Functional architecture of an intracellular membrane t-SNARE

Lipid bilayer fusion is mediated by SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) located on the vesicle membrane (v-SNAREs) and the target membrane (t-SNAREs). The assembled v-SNARE/t-SNARE complex consists of a bundle of four helices, of which one is supplied by the v-SNARE and the other three by the t-SNARE. For t-SNAREs on the plasma membrane, the protein syntaxin supplies one helix and a SNAP-25 protein contributes the other two. Although there are numerous homologues of syntaxin on intracellular membranes, there are only two SNAP-25-related proteins in yeast, Sec9 and Spo20, both of which are localized to the plasma membrane and function in secretion and sporulation, respectively. What replaces SNAP-25 in t-SNAREs of intracellular membranes? Here we show that an intracellular t-SNARE is built from a 'heavy chain' homologous to syntaxin and two separate non-syntaxin 'light chains'. SNAP-25 may thus be the exception rather than the rule, having been derived from genes that encoded separate light chains that fused during evolution to produce a single gene encoding one protein with two helices.     

Myosin V orientates the mitotic spindle in yeast

Coordination of spindle orientation with the axis of cell division is an essential process in all eukaryotes. In addition to ensuring accurate chromosomal segregation, proper spindle orientation also establishes differential cell fates and proper morphogenesis. In both animal and yeast cells, this process is dependent on cytoplasmic microtubules interacting with the cortical actin-based cytoskeleton, although the motive force was unknown. Here we show that yeast Myo2, a myosin V that translocates along polarized actin cables into the bud, orientates the spindle early in the cell cycle by binding and polarizing the microtubule-associated protein Kar9 (refs 7-9). The tail domain of Myo2 that binds Kar9 also interacts with secretory vesicles and vacuolar elements, making it a pivotal component of yeast cell polarization.     

Nicastrin modulates presenilin-mediated notch/glp-1 signal transduction and betaAPP processing

Nicastrin, a transmembrane glycoprotein, forms high molecular weight complexes with presenilin 1 and presenilin 2. Suppression of nicastrin expression in Caenorhabditis elegans embryos induces a subset of notch/glp-1 phenotypes similar to those induced by simultaneous null mutations in both presenilin homologues of C. elegans (sel-12 and hop-1). Nicastrin also binds carboxy-terminal derivatives of beta-amyloid precursor protein (betaAPP), and modulates the production of the amyloid beta-peptide (A beta) from these derivatives. Missense mutations in a conserved hydrophilic domain of nicastrin increase A beta42 and A beta40 peptide secretion. Deletions in this domain inhibit A beta production. Nicastrin and presenilins are therefore likely to be functional components of a multimeric complex necessary for the intramembranous proteolysis of proteins such as Notch/GLP-1 and betaAPP.     

Cloned pigs produced by nuclear transfer from adult somatic cells

Since the first report of live mammals produced by nuclear transfer from a cultured differentiated cell population in 1995 (ref. 1), successful development has been obtained in sheep, cattle, mice and goats using a variety of somatic cell types as nuclear donors. The methodology used for embryo reconstruction in each of these species is essentially similar: diploid donor nuclei have been transplanted into enucleated MII oocytes that are activated on, or after transfer. In sheep and goat pre-activated oocytes have also proved successful as cytoplast recipients. The reconstructed embryos are then cultured and selected embryos transferred to surrogate recipients for development to term. In pigs, nuclear transfer has been significantly less successful; a single piglet was reported after transfer of a blastomere nucleus from a four-cell embryo to an enucleated oocyte; however, no live offspring were obtained in studies using somatic cells such as diploid or mitotic fetal fibroblasts as nuclear donors. The development of embryos reconstructed by nuclear transfer is dependent upon a range of factors. Here we investigate some of these factors and report the successful production of cloned piglets from a cultured adult somatic cell population using a new nuclear transfer procedure.     

Simulating dynamical features of escape panic

One of the most disastrous forms of collective human behaviour is the kind of crowd stampede induced by panic, often leading to fatalities as people are crushed or trampled. Sometimes this behaviour is triggered in life-threatening situations such as fires in crowded buildings; at other times, stampedes can arise during the rush for seats or seemingly without cause. Although engineers are finding ways to alleviate the scale of such disasters, their frequency seems to be increasing with the number and size of mass events. But systematic studies of panic behaviour and quantitative theories capable of predicting such crowd dynamics are rare. Here we use a model of pedestrian behaviour to investigate the mechanisms of (and preconditions for) panic and jamming by uncoordinated motion in crowds. Our simulations suggest practical ways to prevent dangerous crowd pressures. Moreover, we find an optimal strategy for escape from a smoke-filled room, involving a mixture of individualistic behaviour and collective 'herding' instinct.     

The crystal structure of DNA mismatch repair protein MutS binding to a G x T mismatch

DNA mismatch repair ensures genomic integrity on DNA replication. Recognition of a DNA mismatch by a dimeric MutS protein initiates a cascade of reactions and results in repair of the newly synthesized strand; however, details of the molecular mechanism remain controversial. Here we present the crystal structure at 2.2 A of MutS from Escherichia coli bound to a G x T mismatch. The two MutS monomers have different conformations and form a heterodimer at the structural level. Only one monomer recognizes the mismatch specifically and has ADP bound. Mismatch recognition occurs by extensive minor groove interactions causing unusual base pairing and kinking of the DNA. Nonspecific major groove DNA-binding domains from both monomers embrace the DNA in a clamp-like structure. The interleaved nucleotide-binding sites are located far from the DNA. Mutations in human MutS alpha (MSH2/MSH6) that lead to hereditary predisposition for cancer, such as hereditary non-polyposis colorectal cancer, can be mapped to this crystal structure.     

Interconversion of single and double helices formed from synthetic molecular strands

Synthetic single-helical conformations are quite common, but the formation of double helices based on recognition between the two constituent strands is relatively rare. Known examples include duplex formation through base-pair-specific hydrogen bonding and stacking, as found in nucleic acids and their analogues, and polypeptides composed of amino acids with alternating L and D configurations. Some synthetic polymers and self-assembled fibres have double-helical winding induced by van der Waals interactions. A third mode of non-covalent interaction, coordination of organic ligands to metal ions, can give rise to double, triple and quadruple helices, although in this case the assembly is driven by the coordination geometry of the metal and the structure of the ligands, rather than by direct inter-strand complementarity. Here we describe a family of oligomeric molecules with bent conformations, which exhibit dynamic exchange between single and double molecular helices in solution, through spiral sliding of the synthetic oligomer strands. The bent conformations leading to the helical shape of the molecules result from intramolecular hydrogen bonding within 2'-pyridyl-2-pyridinecarboxamide units, with extensive intermolecular aromatic stacking stabilizing the double-stranded helices that form through dimerization.     

Importance of stirring in the development of an iron-fertilized phytoplankton bloom

The growth of populations is known to be influenced by dispersal, which has often been described as purely diffusive. In the open ocean, however, the tendrils and filaments of phytoplankton populations provide evidence for dispersal by stirring. Despite the apparent importance of horizontal stirring for plankton ecology, this process remains poorly characterized. Here we investigate the development of a discrete phytoplankton bloom, which was initiated by the iron fertilization of a patch of water (7 km in diameter) in the Southern Ocean. Satellite images show a striking, 150-km-long bloom near the experimental site, six weeks after the initial fertilization. We argue that the ribbon-like bloom was produced from the fertilized patch through stirring, growth and diffusion, and we derive an estimate of the stirring rate. In this case, stirring acts as an important control on bloom development, mixing phytoplankton and iron out of the patch, but also entraining silicate. This may have prevented the onset of silicate limitation, and so allowed the bloom to continue for as long as there was sufficient iron. Stirring in the ocean is likely to be variable, so blooms that are initially similar may develop very differently.     

arrow encodes an LDL-receptor-related protein essential for Wingless signalling

The Wnt family of secreted molecules functions in cell-fate determination and morphogenesis during development in both vertebrates and invertebrates (reviewed in ref. 1). Drosophila Wingless is a founding member of this family, and many components of its signal transduction cascade have been identified, including the Frizzled class of receptor. But the mechanism by which the Wingless signal is received and transduced across the membrane is not completely understood. Here we describe a gene that is necessary for all Wingless signalling events in Drosophila. We show that arrow gene function is essential in cells receiving Wingless input and that it acts upstream of Dishevelled. arrow encodes a single-pass transmembrane protein, indicating that it may be part of a receptor complex with Frizzled class proteins. Arrow is a low-density lipoprotein (LDL)-receptor-related protein (LRP), strikingly homologous to murine and human LRP5 and LRP6. Thus, our data suggests a new and conserved function for this LRP subfamily in Wingless/Wnt signal reception.