Role of the Spatzle Pro-domain in the generation of an active toll receptor ligand

The cytokine Sp?tzle is the ligand for Drosophila Toll, the prototype of an important family of membrane receptors that function in embryonic patterning and innate immunity. A dimeric precursor of Sp?tzle is processed by an endoprotease to produce a form (C-106) that cross-links Toll receptor ectodomains and establishes signaling. Here we show that before processing the pro-domain of Sp?tzle is required for correct biosynthesis and secretion. We mapped two loss-of-function mutations of Sp?tzle to a discrete site in the pro-domain and showed that the phenotype arises because of a defect in biosynthesis rather than signaling. We also report that the pro-domain and C-106 remain associated after cleavage and that this processed complex signals with the same characteristics as the C-terminal fragment. These results suggest that before activation the determinants on C-106 that bind specifically to Toll are sequestered by the pro-domain and that proteolytic processing causes conformational rearrangements that expose these determinants and enables binding to Toll. Furthermore, we show that the pro-domain is released when the Toll extracellular domain binds to the complex, a finding that has implications for the generation of a signaling-competent Toll dimer.     

Dissociation of conditioned taste avoidance from conditioned disgust reactions induced by wheel running in rats

It is well established that wheel running in rats produces conditioned taste avoidance; that is, rats that run in wheels after consuming a novel-tasting solution later consume less of that solution than rats that do not run. In experiment 1, we found that wheel running also produces conditioned disgust reactions, indicated by gapes elicited by both the taste and context that were experienced before running. Experiment 2 showed that the conditioned disgust reactions were likely not due to running itself but to a by-product of running, the rocking of the wheel that occurs when the running stops. When rocking was reduced, the disgust reactions were also reduced, but consumption of the taste solution was not changed, showing dissociation of conditioned taste avoidance and disgust. These findings indicate that the taste avoidance induced by wheel running itself is more like the taste avoidance produced by rewarding drugs than that produced by nausea-inducing drugs.     

Cofilin-mediated F-actin severing is regulated by the Rap GTPase and controls the cytoskeletal dynamics that drive lymphocyte spreading and BCR microcluster formation

When lymphocytes encounter APCs bearing cognate Ag, they spread across the surface of the APC to scan for additional Ags. This is followed by membrane contraction and the formation of Ag receptor microclusters that initiate the signaling reactions that lead to lymphocyte activation. Breakdown of the submembrane cytoskeleton is likely to be required for the cytoskeleton reorganization that drives cell spreading and for removing physical barriers that limit Ag receptor mobility. In this report, we show that Ag receptor signaling via the Rap GTPases promotes the dephosphorylation and activation of the actin-severing protein cofilin and that this results in increased severing of cellular actin filaments. Moreover, we show that this cofilin-mediated actin severing is critical for the changes in actin dynamics that drive B and T cell spreading, for the formation of BCR microclusters, and for the increased mobility of BCR microclusters within the plasma membrane after BCR engagement. Finally, using a model APC, we show that activation of this Rap-cofilin signaling module controls the amount of Ag that is gathered into BCR microclusters and that this is directly related to the magnitude of the resulting BCR signaling that is initiated during B cell-APC interactions. Thus, Rap-dependent activation of cofilin is critical for the early cytoskeletal changes and BCR reorganization that are involved in APC-dependent lymphocyte activation.     

Determinants of Rad21 localization at the centrosome in human cells

Cohesin proteins help maintain the physical associations between sister chromatids that arise in S-phase and are removed in anaphase. Recent studies found that cohesins also localize to the centrosomes, the organelles that organize the mitotic bipolar spindle. We find that the cohesin protein Rad21 localizes to centrosomes in a manner that is dependent upon known regulators of sister chromatid cohesion as well as regulators of centrosome function. These data suggest that Rad21 functions at the centrosome and that the regulators of Rad21 coordinate the centrosome and chromosomal functions of cohesin.     

Functional specialization of ribosomes?

Ribosomes are highly conserved macromolecular machines that are responsible for protein synthesis in all living organisms. Work published in the past year has shown that changes to the ribosome core can affect the mechanism of translation initiation that is favored in the cell, which potentially leads to specific changes in the relative efficiencies with which different proteins are made. Here, I examine recent data from expression and proteomic studies that suggest that cells make slightly different ribosomes under different growth conditions, and discuss genetic evidence that such differences are functional. In particular, I argue that eukaryotic cells probably produce ribosomes that lack one or more core ribosomal proteins (RPs) under some conditions, and that core RPs contribute differentially to translation of distinct subpopulations of mRNAs.     

Progress toward a general species concept

New insights in the speciation process and the nature of "species" that accumulated in the past decade demand adjustments of the species concept. The standing of some of the most broadly accepted or most innovative species concepts in the light of the growing evidence that reproductive barriers are semipermeable to gene flow, that species can differentiate despite ongoing interbreeding, that a single species can originate polyphyletically by parallel evolution, and that uniparental organisms are organised in units that resemble species of biparental organisms is discussed. As a synthesis of ideas in existing concepts and the new insights, a generalization of the genic concept is proposed that defines species as groups of individuals that are reciprocally characterized by features that would have negative fitness effects in other groups and that cannot be regularly exchanged between groups upon contact. The benefits of this differential fitness species concept are that it classifies groups that keep differentiated and keep on differentiating despite interbreeding as species, that it is not restricted to specific mutations or mechanisms causing speciation, and that it can be applied to the whole spectrum of organisms from uni- to biparentals.     

Optimal Information Transfer in the Cortex through Synchronization

In recent experimental work it has been shown that neuronal interactions are modulated by neuronal synchronization and that this modulation depends on phase shifts in neuronal oscillations. This result suggests that connections in a network can be shaped through synchronization. Here, we test and expand this hypothesis using a model network. We use transfer entropy, an information theoretical measure, to quantify the exchanged information. We show that transferred information depends on the phase relation of the signal, that the amount of exchanged information increases as a function of oscillations in the signal and that the speed of the information transfer increases as a function of synchronization. This implies that synchronization makes information transport more efficient. In summary, our results reinforce the hypothesis that synchronization modulates neuronal interactions and provide further evidence that gamma band synchronization has behavioral relevance.     

Endosomal microdomains: Formation and function

It is widely recognized that after endocytosis, internalized cargo is delivered to endosomes that act as sorting stations. The limiting membrane of endosomes contain specialized subregions, or microdomains, that represent distinct functions of the endosome, including regions competing for cargo capture leading to degradation or recycling. Great progress has been made in defining the endosomal protein coats that sort cargo in these domains, including Retromer that recycles transmembrane cargo, and ESCRT (endosomal sorting complex required for transport) that degrades transmembrane cargo. In this review, we discuss recent work that is beginning to unravel how such coat complexes contribute to the creation and maintenance of endosomal microdomains. We highlight data that indicates that adjacent microdomains do not act independently but rather interact to cross-regulate. We posit that these interactions provide an agile means for the cell to adjust sorting in response to extracellular signals and intracellular metabolic cues.     

FtsZ ring: the eubacterial division apparatus conserved in archaebacteria

FtsZ is a tubulin-like protein that is essential for cell division in eubacteria. It functions by forming a ring at the division site that directs septation. The archaebacteria constitute a kingdom of life separate from eubacteria and eukaryotes. Like eubacteria, archaebacteria are prokaryotes, although they are phylogenetically closer to eukaryotes. Here it is shown that archaebacteria also possess FtsZ and that it is biochemically similar to eubacterial FtsZs. Significantly, FtsZ from the archaebacterium Haloferax volcanii is a GTPase that is localized to a ring that coincides with the division constriction. These results indicate that the FtsZ ring was part of the division apparatus of a common prokaryotic ancestor that was retained by both eubacteria and archaebacteria.     

Primary amphipathic cell-penetrating peptides: structural requirements and interactions with model membranes

To identify rules for the design of efficient cell-penetrating peptides that deliver therapeutic agents into subcellular compartments, we compared the properties of two closely related primary amphipathic peptides that mainly differ by their conformational state. On the basis of a peptide Pbeta that is nonstructured in water and that promotes efficient cellular uptake of nucleic acids through noncovalent association, we have designed a peptide [Palpha] that is predicted to adopt a helical conformation. We show that [Pbeta] undergoes a lipid-induced conformational transition into a sheet structure, while [Palpha] remains helical. Penetration experiments show that both peptides can spontaneously insert into phospholipid membranes. Analysis of compression isotherms indicates that both peptides interact with phospholipids in the liquid expanded and liquid condensed states. AFM observations reveal that the peptides strongly disrupt the lipid organization of the monolayers and that the conformational state can influence the uptake by model membranes.     

Feature-based attention: it is all bottom-up priming

Feature-based attention (FBA) enhances the representation of image characteristics throughout the visual field, a mechanism that is particularly useful when searching for a specific stimulus feature. Even though most theories of visual search implicitly or explicitly assume that FBA is under top-down control, we argue that the role of top-down processing in FBA may be limited. Our review of the literature indicates that all behavioural and neuro-imaging studies investigating FBA suffer from the shortcoming that they cannot rule out an effect of priming. The mere attending to a feature enhances the mandatory processing of that feature across the visual field, an effect that is likely to occur in an automatic, bottom-up way. Studies that have investigated the feasibility of FBA by means of cueing paradigms suggest that the role of top-down processing in FBA is limited (e.g. prepare for red). Instead, the actual processing of the stimulus is needed to cause the mandatory tuning of responses throughout the visual field. We conclude that it is likely that all FBA effects reported previously are the result of bottom-up priming.     

The anarchist's guide to ecological theory. Or, we don't need no stinkin' laws

Several ecologists have recently suggested that ecology has several laws. This conclusion contrasts with the views of some philosophers of science, who have suggested that biology cannot have laws. I argue that the debate has been confused because two very different types of law can be recognised: correlative and causal laws. Once we recognise that there is a difference, the argument against causal laws becomes stronger, and instead I suggest that ecologists should recognise that they can and do produce generalisations that are used to build models – nomological machines – that describe the ecological systems they are studying.     

Stem cells' centrosomes: How can organelles identified 130 years ago contribute to the future of regenerative medicine?

At the core of regenerative medicine lies the expectation of repair or replacement of damaged tissues or whole organs. Donor scarcity and transplant rejection are major obstacles, and exactly the obstacles that stem cell based therapy promises to overcome. These therapies demand a comprehensive understanding of the asymmetric division of stem cells, i.e. their ability to produce cells with identical potency or differentiated cells. It is believed that with better understanding, researchers will be able to direct stem cell differentiation. Here, we describe extraordinary advances in manipulating stem cell fate that show that we need to focus on the centrosome and the centrosome-derived primary cilium. This belief comes from the fact that this organelle is the vehicle that coordinates the asymmetric division of stem cells. This is supported by studies that report the significant role of the centrosome/cilium in orchestrating signaling pathways that dictate stem cell fate. We anticipate that there is sufficient evidence to place this organelle at the center of efforts that will shape the future of regenerative medicine.     

Sex therapy: considerations in the selection of patients.

Sex therapy has proven helpful for many patients with sexual dysfunctions. In judging the appropriateness of this new treatment procedure the clinician should determine the following: that a sexual dysfunction exists, that a stable couple relationship exists, that there are no major marital, psychiatric or physical problems that are etiologically important or that would interfere with treatment, and that both partners are willing to engage in a treatment program.     

Domain structure of secretin PulD revealed by limited proteolysis and electron microscopy

Secretins, a superfamily of multimeric outer membrane proteins, mediate the transport of large macromolecules across the outer membrane of Gram-negative bacteria. Limited proteolysis of secretin PulD from the Klebsiella oxytoca pullulanase secretion pathway showed that it consists of an N-terminal domain and a protease-resistant C-terminal domain that remains multimeric after proteolysis. The stable C-terminal domain starts just before the region in PulD that is highly conserved in the secretin superfamily and apparently lacks the region at the C-terminal end to which the secretin-specific pilot protein PulS binds. Electron microscopy showed that the stable fragment produced by proteolysis is composed of two stacked rings that encircle a central channel and that it lacks the peripheral radial spokes that are seen in the native complex. Moreover, the electron microscopic images suggest that the N-terminal domain folds back into the large cavity of the channel that is formed by the C-terminal domain of the native complex, thereby occluding the channel, consistent with previous electrophysiological studies showing that the channel is normally closed.     

Biosynthesis of cryptic lipopolysaccharide glycoforms in Haemophilus influenzae involves a mechanism similar to that required for O-antigen synthesis

It is generally thought that mucosal bacterial pathogens of the genera Haemophilus, Neisseria, and Moraxella elaborate lipopolysaccharide (LPS) that is fundamentally different from that of enteric organisms that express O-specific polysaccharide side chains. Haemophilus influenzae elaborates short-chain LPS that has a role in the pathogenesis of H. influenzae infections. We show that the synthesis of LPS in this organism can no longer be as clearly distinguished from that in other gram-negative bacteria that express an O antigen. We provide evidence that a region of the H. influenzae genome, the hmg locus, is involved in the synthesis of glycoforms in which tetrasaccharide units are added en bloc, not stepwise, to the normal core glycoforms, similar to the biosynthesis of an O-antigen.     

The dynein heavy chain: structure, mechanics and evolution

The translocation of dynein along microtubules is the basis for a variety of essential cellular movements. Despite a general domain organization that is found in all the cytoskeletal motors, there are structural features of dynein that set it apart from the other motors. These include a track-binding site that is located at the tip of a long projection, and six nucleotide-binding modules that together form the globular head of dynein. These unique features suggest that dynein produces movement by a mechanism that is different from that used by the other motors.     

Mature glycosylation and trafficking of nicastrin modulate its binding to presenilins

Nicastrin is an integral component of the high molecular weight presenilin complexes that control proteolytic processing of the amyloid precursor protein and Notch. We report here that nicastrin is most probably a type 1 transmembrane glycoprotein that is expressed at moderate levels in the brain and in cultured neurons. Immunofluorescence studies demonstrate that nicastrin is localized in the endoplasmic reticulum, Golgi, and a discrete population of vesicles. Glycosidase analyses reveal that endogenous nicastrin undergoes a conventional, trafficking-dependent maturation process. However, when highly expressed in transfected cells, there is a disproportionate accumulation of the endo-beta-N-acetylglucosaminidase H-sensitive, immature form, with no significant increase in the levels of the fully mature species. Immunoprecipitation revealed that presenilin-1 interacts preferentially with mature nicastrin, suggesting that correct trafficking and co-localization of the presenilin complex components are essential for activity. These findings demonstrate that trafficking and post-translational modifications of nicastrin are tightly regulated processes that accompany the assembly of the active presenilin complexes that execute gamma-secretase cleavage. These results also underscore the caveat that simple overexpression of nicastrin in transfected cells may result in the accumulation of large amounts of the immature protein, which is apparently unable to assemble into the active complexes capable of processing amyloid precursor protein and Notch.     

Breaking the ties: epistemic significance, bacilli, and underdetermination

One premise of the underdetermination argument is that entailment of evidence is the only epistemic constraint on theory-choice. I argue that methodological rules can be epistemically significant, both with respect to observables and unobservables. Using an example from the history of medicine -- Koch's 1882 discovery of tuberculosis bacteria -- I argue that even anti-realists ought to accept that these rules can break the tie between theories that are allegedly underdetermined. I then distinguish two types of underdetermination and argue that anti-realists, in order to maintain the underdetermination argument, need to do more than show that theories are empirically equivalent: they need to show that a certain kind of underdetermination obtains.