Improving reservoir conformance using gelled polymer systems. Final report, September 25, 1992--July 31, 1996

The objectives of the research program were to (1) identify and develop polymer systems which have potential to improve reservoir conformance of fluid displacement processes, (2) determine the performance of these systems in bulk and in porous media, and (3) develop methods to predict their performance in field applications. The research focused on four types of gel systems--KUSP1 systems that contain an aqueous polysaccharide designated KUSP1, phenolic-aldehyde systems composed of resorcinol and formaldehyde, colloidal-dispersion systems composed of polyacrylamide and aluminum citrate, and a chromium-based system where polyacrylamide is crosslinked by chromium(III). Gelation behavior of the resorcinol-formaldehyde systems and the KUSP1-borate system was examined. Size distributions of aggregates that form in the polyacrylamide-aluminum colloidal-dispersion gel system were determined. Permeabilities to brine of several rock materials were significantly reduced by gel treatments using the KUSP1 polymer-ester (monoethyl phthalate) system, the KUSP1 polymer-boric acid system, and the sulfomethylated resorcinol-formaldehyde system were also shown to significantly reduce the permeability to supercritical carbon dioxide. A mathematical model was developed to simulate the behavior of a chromium redox-polyacrylamide gel system that is injected through a wellbore into a multi-layer reservoir in which crossflow between layers is allowed. The model describes gelation kinetics and filtration of pre-gel aggregates in the reservoir. Studies using the model demonstrated the effect filtration of gel aggregates has on the placement of gel systems in layered reservoirs

Classification Accuracy of the Quick Interactive Language Screener for Preschool Children with and without Developmental Language Disorder

This research examined the classification accuracy of the Quick Interactive Language Screener (QUILS) for identifying preschool-aged children (3;0 to 6;9) with developmental language disorder (DLD). We present data from two independent samples that varied in prevalence and diagnostic reference standard

Human tribbles-1 controls proliferation and chemotaxis of smooth muscle cells via MAPK signaling pathways

Migration and proliferation of smooth muscle cells are key to a number of physiological and pathological processes, including wound healing and the narrowing of the vessel wall.Previous work has shown links between inflammatory stimuli and vascular smooth muscle cell proliferation and migration through mitogen activated protein kinase (MAPK) activation, though the molecular mechanisms of this process are poorly understood. Here we report that tribbles-1, a recently described modulator of MAPK activation controls vascular smooth muscle cell proliferation and chemotaxis via the Jun Kinase pathway. Our findings demonstrate that this regulation takes place via direct interactions between tribbles-1 and MKK4/SEK1, a Jun activator kinase. The activity of this kinase is dependent on tribbles-1 levels, whilst the activation and the expression of MKK4/SEK1 is not. In addition, tribbles-1 expression is elevated in human atherosclerotic arteries compared to non-atherosclerotic controls, suggesting that this protein may pay a role in disease in vivo. In summary, the data presented here suggest an important regulatory role for trb-1 in vascular smooth muscle cell biology

Mild/moderate haemophilia A: new insights into molecular mechanisms and inhibitor development

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/72987/1/j.1365-2516.2008.01730.x.pd

A Novel Genetic Screen Implicates Elm1 in the Inactivation of the Yeast Transcription Factor SBF

BACKGROUND: Despite extensive large scale analyses of expression and protein-protein interactions (PPI) in the model organism Saccharomyces cerevisiae, over a thousand yeast genes remain uncharacterized. We have developed a novel strategy in yeast that directly combines genetics with proteomics in the same screen to assign function to proteins based on the observation of genetic perturbations of sentinel protein interactions (GePPI). As proof of principle of the GePPI screen, we applied it to identify proteins involved in the regulation of an important yeast cell cycle transcription factor, SBF that activates gene expression during G1 and S phase. METHODOLOGY/PRINCIPLE FINDINGS: The principle of GePPI is that if a protein is involved in a pathway of interest, deletion of the corresponding gene will result in perturbation of sentinel PPIs that report on the activity of the pathway. We created a fluorescent protein-fragment complementation assay (PCA) to detect the interaction between Cdc28 and Swi4, which leads to the inactivation of SBF. The PCA signal was quantified by microscopy and image analysis in deletion strains corresponding to 25 candidate genes that are periodically expressed during the cell cycle and are substrates of Cdc28. We showed that the serine-threonine kinase Elm1 plays a role in the inactivation of SBF and that phosphorylation of Elm1 by Cdc28 may be a mechanism to inactivate Elm1 upon completion of mitosis. CONCLUSIONS/SIGNIFICANCE: Our findings demonstrate that GePPI is an effective strategy to directly link proteins of known or unknown function to a specific biological pathway of interest. The ease in generating PCA assays for any protein interaction and the availability of the yeast deletion strain collection allows GePPI to be applied to any cellular network. In addition, the high degree of conservation between yeast and mammalian proteins and pathways suggest GePPI could be used to generate insight into human disease

The yeast P5 type ATPase, Spf1, regulates manganese transport into the endoplasmic reticulum

The endoplasmic reticulum (ER) is a large, multifunctional and essential organelle. Despite intense research, the function of more than a third of ER proteins remains unknown even in the well-studied model organism Saccharomyces cerevisiae. One such protein is Spf1, which is a highly conserved, ER localized, putative P-type ATPase. Deletion of SPF1 causes a wide variety of phenotypes including severe ER stress suggesting that this protein is essential for the normal function of the ER. The closest homologue of Spf1 is the vacuolar P-type ATPase Ypk9 that influences Mn2+ homeostasis. However in vitro reconstitution assays with Spf1 have not yielded insight into its transport specificity. Here we took an in vivo approach to detect the direct and indirect effects of deleting SPF1. We found a specific reduction in the luminal concentration of Mn2+ in ∆spf1 cells and an increase following it’s overexpression. In agreement with the observed loss of luminal Mn2+ we could observe concurrent reduction in many Mn2+-related process in the ER lumen. Conversely, cytosolic Mn2+-dependent processes were increased. Together, these data support a role for Spf1p in Mn2+ transport in the cell. We also demonstrate that the human sequence homologue, ATP13A1, is a functionally conserved orthologue. Since ATP13A1 is highly expressed in developing neuronal tissues and in the brain, this should help in the study of Mn2+-dependent neurological disorders

The Power of Play: A Pediatric Role in Enhancing Development in Young Children

Children need to develop a variety of skill sets to optimize their development and manage toxic stress. Research demonstrates that developmentally appropriate play with parents and peers is a singular opportunity to promote the social-emotional, cognitive, language, and self-regulation skills that build executive function and a prosocial brain. Furthermore, play supports the formation of the safe, stable, and nurturing relationships with all caregivers that children need to thrive

Laboratory Evolution of Fast-Folding Green Fluorescent Protein Using Secretory Pathway Quality Control

Green fluorescent protein (GFP) has undergone a long history of optimization to become one of the most popular proteins in all of cell biology. It is thermally and chemically robust and produces a pronounced fluorescent phenotype when expressed in cells of all types. Recently, a superfolder GFP was engineered with increased resistance to denaturation and improved folding kinetics. Here we report that unlike other well-folded variants of GFP (e.g., GFPmut2), superfolder GFP was spared from elimination when targeted for secretion via the SecYEG translocase. This prompted us to hypothesize that the folding quality control inherent to this secretory pathway could be used as a platform for engineering similar ‘superfolded’ proteins. To test this, we targeted a combinatorial library of GFPmut2 variants to the SecYEG translocase and isolated several superfolded variants that accumulated in the cytoplasm due to their enhanced folding properties. Each of these GFP variants exhibited much faster folding kinetics than the parental GFPmut2 protein and one of these, designated superfast GFP, folded at a rate that even exceeded superfolder GFP. Remarkably, these GFP variants exhibited little to no loss in specific fluorescence activity relative to GFPmut2, suggesting that the process of superfolding can be accomplished without altering the proteins' normal function. Overall, we demonstrate that laboratory evolution combined with secretory pathway quality control enables sampling of largely unexplored amino-acid sequences for the discovery of artificial, high-performance proteins with properties that are unparalleled in their naturally occurring analogues

Use of cDNA Tiling Arrays for Identifying Protein Interactions Selected by In Vitro Display Technologies

In vitro display technologies such as mRNA display are powerful screening tools for protein interaction analysis, but the final cloning and sequencing processes represent a bottleneck, resulting in many false negatives. Here we describe an application of tiling array technology to identify specifically binding proteins selected with the in vitro virus (IVV) mRNA display technology. We constructed transcription-factor tiling (TFT) arrays containing ∼1,600 open reading frame sequences of known and predicted mouse transcription-regulatory factors (334,372 oligonucleotides, 50-mer in length) to analyze cDNA fragments from mRNA-display screening for Jun-associated proteins. The use of the TFT arrays greatly increased the coverage of known Jun-interactors to 28% (from 14% with the cloning and sequencing approach), without reducing the accuracy (∼75%). This method could detect even targets with extremely low expression levels (less than a single mRNA copy per cell in whole brain tissue). This highly sensitive and reliable method should be useful for high-throughput protein interaction analysis on a genome-wide scale

New insights into protein-protein interaction data lead to increased estimates of the S. cerevisiae interactome size

<p>Abstract</p> <p>Background</p> <p>As protein interactions mediate most cellular mechanisms, protein-protein interaction networks are essential in the study of cellular processes. Consequently, several large-scale interactome mapping projects have been undertaken, and protein-protein interactions are being distilled into databases through literature curation; yet protein-protein interaction data are still far from comprehensive, even in the model organism <it>Saccharomyces cerevisiae</it>. Estimating the interactome size is important for evaluating the completeness of current datasets, in order to measure the remaining efforts that are required.</p> <p>Results</p> <p>We examined the yeast interactome from a new perspective, by taking into account how thoroughly proteins have been studied. We discovered that the set of literature-curated protein-protein interactions is qualitatively different when restricted to proteins that have received extensive attention from the scientific community. In particular, these interactions are less often supported by yeast two-hybrid, and more often by more complex experiments such as biochemical activity assays. Our analysis showed that high-throughput and literature-curated interactome datasets are more correlated than commonly assumed, but that this bias can be corrected for by focusing on well-studied proteins. We thus propose a simple and reliable method to estimate the size of an interactome, combining literature-curated data involving well-studied proteins with high-throughput data. It yields an estimate of at least 37, 600 direct physical protein-protein interactions in <it>S. cerevisiae</it>.</p> <p>Conclusions</p> <p>Our method leads to higher and more accurate estimates of the interactome size, as it accounts for interactions that are genuine yet difficult to detect with commonly-used experimental assays. This shows that we are even further from completing the yeast interactome map than previously expected.</p