above; dry weight, particulate organic material (POM), particulate organic nitrogen and carbon (PON and POC), ash, and particulate phosphorus were also measured. The phytoplankton community exhibited three main phases: During the first (02–15 April, chiefly surface biomass) and the second (20 April–10 May, deep biomass-maximum and spring bloom peak) periods, Phaeocystis pouchetii dominated biomass (ca. 50% of PPC) followed by vegetative cells of Chaetoceros socialis. In the third period (10 May onwards, characterized by surface estuarinecir-culation), dino- and microflagellates dominated the low post-bloom biomass. Protozooplankton comprising tintinnids, other ciliates and heterotrophic dinoflagellates increased in abundance. Vegetative cells of phytoplankton were scarce in trap collections at 50 m or below; resting cells of Chaetoceros comprised nearly all the intact sedimenting phytoplankton. Krill faeces accounted for >90% by volume of the total faecal material trapped, despite a >21 biomass dominance of copepods in the fjord. The greatest sedimentation rates of krill faeces were at > 100 m, reflecting the downward migration of krill during the day. In all, 2–3 g Cm^–2 of krill faeces were collected, representing ca. twice that from intact phytoplankton cells. POC in the traps at 50 m was ca. 11 gm^–2, accounting for ca. 17% of the estimated primary production during the study period. As the secondary production is high, a large proportion of the production of P. pouchetii must be grazed by herbivores. Copepod faeces are probably remineralized in the euphotic zone, while those of krill provide the major coupling between the pelagial and the benthos. The implications of such a sedimentation model for partitioning energy flow between the pelagial and the benthos is discussed.     

The molecular mechanism of induction of unfolded protein response by Chlamydia

The unfolded protein response (UPR) contributes to chlamydial pathogenesis, as a source of lipids and ATP during replication, and for establishing the initial anti-apoptotic state of host cell that ensures successful inclusion development. The molecular mechanism(s) of UPR induction by Chlamydia is unknown. Chlamydia use type III secretion system (T3SS) effector proteins (e.g, the Translocated Actin-Recruiting Phosphoprotein (Tarp) to stimulate host cell's cytoskeletal reorganization that facilitates invasion and inclusion development. We investigated the hypothesis that T3SS effector-mediated assembly of myosin-II complex produces activated non-muscle myosin heavy chain II (NMMHC-II), which then binds the UPR master regulator (BiP) and/or transducers to induce UPR. Our results revealed the interaction of the chlamydial effector proteins (CT228 and Tarp) with components of the myosin II complex and UPR regulator and transducer during infection. These interactions caused the activation and binding of NMMHC-II to BiP and IRE1α leading to UPR induction. In addition, specific inhibitors of myosin light chain kinase, Tarp oligomerization and myosin ATPase significantly reduced UPR activation and Chlamydia replication. Thus, Chlamydia induce UPR through T3SS effector-mediated activation of NMMHC-II components of the myosin complex to facilitate infectivity. The finding provides greater insights into chlamydial pathogenesis with the potential to identify therapeutic targets and formulations.