SUMMARYPeroxidasins form a highly conserved family of extracellular peroxidases of unknown cellular function. We identified the C. elegans peroxidasin PXN-2 in screens for mutants defective in embryonic morphogenesis. We find that PXN-2 is essential for specific stages of embryonic morphogenesis and muscle-epidermal attachment, and is also required postembryonically for basement membrane integrity. The peroxidase catalytic activity of PXN-2 is necessary for these developmental roles. pxn-2 mutants display aberrant ultrastructure of the extracellular matrix, suggesting a role in basement membrane consolidation. PXN-2 affects specific axon guidance choice points in the developing nervous system but is dispensable for maintenance of process positions. In adults, loss of pxn-2 function promotes regrowth of axons after injury, providing the first evidence that C. elegans extracellular matrix can play an inhibitory role in axon regeneration. Loss of function in the closely related C. elegans peroxidasin pxn-1 does not cause overt developmental defects. Unexpectedly, pxn-2 mutant phenotypes are suppressed by loss of function in pxn-1 and exacerbated by overexpression of wild-type pxn-1, indicating that PXN-1 and PXN-2 have antagonistic functions. These results demonstrate that peroxidasins play crucial roles in development and reveal a new role for peroxidasins as extracellular inhibitors of axonal regeneration.

A quantitative understanding of tissue morphogenesis requires description of the movements of individual cells in space and over time. In transparent embryos, such as C. elegans, fluorescently labeled nuclei can be imaged in three-dimensional time-lapse (4D) movies and automatically tracked through early cleavage divisions up to ~350 nuclei. A similar analysis of later stages of C. elegans development has been challenging owing to the increased error rates of automated tracking of large numbers of densely packed nuclei. We present Nucleitracker4D, a freely available software solution for tracking nuclei in complex embryos that integrates automated tracking of nuclei in local searches with manual curation. Using these methods, we have been able to track >99% of all nuclei generated in the C. elegans embryo. Our analysis reveals that ventral enclosure of the epidermis is accompanied by complex coordinated migration of the neuronal substrate. We can efficiently track large numbers of migrating nuclei in 4D movies of zebrafish cardiac morphogenesis, suggesting that this approach is generally useful in situations in which the number, packing or dynamics of nuclei present challenges for automated tracking.

During vulval development in

We have used Car-Parrinello constrained molecular dynamics to study the coupling of two molecules of methanol in the zeolite chabazite to form ethanol and water. We have chosen to study this reaction because it represents the formation of the first C-C bond, which is thought to be the rate limiting step for the MTO and MTG processes. We have elucidated a new mechanism for this reaction that does not require the prior formation of surface methoxy groups or dimethyl ether intermediates. The mechanism involves stable intermediates of methane and protonated formaldehyde. We have also calculated an upper bound of the free energy barrier for the overall reaction, and found that it compares favourably with the rough experimental measurements available. Finally, we consider what are the natural reaction coordinates for the methanol-methanol coupling process.

Eph receptors and their ephrin ligands are key conserved regulators of axon guidance and can function in a variety of signaling modes. Here we analyze the genetic and cellular requirements for Eph signaling in a Caenorhabditis elegans axon guidance choice point, the ventral guidance of axons in the amphid commissure. The C. elegans Eph receptor EFN-1 has both kinase-dependent and kinase-independent roles in amphid ventral guidance. Of the four C. elegans ephrins, we find that only EFN-1 has a major role in amphid axon ventral guidance, and signals in both a receptor kinase-dependent and kinase-independent manner. Analysis of EFN-1 and EFN-1 expression and tissue-specific requirements is consistent with a model in which VAB-1 acts in amphid neurons, interacting with EFN-1 expressed on surrounding cells. Unexpectedly, left-hand neurons are more strongly affected than right-hand neurons by loss of Eph signaling, indicating a previously undetected left-right asymmetry in the requirement for Eph signaling. By screening candidate genes involved in Eph signaling, we find that the Eph kinase-independent pathway involves the ABL-1 nonreceptor tyrosine kinase and possibly the phosphatidylinositol 3-kinase pathway. Overexpression of ABL-1 is sufficient to rescue EFN-1 ventral guidance defects cell autonomously. Our results reveal new aspects of Eph signaling in a single axon guidance decision in vivo.E PHRINS and their cell surface receptors, the Eph receptor tyrosine kinases (EphR), play critical roles in many axon guidance processes, including midline guidance and growth cone collapse (Drescher et al. 1995;Cowan et al. 2000). In contrast to long-range guidance cues, Eph signaling involves short-range interactions between a transmembrane receptor and transmembrane (ephrin-B) or GPI-linked (ephrin-A) ligands. Eph signaling is complex and multifunctional, capable of mediating both repulsion and attraction depending on ephrin concentration even in the same neurons (Hansen et al. 2004). Many of the signaling pathways downstream of Eph receptors and ephrins regulate cell movement or cell adhesion (Kullander and Klein 2002;Pasquale 2005).Because Eph receptors and ephrins are cell surface molecules, they can operate in a variety of signaling modes (Kullander and Klein 2002;Egea and Klein 2007;Pasquale 2008). Eph receptors can generate kinase-dependent "forward" signals, in which ligand binding triggers receptor dimerization, activating the intrinsic kinase activity of the receptor, and initiating responses in the receptor-expressing cell. Kinase-dependent forward Eph signaling contributes to many processes including retinotopic mapping (Hindges et al. 2002), axonal midline avoidance after crossing (Yokoyama et al. 2001), neural crest cell migration (Smith et al. 1997), and migration of neural progenitors (Catchpole and Henkemeyer 2011). This regulation of diverse developmental processes occurs in part via kinase-dependent interactions with downstream effectors including Src-family kinases (Zisch et al. 1998;Knoll and Dr...