Functional diversification of closely related ARF-GEFs in protein secretion and recycling

Guanine-nucleotide exchange factors on ADP-ribosylation factor GTPases (ARF-GEFs) regulate vesicle formation in time and space by activating ARF substrates on distinct donor membranes. Mammalian GBF1 (ref. 2) and yeast Gea1/2 (ref. 3) ARF-GEFs act at Golgi membranes, regulating COPI-coated vesicle formation. In contrast, their Arabidopsis thaliana homologue GNOM (GN) is required for endosomal recycling, playing an important part in development. This difference indicates an evolutionary divergence of trafficking pathways between animals and plants, and raised the question of how endoplasmic reticulum-Golgi transport is regulated in plants. Here we demonstrate that the closest homologue of GNOM in Arabidopsis, GNOM-LIKE1 (GNL1; NM_123312; At5g39500), performs this ancestral function. GNL1 localizes to and acts primarily at Golgi stacks, regulating COPI-coated vesicle formation. Surprisingly, GNOM can functionally substitute for GNL1, but not vice versa. Our results suggest that large ARF-GEFs of the GBF1 class perform a conserved role in endoplasmic reticulum-Golgi trafficking and secretion, which is done by GNL1 and GNOM in Arabidopsis, whereas GNOM has evolved to perform an additional plant-specific function of recycling from endosomes to the plasma membrane. Duplication and diversification of ARF-GEFs in plants contrasts with the evolution of entirely new classes of ARF-GEFs for endosomal trafficking in animals, which illustrates the independent evolution of complex endosomal pathways in the two kingdoms.     

A flagellin-induced complex of the receptor FLS2 and BAK1 initiates plant defence

Plants sense potential microbial invaders by using pattern-recognition receptors to recognize pathogen-associated molecular patterns (PAMPs). In Arabidopsis thaliana, the leucine-rich repeat receptor kinases flagellin-sensitive 2 (FLS2) (ref. 2) and elongation factor Tu receptor (EFR) (ref. 3) act as pattern-recognition receptors for the bacterial PAMPs flagellin and elongation factor Tu (EF-Tu) (ref. 5) and contribute to resistance against bacterial pathogens. Little is known about the molecular mechanisms that link receptor activation to intracellular signal transduction. Here we show that BAK1 (BRI1-associated receptor kinase 1), a leucine-rich repeat receptor-like kinase that has been reported to regulate the brassinosteroid receptor BRI1 (refs 6,7), is involved in signalling by FLS2 and EFR. Plants carrying bak1 mutations show normal flagellin binding but abnormal early and late flagellin-triggered responses, indicating that BAK1 acts as a positive regulator in signalling. The bak1-mutant plants also show a reduction in early, but not late, EF-Tu-triggered responses. The decrease in responses to PAMPs is not due to reduced sensitivity to brassinosteroids. We provide evidence that FLS2 and BAK1 form a complex in vivo, in a specific ligand-dependent manner, within the first minutes of stimulation with flagellin. Thus, BAK1 is not only associated with developmental regulation through the plant hormone receptor BRI1 (refs 6,7), but also has a functional role in PRR-dependent signalling, which initiates innate immunity.     

Aggregation and vesiculation of membrane proteins by curvature-mediated interactions

Membrane remodelling plays an important role in cellular tasks such as endocytosis, vesiculation and protein sorting, and in the biogenesis of organelles such as the endoplasmic reticulum or the Golgi apparatus. It is well established that the remodelling process is aided by specialized proteins that can sense as well as create membrane curvature, and trigger tubulation when added to synthetic liposomes. Because the energy needed for such large-scale changes in membrane geometry significantly exceeds the binding energy between individual proteins and between protein and membrane, cooperative action is essential. It has recently been suggested that curvature-mediated attractive interactions could aid cooperation and complement the effects of specific binding events on membrane remodelling. But it is difficult to experimentally isolate curvature-mediated interactions from direct attractions between proteins. Moreover, approximate theories predict repulsion between isotropically curving proteins. Here we use coarse-grained membrane simulations to show that curvature-inducing model proteins adsorbed on lipid bilayer membranes can experience attractive interactions that arise purely as a result of membrane curvature. We find that once a minimal local bending is realized, the effect robustly drives protein cluster formation and subsequent transformation into vesicles with radii that correlate with the local curvature imprint. Owing to its universal nature, curvature-mediated attraction can operate even between proteins lacking any specific interactions, such as newly synthesized and still immature membrane proteins in the endoplasmic reticulum.     

A high-resolution environmental change record since 19 cal ka BP in Pumoyum Co,southern Tibet

A 380-cm-long sediment core was acquired from the deep water area of Pumoyum Co, southern Tibet. Twenty-five plant residue samples were selected, and organic carbon stable isotopes were obtained using the AMS ¹⁴C chronological method. The ¹⁴C age and carbon reservoir effect were calibrated with surface sedimentation rate measurements using ²¹⁰Pb dating. Results showed that the core sediment deposited over 19 cal ka BP. Based on a multi-proxy analysis of TOC and IC contents, grain size and pollen assemblage data, the palaeoclimatic evolution of Pumoyum Co was reconstructed since the last glacial. Pumoyum Co was a shallow lake prior to 16.2 cal ka BP; although the glacier around the lake began to melt due to increasing temperatures, climate was still cold and dry. In the interval of 16.2–11.8 cal ka BP, the sedimentary environment fluctuated drastically and frequently. Two cold-events occurred at 14.2 and 11.8 cal ka BP, and these may correspond to the Older Dryas and the Younger Dryas events, respectively. After 11.8 cal ka BP, Pumoyun Co developed into the deep lake as it is now. The lake water temperature was relatively lower at that time because of influx of cold water from glacial meltwater entering the lake. As a result, the multi-proxy indicators showed no sign of warm conditions. Comparisons between the sedimentary record of Pumoyum Co with that of other lakes of the same age in southern Tibet indicate a warmer climate following the last deglaciation influenced the southeastern Tibetan Plateau. These results imply that the southwest Asian monsoon gradually became stronger since the deglaciation during its expansion to the inner plateau. The glacial-supplied water of the lake responded sensitively to cold-events. The entire southern Tibet region was dominantly influenced climatically by the southwest Asian monsoon during the Holocene.     

Genome sequence and analysis of the tuber crop potato

Potato (Solanum tuberosum L.) is the world's most important non-grain food crop and is central to global food security. It is clonally propagated, highly heterozygous, autotetraploid, and suffers acute inbreeding depression. Here we use a homozygous doubled-monoploid potato clone to sequence and assemble 86% of the 844-megabase genome. We predict 39,031 protein-coding genes and present evidence for at least two genome duplication events indicative of a palaeopolyploid origin. As the first genome sequence of an asterid, the potato genome reveals 2,642 genes specific to this large angiosperm clade. We also sequenced a heterozygous diploid clone and show that gene presence/absence variants and other potentially deleterious mutations occur frequently and are a likely cause of inbreeding depression. Gene family expansion, tissue-specific expression and recruitment of genes to new pathways contributed to the evolution of tuber development. The potato genome sequence provides a platform for genetic improvement of this vital crop.     

Acetylation-dependent regulation of endothelial Notch signalling by the SIRT1 deacetylase

Notch signalling is a key intercellular communication mechanism that is essential for cell specification and tissue patterning, and which coordinates critical steps of blood vessel growth. Although subtle alterations in Notch activity suffice to elicit profound differences in endothelial behaviour and blood vessel formation, little is known about the regulation and adaptation of endothelial Notch responses. Here we report that the NAD(+)-dependent deacetylase SIRT1 acts as an intrinsic negative modulator of Notch signalling in endothelial cells. We show that acetylation of the Notch1 intracellular domain (NICD) on conserved lysines controls the amplitude and duration of Notch responses by altering NICD protein turnover. SIRT1 associates with NICD and functions as a NICD deacetylase, which opposes the acetylation-induced NICD stabilization. Consequently, endothelial cells lacking SIRT1 activity are sensitized to Notch signalling, resulting in impaired growth, sprout elongation and enhanced Notch target gene expression in response to DLL4 stimulation, thereby promoting a non-sprouting, stalk-cell-like phenotype. In vivo, inactivation of Sirt1 in zebrafish and mice causes reduced vascular branching and density as a consequence of enhanced Notch signalling. Our findings identify reversible acetylation of the NICD as a molecular mechanism to adapt the dynamics of Notch signalling, and indicate that SIRT1 acts as rheostat to fine-tune endothelial Notch responses.     

Observed increase in local cooling effect of deforestation at higher latitudes

Deforestation in mid- to high latitudes is hypothesized to have the potential to cool the Earth's surface by altering biophysical processes. In climate models of continental-scale land clearing, the cooling is triggered by increases in surface albedo and is reinforced by a land albedo-sea ice feedback. This feedback is crucial in the model predictions; without it other biophysical processes may overwhelm the albedo effect to generate warming instead. Ongoing land-use activities, such as land management for climate mitigation, are occurring at local scales (hectares) presumably too small to generate the feedback, and it is not known whether the intrinsic biophysical mechanism on its own can change the surface temperature in a consistent manner. Nor has the effect of deforestation on climate been demonstrated over large areas from direct observations. Here we show that surface air temperature is lower in open land than in nearby forested land. The effect is 0.85 ± 0.44 K (mean ± one standard deviation) northwards of 45° N and 0.21 ± 0.53 K southwards. Below 35° N there is weak evidence that deforestation leads to warming. Results are based on comparisons of temperature at forested eddy covariance towers in the USA and Canada and, as a proxy for small areas of cleared land, nearby surface weather stations. Night-time temperature changes unrelated to changes in surface albedo are an important contributor to the overall cooling effect. The observed latitudinal dependence is consistent with theoretical expectation of changes in energy loss from convection and radiation across latitudes in both the daytime and night-time phase of the diurnal cycle, the latter of which remains uncertain in climate models.     

Genetic variants in novel pathways influence blood pressure and cardiovascular disease risk

Blood pressure is a heritable trait influenced by several biological pathways and responsive to environmental stimuli. Over one billion people worldwide have hypertension (≥140?mm?Hg systolic blood pressure or ≥90?mm?Hg diastolic blood pressure). Even small increments in blood pressure are associated with an increased risk of cardiovascular events. This genome-wide association study of systolic and diastolic blood pressure, which used a multi-stage design in 200,000 individuals of European descent, identified sixteen novel loci: six of these loci contain genes previously known or suspected to regulate blood pressure (GUCY1A3-GUCY1B3, NPR3-C5orf23, ADM, FURIN-FES, GOSR2, GNAS-EDN3); the other ten provide new clues to blood pressure physiology. A genetic risk score based on 29 genome-wide significant variants was associated with hypertension, left ventricular wall thickness, stroke and coronary artery disease, but not kidney disease or kidney function. We also observed associations with blood pressure in East Asian, South Asian and African ancestry individuals. Our findings provide new insights into the genetics and biology of blood pressure, and suggest potential novel therapeutic pathways for cardiovascular disease prevention.     

CENP-B preserves genome integrity at replication forks paused by retrotransposon LTR

Centromere-binding protein B (CENP-B) is a widely conserved DNA binding factor associated with heterochromatin and centromeric satellite repeats. In fission yeast, CENP-B homologues have been shown to silence long terminal repeat (LTR) retrotransposons by recruiting histone deacetylases. However, CENP-B factors also have unexplained roles in DNA replication. Here we show that a molecular function of CENP-B is to promote replication-fork progression through the LTR. Mutants have increased genomic instability caused by replication-fork blockage that depends on the DNA binding factor switch-activating protein 1 (Sap1), which is directly recruited by the LTR. The loss of Sap1-dependent barrier activity allows the unhindered progression of the replication fork, but results in rearrangements deleterious to the retrotransposon. We conclude that retrotransposons influence replication polarity through recruitment of Sap1 and transposition near replication-fork blocks, whereas CENP-B counteracts this activity and promotes fork stability. Our results may account for the role of LTR in fragile sites, and for the association of CENP-B with pericentromeric heterochromatin and tandem satellite repeats.