Essential role for induced Ca2+ influx followed by [Ca2+]i rise in maintaining viability of yeast cells late in the mating pheromone response pathway. A study of [Ca2+]i in single Saccharomyces cerevisiae cells with imaging of fura-2

We established an experimental system for measuring the cytosolic-free Ca2+ concentration ([Ca2+]i) in individual Saccharomyces cerevisiae cells using fura-2 as a Ca2(+)-specific probe in conjunction with digital image processing and examined changes in [Ca2+]i in response to alpha-factor in single cells of a mating type. The addition of alpha-factor to a cells raised [Ca2+]i to several hundred nanomolar in the cells from a basal level of approximately 100 nM, simultaneous with the induction of Ca2+ influx. When the cells were incubated with alpha-factor in a Ca2(+)-deficient medium, Ca2+ influx was greatly reduced, and the rise in [Ca2+]i was not detected. This indicates that the alpha-factor-induced rise in [Ca2+]i is generated by Ca2+ influx through the plasma membrane and not by release from internal stores. In the Ca2(+)-deficient medium, a cells died specifically after they had changed into cells with one projection on the cell surface. This indicates that the rise in [Ca2+]i is essential for the late response to alpha-factor. The duration of Ca2+ requirement for maintaining viability was limited to this stage, and the earlier and later stages were not affected by Ca2+ deprivation. Mating between a and alpha mating type cells was impaired in this medium due to cell death at and before the stage of conjugation. These findings are the first evidence for an essential role for mobilized Ca2+ in the yeast life cycle.     

Proton magnetic resonance identification and discrimination of stereoisomers of C27 steroids using lanthanide shift reagents.

A simple proton magnetic resonance spectroscopic method is described for the identification and and confirmation of several stereoisomeric paris of C27 stanols as well as their keto and acetate derivatives related to cholesterol. The method, which involves the use of lanthanide shift reagents, is useful in distinguishing clearly between the isomeric pairs differing only in the geometry of a functional group and/or of the A/B-ring junction in the steroid skeleton.     

Active Sensing System with In Situ Adjustable Sensor Morphology

Background

Despite the widespread use of sensors in engineering systems like robots and automation systems, the common paradigm is to have fixed sensor morphology tailored to fulfill a specific application. On the other hand, robotic systems are expected to operate in ever more uncertain environments. In order to cope with the challenge, it is worthy of note that biological systems show the importance of suitable sensor morphology and active sensing capability to handle different kinds of sensing tasks with particular requirements.

Methodology

This paper presents a robotics active sensing system which is able to adjust its sensor morphology in situ in order to sense different physical quantities with desirable sensing characteristics. The approach taken is to use thermoplastic adhesive material, i.e. Hot Melt Adhesive (HMA). It will be shown that the thermoplastic and thermoadhesive nature of HMA enables the system to repeatedly fabricate, attach and detach mechanical structures with a variety of shape and size to the robot end effector for sensing purposes. Via active sensing capability, the robotic system utilizes the structure to physically probe an unknown target object with suitable motion and transduce the arising physical stimuli into information usable by a camera as its only built-in sensor.

Conclusions/Significance

The efficacy of the proposed system is verified based on two results. Firstly, it is confirmed that suitable sensor morphology and active sensing capability enables the system to sense different physical quantities, i.e. softness and temperature, with desirable sensing characteristics. Secondly, given tasks of discriminating two visually indistinguishable objects with respect to softness and temperature, it is confirmed that the proposed robotic system is able to autonomously accomplish them. The way the results motivate new research directions which focus on in situ adjustment of sensor morphology will also be discussed.     

Quantitative Proteomic Profiling Identifies DPYSL3 as Pancreatic Ductal Adenocarcinoma-Associated Molecule That Regulates Cell Adhesion and Migration by Stabilization of Focal Adhesion Complex

Elucidation of how pancreatic cancer cells give rise to distant metastasis is urgently needed in order to provide not only a better understanding of the underlying molecular mechanisms, but also to identify novel targets for greatly improved molecular diagnosis and therapeutic intervention. We employed combined proteomic technologies including mass spectrometry and isobaric tags for relative and absolute quantification peptide tagging to analyze protein profiles of surgically resected human pancreatic ductal adenocarcinoma tissues. We identified a protein, dihydropyrimidinase-like 3, as highly expressed in human pancreatic ductal adenocarcinoma tissues as well as pancreatic cancer cell lines. Characterization of the roles of dihydropyrimidinase-like 3 in relation to cancer cell adhesion and migration in vitro, and metastasis in vivo was performed using a series of functional analyses, including those employing multiple reaction monitoring proteomic analysis. Furthermore, dihydropyrimidinase-like 3 was found to interact with Ezrin, which has important roles in cell adhesion, motility, and invasion, while that interaction promoted stabilization of an adhesion complex consisting of Ezrin, c-Src, focal adhesion kinase, and Talin1. We also found that exogenous expression of dihydropyrimidinase-like 3 induced activating phosphorylation of Ezrin and c-Src, leading to up-regulation of the signaling pathway. Taken together, the present results indicate successful application of combined proteomic approaches to identify a novel key player, dihydropyrimidinase-like 3, in pancreatic ductal adenocarcinoma tumorigenesis, which may serve as an important biomarker and/or drug target to improve therapeutic strategies.     

Facilitated Hyperpolarization Signaling in Vascular Smooth Muscle-overexpressing TRIC-A Channels

The TRIC channel subtypes, namely TRIC-A and TRIC-B, are intracellular monovalent cation-specific channels and likely mediate counterion movements to support efficient Ca²⁺ release from the sarco/endoplasmic reticulum. Vascular smooth muscle cells (VSMCs) contain both TRIC subtypes and two Ca²⁺ release mechanisms; incidental opening of ryanodine receptors (RyRs) generates local Ca²⁺ sparks to induce hyperpolarization and relaxation, whereas agonist-induced activation of inositol trisphosphate receptors produces global Ca²⁺ transients causing contraction. Tric-a knock-out mice develop hypertension due to insufficient RyR-mediated Ca²⁺ sparks in VSMCs. Here we describe transgenic mice overexpressing TRIC-A channels under the control of a smooth muscle cell-specific promoter. The transgenic mice developed congenital hypotension. In Tric-a-overexpressing VSMCs from the transgenic mice, the resting membrane potential decreased because RyR-mediated Ca²⁺ sparks were facilitated and cell surface Ca²⁺-dependent K⁺ channels were hyperactivated. Under such hyperpolarized conditions, L-type Ca²⁺ channels were inactivated, and thus, the resting intracellular Ca²⁺ levels were reduced in Tric-a-overexpressing VSMCs. Moreover, Tric-a overexpression impaired inositol trisphosphate-sensitive stores to diminish agonist-induced Ca²⁺ signaling in VSMCs. These altered features likely reduced vascular tonus leading to the hypotensive phenotype. Our Tric-a-transgenic mice together with Tric-a knock-out mice indicate that TRIC-A channel density in VSMCs is responsible for controlling basal blood pressure at the whole-animal level.