Structure-function study of maize ribosome-inactivating protein: implications for the internal inactivation region and the sole glutamate in the active site

Maize ribosome-inactivating protein is classified as a class III or an atypical RNA N-glycosidase. It is synthesized as an inactive precursor with a 25-amino acid internal inactivation region, which is removed in the active form. As the first structural example of this class of proteins, crystals of the precursor and the active form were diffracted to 2.4 and 2.5 Å, respectively. The two proteins are similar, with main chain root mean square deviation (RMSD) of 0.519. In the precursor, the inactivation region is found on the protein surface and consists of a flexible loop followed by a long α-helix. This region diminished both the interaction with ribosome and cytotoxicity, but not cellular uptake. Like bacterial ribosome-inactivating proteins, maize ribosome-inactivating protein does not have a back-up glutamate in the active site, which helps the protein to retain some activity if the catalytic glutamate is mutated. The structure reveals that the active site is too small to accommodate two glutamate residues. Our structure suggests that maize ribosome-inactivating protein may represent an intermediate product in the evolution of ribosome-inactivating proteins. © 2007 The Author(s).published_or_final_versio

The Significance of Bladder Trabeculation in the Female Lower Urinary System: An Objective Evaluation by Urodynamic Studies

This study aimed to investigate the relationship between bladder trabeculation, urinary function, and the stage of pelvic organ prolapse (POP). The medical records of 104 patients with POP who underwent cystoscopies and urodynamic studies were reviewed retrospectively. Age, incidence of detrusor instability, stage and site of POP, and the parameters of urodynamic studies of patients with and without bladder trabeculation were compared. The difference in the incidence of bladder trabeculation was estimated between patients with and without a suspected bladder outlet obstruction. There were significant differences in the patients' age, stage of POP, and maximal voiding velocity. Patients with a suspected bladder outlet obstruction had a significantly higher incidence of bladder trabeculation. In addition, patients with advanced stages of POP were also found to have a higher incidence of bladder trabeculation

TSPYL2 Is Important for G1 Checkpoint Maintenance upon DNA Damage

Nucleosome assembly proteins play important roles in chromatin remodeling, which determines gene expression, cell proliferation and terminal differentiation. Testis specific protein, Y-encoded-like 2 (TSPYL2) is a nucleosome assembly protein expressed in neuronal precursors and mature neurons. Previous studies have shown that TSPYL2 binds cyclin B and inhibits cell proliferation in cultured cells suggesting a role in cell cycle regulation. To investigate the physiological significance of TSPYL2 in the control of cell cycle, we generated mice with targeted disruption of Tspyl2. These mutant mice appear grossly normal, have normal life span and do not exhibit increased tumor incidence. To define the role of TSPYL2 in DNA repair, checkpoint arrest and apoptosis, primary embryonic fibroblasts and thymocytes from Tspyl2 deficient mice were isolated and examined under unperturbed and stressed conditions. We show that mutant fibroblasts are impaired in G1 arrest under the situation of DNA damage induced by gamma irradiation. This is mainly attributed to the defective activation of p21 transcription despite proper p53 protein accumulation, suggesting that TSPYL2 is additionally required for p21 induction. TSPYL2 serves a biological role in maintaining the G1 checkpoint under stress condition

Overview of Bayesian sequential Monte Carlo methods for group and extended object tracking

This work presents the current state-of-the-art in techniques for tracking a number of objects moving in a coordinated and interacting fashion. Groups are structured objects characterized with particular motion patterns. The group can be comprised of a small number of interacting objects (e.g. pedestrians, sport players, convoy of cars) or of hundreds or thousands of components such as crowds of people. The group object tracking is closely linked with extended object tracking but at the same time has particular features which differentiate it from extended objects. Extended objects, such as in maritime surveillance, are characterized by their kinematic states and their size or volume. Both group and extended objects give rise to a varying number of measurements and require trajectory maintenance. An emphasis is given here to sequential Monte Carlo (SMC) methods and their variants. Methods for small groups and for large groups are presented, including Markov Chain Monte Carlo (MCMC) methods, the random matrices approach and Random Finite Set Statistics methods. Efficient real-time implementations are discussed which are able to deal with the high dimensionality and provide high accuracy. Future trends and avenues are traced. © 2013 Elsevier Inc. All rights reserved

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

MCMC-Based Tracking and Identification of Leaders in Groups

We present a novel framework for identifying and tracking dominant agents in groups. Our proposed approach relies on a causality detection scheme that is capable of ranking agents with respect to their contribution in shaping the system’s collective behaviour based exclusively on the agents’ observed trajectories. Further, the reasoning paradigm is made robust to multiple emissions and clutter by employing a class of recently introduced Markov chain Monte Carlo-based group tracking methods. Examples are provided that demonstrate the strong potential of the proposed scheme in identifying actual leaders in swarms of interacting agents and moving crowds