Inhibitory GEF Phosphorylation Provides Negative Feedback in the Yeast Polarity Circuit

Cell polarity is critical for the form and function of many celltypes. During polarity establishment, cells define a cortical‘‘front’’ that behaves differently from the rest of the cortex.The front accumulates high levels of the active form ofa polarity-determining Rho-family GTPase (Cdc42, Rac, orRop) that then orients cytoskeletal elements through variouseffectors to generate the polarized morphology appropriateto the particular cell type [1, 2]. GTPase accumulation isthought to involve positive feedback, such that activeGTPase promotes further delivery and/or activation ofmore GTPase in its vicinity [3]. Recent studies suggest thatonce a front forms, the concentration of polarity factors atthe front can increase and decrease periodically, first clus-tering the factors at the cortex and then dispersing themback to the cytoplasm [4–7]. Such oscillatory behaviorimplies the presence of negative feedback in the polaritycircuit [8], but the mechanism of negative feedback wasnot known. Here we show that, in the budding yeastSaccha-romyces cerevisiae, the catalytic activity of the Cdc42-directed GEF is inhibited by Cdc42-stimulated effectorkinases, thus providing negative feedback. We furthershow that replacing the GEF with a phosphosite mutantGEF abolishes oscillations and leads to the accumulationof excess GTP-Cdc42 and other polarity factors at the front.These findings reveal a mechanism for negative feedbackand suggest that the function of negative feedback via GEFinhibition is to buffer the level of Cdc42 at the polarity site

Tracking Shallow Chemical Gradients by Actin-Driven Wandering of the Polarization Site

BACKGROUND: Many cells are remarkably proficient at tracking very shallow chemical gradients, despite considerable noise from stochastic receptor-ligand interactions. Motile cells appear to undergo a biased random walk: spatial noise in receptor activity may determine the instantaneous direction, but because noise is spatially unbiased it is filtered out by time-averaging, resulting in net movement up-gradient. How non-motile cells might filter out noise is unknown. RESULTS: Using yeast chemotropic mating as a model, we demonstrate that a polarized patch of polarity regulators “wanders” along the cortex during gradient tracking. Computational and experimental findings suggest that actin-directed membrane traffic contributes to wandering by diluting local polarity factors. The pheromone gradient appears to bias wandering via interactions between receptor-activated Gβγ and polarity regulators. Artificially blocking patch wandering impairs gradient tracking. CONCLUSIONS: We suggest that the polarity patch undergoes an intracellular biased random walk that enables noise filtering by time-averaging, allowing non-motile cells to track shallow gradients

Septin ring assembly involves cycles of GTP loading and hydrolysis by Cdc42p

At the beginning of the budding yeast cell cycle, the GTPase Cdc42p promotes the assembly of a ring of septins at the site of future bud emergence. Here, we present an analysis of cdc42 mutants that display specific defects in septin organization, which identifies an important role for GTP hydrolysis by Cdc42p in the assembly of the septin ring. The mutants show defects in basal or stimulated GTP hydrolysis, and the septin misorganization is suppressed by overexpression of a Cdc42p GTPase-activating protein (GAP). Other mutants known to affect GTP hydrolysis by Cdc42p also caused septin misorganization, as did deletion of Cdc42p GAPs. In performing its roles in actin polarization and transcriptional activation, GTP-Cdc42p is thought to function by activating and/or recruiting effectors to the site of polarization. Excess accumulation of GTP-Cdc42p due to a defect in GTP hydrolysis by the septin-specific alleles might cause unphysiological activation of effectors, interfering with septin assembly. However, the recessive and dose-sensitive genetic behavior of the septin-specific cdc42 mutants is inconsistent with the septin defect stemming from a dominant interference of this type. Instead, we suggest that assembly of the septin ring involves repeated cycles of GTP loading and GTP hydrolysis by Cdc42p. These results suggest that a single GTPase, Cdc42p, can act either as a ras-like GTP-dependent “switch” to turn on effectors or as an EF-Tu–like “assembly factor” using the GTPase cycle to assemble a macromolecular structure

Role of Polarized G Protein Signaling in Tracking Pheromone Gradients

SummaryYeast cells track gradients of pheromones to locate mating partners. Intuition suggests that uniform distribution of pheromone receptors over the cell surface would yield optimal gradient sensing. However, yeast cells display polarized receptors. The benefit of such polarization was unknown. During gradient tracking, cell growth is directed by a patch of polarity regulators that wanders around the cortex. Patch movement is sensitive to pheromone dose, with wandering reduced on the up-gradient side of the cell, resulting in net growth in that direction. Mathematical modeling suggests that active receptors and associated G proteins lag behind the polarity patch and act as an effective drag on patch movement. In vivo, the polarity patch is trailed by a G protein-rich domain, and this polarized distribution of G proteins is required to constrain patch wandering. Our findings explain why G protein polarization is beneficial and illuminate a novel mechanism for gradient tracking

Metabolite changes in blood predict the onset of tuberculosis

Immunogenetics and cellular immunology of bacterial infectious disease

Search for the Lepton-Number-Violating Decay Ξ−→pμ−μ−\Xi^- \to p \mu^- \mu^-

A sensitive search for the lepton-number-violating decay $\Xi^-\to p \mu^-\mu^-$ has been performed using a sample of $\sim10^9$ $\Xi^-$ hyperons produced in 800 GeV/$c$ $p$-Cu collisions. We obtain $\mathcal{B}(\Xi^-\to p \mu^-\mu^-)< 4.0\times 10^{-8}$ at 90% confidence, improving on the best previous limit by four orders of magnitude.Comment: 9 pages, 5 figures, to be published in Phys. Rev. Let

Observation of Parity Violation in the Omega-minus -> Lambda + K-minus Decay

The alpha decay parameter in the process Omega-minus -> Lambda + K-minus has been measured from a sample of 4.50 million unpolarized Omega-minus decays recorded by the HyperCP (E871) experiment at Fermilab and found to be [1.78 +/- 0.19(stat) +/- 0.16(syst)]{\times}10^{-2}. This is the first unambiguous evidence for a nonzero alpha decay parameter, and hence parity violation, in the Omega-minus -> Lambda + K-minus decay.Comment: 10 pages, 7 figure