Multi-domain conformational selection underlies pre-mRNA splicing regulation by U2AF

Many cellular functions involve multi-domain proteins, which are composed of structurally independent modules connected by flexible linkers. Although it is often well understood how a given domain recognizes a cognate oligonucleotide or peptide motif, the dynamic interaction of multiple domains in the recognition of these ligands remains to be characterized. Here we have studied the molecular mechanisms of the recognition of the 3'-splice-site-associated polypyrimidine tract RNA by the large subunit of the human U2 snRNP auxiliary factor (U2AF65) as a key early step in pre-mRNA splicing. We show that the tandem RNA recognition motif domains of U2AF65 adopt two remarkably distinct domain arrangements in the absence or presence of a strong (that is, high affinity) polypyrimidine tract. Recognition of sequence variations in the polypyrimidine tract RNA involves a population shift between these closed and open conformations. The equilibrium between the two conformations functions as a molecular rheostat that quantitatively correlates the natural variations in polypyrimidine tract nucleotide composition, length and functional strength to the efficiency to recruit U2 snRNP to the intron during spliceosome assembly. Mutations that shift the conformational equilibrium without directly affecting RNA binding modulate splicing activity accordingly. Similar mechanisms of cooperative multi-domain conformational selection may operate more generally in the recognition of degenerate nucleotide or amino acid motifs by multi-domain proteins.     

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.     

Comprehensive analysis of the chromatin landscape in Drosophila melanogaster

Chromatin is composed of DNA and a variety of modified histones and non-histone proteins, which have an impact on cell differentiation, gene regulation and other key cellular processes. Here we present a genome-wide chromatin landscape for Drosophila melanogaster based on eighteen histone modifications, summarized by nine prevalent combinatorial patterns. Integrative analysis with other data (non-histone chromatin proteins, DNase I hypersensitivity, GRO-Seq reads produced by engaged polymerase, short/long RNA products) reveals discrete characteristics of chromosomes, genes, regulatory elements and other functional domains. We find that active genes display distinct chromatin signatures that are correlated with disparate gene lengths, exon patterns, regulatory functions and genomic contexts. We also demonstrate a diversity of signatures among Polycomb targets that include a subset with paused polymerase. This systematic profiling and integrative analysis of chromatin signatures provides insights into how genomic elements are regulated, and will serve as a resource for future experimental investigations of genome structure and function.     

Senescence surveillance of pre-malignant hepatocytes limits liver cancer development

Upon the aberrant activation of oncogenes, normal cells can enter the cellular senescence program, a state of stable cell-cycle arrest, which represents an important barrier against tumour development in vivo. Senescent cells communicate with their environment by secreting various cytokines and growth factors, and it was reported that this 'secretory phenotype' can have pro- as well as anti-tumorigenic effects. Here we show that oncogene-induced senescence occurs in otherwise normal murine hepatocytes in vivo. Pre-malignant senescent hepatocytes secrete chemo- and cytokines and are subject to immune-mediated clearance (designated as 'senescence surveillance'), which depends on an intact CD4(+) T-cell-mediated adaptive immune response. Impaired immune surveillance of pre-malignant senescent hepatocytes results in the development of murine hepatocellular carcinomas (HCCs), thus showing that senescence surveillance is important for tumour suppression in vivo. In accordance with these observations, ras-specific Th1 lymphocytes could be detected in mice, in which oncogene-induced senescence had been triggered by hepatic expression of Nras(G12V). We also found that CD4(+) T cells require monocytes/macrophages to execute the clearance of senescent hepatocytes. Our study indicates that senescence surveillance represents an important extrinsic component of the senescence anti-tumour barrier, and illustrates how the cellular senescence program is involved in tumour immune surveillance by mounting specific immune responses against antigens expressed in pre-malignant senescent cells.     

Detection of prokaryotic mRNA signifies microbial viability and promotes immunity

Live vaccines have long been known to trigger far more vigorous immune responses than their killed counterparts. This has been attributed to the ability of live microorganisms to replicate and express specialized virulence factors that facilitate invasion and infection of their hosts. However, protective immunization can often be achieved with a single injection of live, but not dead, attenuated microorganisms stripped of their virulence factors. Pathogen-associated molecular patterns (PAMPs), which are detected by the immune system, are present in both live and killed vaccines, indicating that certain poorly characterized aspects of live microorganisms, not incorporated in dead vaccines, are particularly effective at inducing protective immunity. Here we show that the mammalian innate immune system can directly sense microbial viability through detection of a special class of viability-associated PAMPs (vita-PAMPs). We identify prokaryotic messenger RNA as a vita-PAMP present only in viable bacteria, the recognition of which elicits a unique innate response and a robust adaptive antibody response. Notably, the innate response evoked by viability and prokaryotic mRNA was thus far considered to be reserved for pathogenic bacteria, but we show that even non-pathogenic bacteria in sterile tissues can trigger similar responses, provided that they are alive. Thus, the immune system actively gauges the infectious risk by searching PAMPs for signatures of microbial life and thus infectivity. Detection of vita-PAMPs triggers a state of alert not warranted for dead bacteria. Vaccine formulations that incorporate vita-PAMPs could thus combine the superior protection of live vaccines with the safety of dead vaccines.     

Heart regeneration

Heart failure plagues industrialized nations, killing more people than any other disease. It usually results from a deficiency of specialized cardiac muscle cells known as cardiomyocytes, and a robust therapy to regenerate lost myocardium could help millions of patients every year. Heart regeneration is well documented in amphibia and fish and in developing mammals. After birth, however, human heart regeneration becomes limited to very slow cardiomyocyte replacement. Several experimental strategies to remuscularize the injured heart using adult stem cells and pluripotent stem cells, cellular reprogramming and tissue engineering are in progress. Although many challenges remain, these interventions may eventually lead to better approaches to treat or prevent heart failure.     

Corridors of migrating neurons in the human brain and their decline during infancy

The subventricular zone of many adult non-human mammals generates large numbers of new neurons destined for the olfactory bulb. Along the walls of the lateral ventricles, immature neuronal progeny migrate in tangentially oriented chains that coalesce into a rostral migratory stream (RMS) connecting the subventricular zone to the olfactory bulb. The adult human subventricular zone, in contrast, contains a hypocellular gap layer separating the ependymal lining from a periventricular ribbon of astrocytes. Some of these subventricular zone astrocytes can function as neural stem cells in vitro, but their function in vivo remains controversial. An initial report found few subventricular zone proliferating cells and rare migrating immature neurons in the RMS of adult humans. In contrast, a subsequent study indicated robust proliferation and migration in the human subventricular zone and RMS. Here we find that the infant human subventricular zone and RMS contain an extensive corridor of migrating immature neurons before 18 months of age but, contrary to previous reports, this germinal activity subsides in older children and is nearly extinct by adulthood. Surprisingly, during this limited window of neurogenesis, not all new neurons in the human subventricular zone are destined for the olfactory bulb--we describe a major migratory pathway that targets the prefrontal cortex in humans. Together, these findings reveal robust streams of tangentially migrating immature neurons in human early postnatal subventricular zone and cortex. These pathways represent potential targets of neurological injuries affecting neonates.