New Mouse Lines for the Analysis of Neuronal Morphology Using CreER(T)/loxP-Directed Sparse Labeling

BACKGROUND: Pharmacologic control of Cre-mediated recombination using tamoxifen-dependent activation of a Cre-estrogen receptor ligand binding domain fusion protein [CreER(T)] is widely used to modify and/or visualize cells in the mouse. METHODS AND FINDINGS: We describe here two new mouse lines, constructed by gene targeting to the Rosa26 locus to facilitate Cre-mediated cell modification. These lines should prove particularly useful in the context of sparse labeling experiments. The R26rtTACreER line provides ubiquitous expression of CreER under transcriptional control by the tetracycline reverse transactivator (rtTA); dual control by doxycycline and tamoxifen provides an extended dynamic range of Cre-mediated recombination activity. The R26IAP line provides high efficiency Cre-mediated activation of human placental alkaline phosphatase (hPLAP), complementing the widely used, but low efficiency, Z/AP line. By crossing with mouse lines that direct cell-type specific CreER expression, the R26IAP line has been used to produce atlases of labeled cholinergic and catecholaminergic neurons in the mouse brain. The R26IAP line has also been used to visualize the full morphologies of retinal dopaminergic amacrine cells, among the largest neurons in the mammalian retina. CONCLUSIONS: The two new mouse lines described here expand the repertoire of genetically engineered mice available for controlled in vivo recombination and cell labeling using the Cre-lox system

Helicobacter pylori versus the Host: Remodeling of the Bacterial Outer Membrane Is Required for Survival in the Gastric Mucosa

Modification of bacterial surface structures, such as the lipid A portion of lipopolysaccharide (LPS), is used by many pathogenic bacteria to help evade the host innate immune response. Helicobacter pylori, a gram-negative bacterium capable of chronic colonization of the human stomach, modifies its lipid A by removal of phosphate groups from the 1- and 4′-positions of the lipid A backbone. In this study, we identify the enzyme responsible for dephosphorylation of the lipid A 4′-phosphate group in H. pylori, Jhp1487 (LpxF). To ascertain the role these modifications play in the pathogenesis of H. pylori, we created mutants in lpxE (1-phosphatase), lpxF (4′-phosphatase) and a double lpxE/F mutant. Analysis of lipid A isolated from lpxE and lpxF mutants revealed lipid A species with a 1 or 4′-phosphate group, respectively while the double lpxE/F mutant revealed a bis-phosphorylated lipid A. Mutants lacking lpxE, lpxF, or lpxE/F show a 16, 360 and 1020 fold increase in sensitivity to the cationic antimicrobial peptide polymyxin B, respectively. Moreover, a similar loss of resistance is seen against a variety of CAMPs found in the human body including LL37, β-defensin 2, and P-113. Using a fluorescent derivative of polymyxin we demonstrate that, unlike wild type bacteria, polymyxin readily associates with the lpxE/F mutant. Presumably, the increase in the negative charge of H. pylori LPS allows for binding of the peptide to the bacterial surface. Interestingly, the action of LpxE and LpxF was shown to decrease recognition of Helicobacter LPS by the innate immune receptor, Toll-like Receptor 4. Furthermore, lpxE/F mutants were unable to colonize the gastric mucosa of C57BL/6J and C57BL/6J tlr4 -/- mice when compared to wild type H. pylori. Our results demonstrate that dephosphorylation of the lipid A domain of H. pylori LPS by LpxE and LpxF is key to its ability to colonize a mammalian host

Single-cell analysis tools for drug discovery and development

The genetic, functional or compositional heterogeneity of healthy and diseased tissues presents major challenges in drug discovery and development. Such heterogeneity hinders the design of accurate disease models and can confound the interpretation of biomarker levels and of patient responses to specific therapies. The complex nature of virtually all tissues has motivated the development of tools for single-cell genomic, transcriptomic and multiplex proteomic analyses. Here, we review these tools and assess their advantages and limitations. Emerging applications of single cell analysis tools in drug discovery and development, particularly in the field of oncology, are discussed

Constraints on the origin and evolution of the layered mound in Gale Crater, Mars using Mars Reconnaissance Orbiter data

Gale Crater contains a 5.2 km-high central mound of layered material that is largely sedimentary in origin and has been considered as a potential landing site for both the MER (Mars Exploration Rover) and MSL (Mars Science Laboratory) missions. We have analyzed recent data from Mars Reconnaissance Orbiter to help unravel the complex geologic history evidenced by these layered deposits and other landforms in the crater. Results from imaging data from the High Resolution Imaging Science Experiment (HiRISE) and Context Camera (CTX) confirm geomorphic evidence for fluvial activity and may indicate an early lacustrine phase. Analysis of spectral data from the CRISM (Compact Reconnaissance Imaging Spectrometer for Mars) instrument shows clay-bearing units interstratified with sulfate-bearing strata in the lower member of the layered mound, again indicative of aqueous activity. The formation age of the layered mound, derived from crater counts and superposition relationships, is similar to 3.6-3.8 Ga and straddles the Noachian-Hesperian time-stratigraphic boundary. Thus Gale provides a unique opportunity to investigate global environmental change on Mars during a period of transition from an environment that favored phyllosilicate deposition to a later one that was dominated by sulfate formation. (C) 2011 Elsevier Inc. All rights reserved.2142413432NAS

Contemporaneous deposition of phyllosilicates and sulfates: Using Australian acidic saline lake deposits to describe geochemical variability on Mars

Studies of the origin of the Martian sulfate and phyllosilicate deposits have led to the hypothesis that there was a marked, global-scale change in the Mars environment from circum-neutral pH aqueous alteration in the Noachian to an acidic evaporitic system in the late Noachian to Hesperian. However, terrestrial studies suggest that two different geochemical systems need not be invoked to explain such geochemical variation. Western Australian acidic playa lakes have large pH differences separated vertically and laterally by only a few tens of meters, demonstrating how highly variable chemistries can coexist over short distances in natural environments. We suggest diverse and variable Martian aqueous environments where the coetaneous formation of phyllosilicates and sulfates at the Australian sites are analogs for regions where phyllosilicates and sulfates coexist on Mars. In these systems, Fe and alkali earth phyllosilicates represent deep facies associated with upwelling neutral to alkaline groundwater, whereas aluminous phyllosilicates and sulfates represent near-surface evaporitic facies formed from more acidic brines. Citation: Baldridge, A. M., S. J. Hook, J. K. Crowley, G. M. Marion, J. S. Kargel, J. L. Michalski, B. J. Thomson, C. R. de Souza Filho, N. T. Bridges, and A. J. Brown (2009), Contemporaneous deposition of phyllosilicates and sulfates: Using Australian acidic saline lake deposits to describe geochemical variability on Mars, Geophys. Res. Lett., 36, L19201, doi:10.1029/2009GL040069.O TEXTO COMPLETO DESTE ARTIGO, ESTARÁ DISPONÍVEL À PARTIR DE FEVEREIRO DE 2015.3