Stephanie Brady

Poster presented at the annual Nebraska Physiological Society meeting on September 8, 2007 and won second place.

Abstract:
Lysophosphatidic acid (LPA) is a lipid mediator that is increasingly implicated in various diseases involving airway wall remodeling and fibrosis. Lung fibroblast-mediated contraction of collagen gels is a widely used assay to assess mechanisms by which these cells may contribute to remodeling and fibrosis. Previous studies from the Toews and Rennard labs [J. Lab. Clin. Med. 139:20, 2002] showed that LPA stimulated gel contraction, but the LPA receptor subtypes mediating this response were not addressed. The current studies were designed to use pharmacological probes to identify the LPA receptor subtypes involved in gel contraction and to assess the importance of LPA in mediating stimulation of gel contraction by serum. HFL-1 human fetal lung fibroblasts cast into collagen gels were exposed to LPA, serum, and/or various LPA receptor-selective drugs, and the rate of gel contraction was monitored over a 6-hr time course. LPA stimulated gel contraction similarly to serum. Stimulation by LPA was blocked by the LPA1+LPA3 blocker Ki16425, implicating LPA1 and/or LPA3. In agreement with these data, gel contraction was stimulated by the LPA1-preferring agonist NAEPA and by the LPA3- preferring agonist OMPT, both of which were blocked by Ki16425. The LPA2-preferring agonist FAP12 stimulated contraction only weakly, and its effects were also blocked by Ki16425.

Stimulation of gel contraction by serum was also completely blocked by Ki16425, implicating LPA acting through LPA1 and/or LPA3 as the critical serum factor. Activation of either LPA1 or LPA3 appears to be both necessary and sufficient for LPA stimulation of fibroblast-mediated collagen gel contraction. LPA, which is present in serum at 10-20 μM, appears to be the primary gel contraction factor in serum, and LPA1 and/or LPA3 receptors mediate the gel contraction in response to serum. Drugs targeting LPA1 and LPA3 receptors could have beneficial effects to reduce or relieve airway wall remodeling. (Supported by Nebraska INBRE).

Adam Cornish

Presentation given at annual NE-INBRE meeting in Grand Island, Nebraska, August, 2007.
Entitled: “Reducing Bottlenecks in Proteomic Data Analysis ”.

Laura Heuermann

Presentation given at annual NE-INBRE meeting in Grand Island, Nebraska, August, 2007.
Entitled: “Gene Mapping NMF 193”.

Jonathan DeMuth

CYTOPLASMIC N-ACETYLGALACTOSAMINE MODIFICATION OF CYTOKERATINS AND THE IDENTIFICATION OF CANDIDATE GLYCOSYLTRANSFERASES

Previous studies by Goeltz et al (J Cell Sci. 110:1585-96) demonstrated the post-translational modification of mammalian cytokeratins (CK) by terminal N-acetylgalactosamine(GalNAc)-bearing structures, as detected by the binding of VVA, a GalNAc-binding plant lectin. The present study has extended this observation to cytokeratins from Lec8 Chinese Hamster Ovary cells, which are deficient in cell-surface glycosylation due to a defect in the delivery of UDP-N-GalNAc to the Golgi. These observations support the hypothesis that a novel intracellular glycosylation pathway exists in cells.

Deletion constructs of human cytokeratins 8 and 18 have been created, transfected into human Panc-1 cells, purified, and tested for VVA reactivity. The N-terminal halves of CK8 and CK18 bind VVA, as does the C-terminal half of CK18, indicating the presence of multiple sites of carbohydrate modification in CK18. The C-terminal half of CK8 does not bind VVA, indicating that this region lacks terminal GalNAc residues.

Screening of the Human Genome Database at the National Center for Bioinformatics Information for proteins with homology to known GalNAc-transferases, but that do not contain a signal sequence for secretion (as expected for a putative cytoplasmic glycosyltransferase), has resulted in the identification of 29 candidate genes for a cytoplasmic glycosylation pathway. This research is supported by NIH grant P20 RR016469 from the INBRE Program of the National Center for Research Resources.

Daniel Recek

Presentation given at Nebraska Academy of Sciences Annual Meeting in Lincoln, Nebraska, April 2007.
Entitled: “Web-Based Alignment Matrix Creation Tool”.

Abstract:
Sequence alignment is a research tool of fundamental importance for biologists and bioinformaticians alike. It is important that bioinformaticians understand not only the motivation behind sequence alignment, but also how it works. Alignment matrices must be computed in order to find the best possible sequence alignment in a fast and efficient manner. However, it is often difficult to create alignment matrices by hand, especially if the sequences become quite large. Therefore, I have created a web-based alignment matrix creation tool. My tool requires the user input two sequences of undetermined length, match score, mismatch penalty, and gap penalty. My program then computes the alignment on the fly and outputs the resulting matrix, including the traceback and optimal alignment, in a graphical format which is output to a PDF file. The matrix allows computation of every possible alignment between the two sequences and with the traceback, the user can find exactly how any alignment score was calculated. This program is hosted on the UNO Bioinformatics website and the back-end of the program is written in Perl and is hosted on the UNO Bioinformatics Cluster. My alignment matrix tool has been used and evaluated by several students and bioniformatics professors and they have found it useful.

Poster presented at annual NE-INBRE meeting in Grand Island, Nebraska, August, 2007.
Entitled: “Web-Based Alignment Matrix Creation Tool”.

Bonnie Errett

MUTAGENESIS OF A PUTATIVE LONG RANGE INTERACTION IN THE 5’ NTR OF COXSACKIEVIRUS B3 RNA GENOME

The overall goal of the lab is to understand the role of RNA structure in the virulence phenotype of Coxsackievirus B3. The primary focus is on the 5’ non-translated region (5’NTR) of the viral genome. Previous work in the laboratory has identified a potential long range interaction involving positions 113-118 and 561-566. My project is to further investigate this potential interaction. I am performing site directed mutagenesis to change nucleotides 113 through 118 so that they will no longer have the potential to base pair with 561-566. Once mutated, chemical modification analysis will be used to investigate the structural characteristics of the altered 5’ NTR RNA.

Corey Lawson

Presentation given at annual INBRE meeting in Grand Island, Nebraska, during the summer of 2005
Entitled: “Development of small animal model for HIV-1-induced alveolitis: Implications for pathogenesis and novel treatment approaches”

Abstract:
HIV-infected patients may present with respiratory complications related to alterations in pulmonary immune regulation. HIV-1 can cause an interstitial lung disease associated with a marked infiltration of T cells in the lung interstitium and alveolar spaces, lymphocytic alveolitis. We tested the idea that activation of peroxisome proliferator-activated receptor gamma (PPAR- g , regulating pleiotropic anti-inflammatory pathway) may affect inflammatory cells in the lung and attenuate tissue injury .

Preliminary results indicate that (1) CD8+ lymphocytes were predominant cell type infiltrating lung parenchyma; (2) increase in lymphocytic infiltrate paralleled disruption of endothelial barrier in the lung; (3) PPAR-g agonists prevented endothelial cell injury and possibly decreased levels of lymphocyte infiltration; (4) PPAR-g agonists diminished viremia levels. In toto , our data indicate that lymphocytic alveolitis can be reproduced in hu-PBL-NOD/SCID model for studies of pathogenesis and testing of new therapeutic paradigms.

Kate Dempsey

Presentation given at annual INBRE meeting in Grand Island, Nebraska, during the summer of 2005.
Entitled: “The Search for Virus-Like Copia and Other Currents Interests in Drosophilia Melnanogaster”.

Nicholas Palermo

Poster presented at the World Association of Theoretical and Computational Chemists (WATOC) meeting in Capetown, South Africa during January 16-21, 2005.
Entitled: “Weakly Polar Interactions in Model Helical Peptides”.

Presentation given at annual BRIN meeting in Grand Island, NE, during the summer of 2004
Entitled: “Theoretical Study of Aromatic-amide Interactions in Alpha Helices”.

Sarfraz Chandio

Presentation given at annual BRIN meeting in Grand Island, NE, during during the summer of 2004
Entitled: “Rapid Molecular Identification of Medically Important Fungi using RFLP Database”.