The reversibility of mitotic exit in vertebrate cells

A guiding hypothesis for cell-cycle regulation asserts that regulated proteolysis constrains the directionality of certain cell-cycle transitions. Here we test this hypothesis for mitotic exit, which is regulated by degradation of the cyclin-dependent kinase 1 (Cdk1) activator, cyclin B. Application of chemical Cdk1 inhibitors to cells in mitosis induces cytokinesis and other normal aspects of mitotic exit, including cyclin B degradation. However, chromatid segregation fails, resulting in entrapment of chromatin in the midbody. If cyclin B degradation is blocked with a proteasome inhibitor or by expression of non-degradable cyclin B, Cdk inhibitors will nonetheless induce mitotic exit and cytokinesis. However, if after mitotic exit, the Cdk1 inhibitor is washed free from cells in which cyclin B degradation is blocked, the cells can revert back to M phase. This reversal is characterized by chromosome recondensation, nuclear envelope breakdown, assembly of microtubules into a mitotic spindle, and in most cases, dissolution of the midbody, reopening of the cleavage furrow, and realignment of chromosomes at the metaphase plate. These findings demonstrate that proteasome-dependent degradation of cyclin B provides directionality for the M phase to G1 transition.     

Inhibition of Dll4 signalling inhibits tumour growth by deregulating angiogenesis

Haploinsufficiency of Dll4, a vascular-specific Notch ligand, has shown that it is essential for embryonic vascular development and arteriogenesis. Mechanistically, it is unclear how the Dll4-mediated Notch pathway contributes to complex vascular processes that demand meticulous coordination of multiple signalling pathways. Here we show that Dll4-mediated Notch signalling has a unique role in regulating endothelial cell proliferation and differentiation. Neutralizing Dll4 with a Dll4-selective antibody rendered endothelial cells hyperproliferative, and caused defective cell fate specification or differentiation both in vitro and in vivo. In addition, blocking Dll4 inhibited tumour growth in several tumour models. Remarkably, antibodies against Dll4 and antibodies against vascular endothelial growth factor (VEGF) had paradoxically distinct effects on tumour vasculature. Our data also indicate that Dll4-mediated Notch signalling is crucial during active vascularization, but less important for normal vessel maintenance. Furthermore, unlike blocking Notch signalling globally, neutralizing Dll4 had no discernable impact on intestinal goblet cell differentiation, supporting the idea that Dll4-mediated Notch signalling is largely restricted to the vascular compartment. Therefore, targeting Dll4 might represent a broadly efficacious and well-tolerated approach for the treatment of solid tumours.     

Palaeoceanography: methane release in the Early Jurassic period

Dramatic global warming, triggered by release of methane from clathrates, has been postulated to have occurred during the early Toarcian age in the Early Jurassic period. Kemp et al. claim that this methane was released at three points, as recorded by three sharp excursions of delta13C(org) of up to 3 per thousand magnitude. But they discount another explanation for the excursions: namely that some, perhaps all, of the rapid excursions could be a local signature of a euxinic basin caused by recycling of isotopically light carbon from the lower water column. This idea has been proposed previously (see ref. 3, for example) and is supported by the lack evidence for negative delta13C excursions in coeval belemnite rostra. Kemp et al. dismiss this alternative, claiming that each abrupt shift would have required the recycling of about double the amount of organic carbon that is currently present in the modern ocean; however, their measurements are not from an ocean but from a restricted, epicontinental seaway and so would not require whole-ocean mixing to achieve the excursions.     

Disruption of extended defects in solid oxide fuel cell anodes for methane oxidation

Point defects largely govern the electrochemical properties of oxides: at low defect concentrations, conductivity increases with concentration; however, at higher concentrations, defect-defect interactions start to dominate. Thus, in searching for electrochemically active materials for fuel cell anodes, high defect concentration is generally avoided. Here we describe an oxide anode formed from lanthanum-substituted strontium titanate (La-SrTiO3) in which we control the oxygen stoichiometry in order to break down the extended defect intergrowth regions and create phases with considerable disordered oxygen defects. We substitute Ti in these phases with Ga and Mn to induce redox activity and allow more flexible coordination. The material demonstrates impressive fuel cell performance using wet hydrogen at 950 degrees C. It is also important for fuel cell technology to achieve efficient electrode operation with different hydrocarbon fuels, although such fuels are more demanding than pure hydrogen. The best anode materials to date--Ni-YSZ (yttria-stabilized zirconia) cermets--suffer some disadvantages related to low tolerance to sulphur, carbon build-up when using hydrocarbon fuels (though device modifications and lower temperature operation can avoid this) and volume instability on redox cycling. Our anode material is very active for methane oxidation at high temperatures, with open circuit voltages in excess of 1.2 V. The materials design concept that we use here could lead to devices that enable more-efficient energy extraction from fossil fuels and carbon-neutral fuels.     

A single amino acid governs enhanced activity of DinB DNA polymerases on damaged templates

Translesion synthesis (TLS) by Y-family DNA polymerases is a chief mechanism of DNA damage tolerance. Such TLS can be accurate or error-prone, as it is for bypass of a cyclobutane pyrimidine dimer by DNA polymerase eta (XP-V or Rad30) or bypass of a (6-4) TT photoproduct by DNA polymerase V (UmuD'2C), respectively. Although DinB is the only Y-family DNA polymerase conserved among all domains of life, the biological rationale for this striking conservation has remained enigmatic. Here we report that the Escherichia coli dinB gene is required for resistance to some DNA-damaging agents that form adducts at the N2-position of deoxyguanosine (dG). We show that DinB (DNA polymerase IV) catalyses accurate TLS over one such N2-dG adduct (N2-furfuryl-dG), and that DinB and its mammalian orthologue, DNA polymerase kappa, insert deoxycytidine (dC) opposite N2-furfuryl-dG with 10-15-fold greater catalytic proficiency than opposite undamaged dG. We also show that mutating a single amino acid, the 'steric gate' residue of DinB (Phe13 --> Val) and that of its archaeal homologue Dbh (Phe12 --> Ala), separates the abilities of these enzymes to perform TLS over N2-dG adducts from their abilities to replicate an undamaged template. We propose that DinB and its orthologues are specialized to catalyse relatively accurate TLS over some N2-dG adducts that are ubiquitous in nature, that lesion bypass occurs more efficiently than synthesis on undamaged DNA, and that this specificity may be achieved at least in part through a lesion-induced conformational change.     

The PerR transcription factor senses H2O2 by metal-catalysed histidine oxidation

The sensing of reactive oxygen species is essential for cellular responses to oxidative stress. The sensing of peroxides is typically mediated by redox-active cysteines in sensors such as the bacterial OxyR, OhrR, and Hsp33 proteins. Bacillus subtilis PerR is the prototype for a widespread family of metal-dependent peroxide sensors that regulate inducible peroxide-defence genes. Here we show that PerR senses peroxides by metal-catalysed oxidation. PerR contains two metal-binding sites: a structural Zn2+ site and a regulatory divalent metal ion site that preferentially binds Fe2+ or Mn2+ (ref. 5). Protein oxidation, catalysed by a bound ferrous ion, leads to the rapid and direct incorporation of one oxygen atom into histidine 37 (H37) or H91, two of the residues that coordinate the bound Fe2+. This mechanism accounts for the ability of PerR to sense low levels of hydrogen peroxide in vivo. The reduction of hydrogen peroxide by metal ions to generate highly reactive hydroxyl radicals underlies the genotoxic effects of peroxides, and has been shown to contribute to enzyme inactivation, but has not previously been shown to provide a regulatory mechanism for peroxide sensing.     

Editing-defective tRNA synthetase causes protein misfolding and neurodegeneration

Misfolded proteins are associated with several pathological conditions including neurodegeneration. Although some of these abnormally folded proteins result from mutations in genes encoding disease-associated proteins (for example, repeat-expansion diseases), more general mechanisms that lead to misfolded proteins in neurons remain largely unknown. Here we demonstrate that low levels of mischarged transfer RNAs (tRNAs) can lead to an intracellular accumulation of misfolded proteins in neurons. These accumulations are accompanied by upregulation of cytoplasmic protein chaperones and by induction of the unfolded protein response. We report that the mouse sticky mutation, which causes cerebellar Purkinje cell loss and ataxia, is a missense mutation in the editing domain of the alanyl-tRNA synthetase gene that compromises the proofreading activity of this enzyme during aminoacylation of tRNAs. These findings demonstrate that disruption of translational fidelity in terminally differentiated neurons leads to the accumulation of misfolded proteins and cell death, and provide a novel mechanism underlying neurodegeneration.     

The structure of H5N1 avian influenza neuraminidase suggests new opportunities for drug design

The worldwide spread of H5N1 avian influenza has raised concerns that this virus might acquire the ability to pass readily among humans and cause a pandemic. Two anti-influenza drugs currently being used to treat infected patients are oseltamivir (Tamiflu) and zanamivir (Relenza), both of which target the neuraminidase enzyme of the virus. Reports of the emergence of drug resistance make the development of new anti-influenza molecules a priority. Neuraminidases from influenza type A viruses form two genetically distinct groups: group-1 contains the N1 neuraminidase of the H5N1 avian virus and group-2 contains the N2 and N9 enzymes used for the structure-based design of current drugs. Here we show by X-ray crystallography that these two groups are structurally distinct. Group-1 neuraminidases contain a cavity adjacent to their active sites that closes on ligand binding. Our analysis suggests that it may be possible to exploit the size and location of the group-1 cavity to develop new anti-influenza drugs.     

Structural asymmetry and the stability of diverse food webs

Untangling the influence of human activities on food-web stability and persistence is complex given the large numbers of species and overwhelming number of interactions within ecosystems. Although biodiversity has been associated with stability, the actual structures and processes that confer stability to diverse food webs remain largely unknown. Here we show that real food webs are structured such that top predators act as couplers of distinct energy channels that differ in both productivity and turnover rate. Our theoretical analysis shows that coupled fast and slow channels convey both local and non-local stability to food webs. Alarmingly, the same human actions that have been implicated in the loss of biodiversity also directly erode the very structures and processes that we show to confer stability on food webs.     

Fishing elevates variability in the abundance of exploited species

The separation of the effects of environmental variability from the impacts of fishing has been elusive, but is essential for sound fisheries management. We distinguish environmental effects from fishing effects by comparing the temporal variability of exploited versus unexploited fish stocks living in the same environments. Using the unique suite of 50-year-long larval fish surveys from the California Cooperative Oceanic Fisheries Investigations we analyse fishing as a treatment effect in a long-term ecological experiment. Here we present evidence from the marine environment that exploited species exhibit higher temporal variability in abundance than unexploited species. This remains true after accounting for life-history effects, abundance, ecological traits and phylogeny. The increased variability of exploited populations is probably caused by fishery-induced truncation of the age structure, which reduces the capacity of populations to buffer environmental events. Therefore, to avoid collapse, fisheries must be managed not only to sustain the total viable biomass but also to prevent the significant truncation of age structure. The double jeopardy of fishing to potentially deplete stock sizes and, more immediately, to amplify the peaks and valleys of population variability, calls for a precautionary management approach.