Department of Biological Sciences
 

Research Day - Saturday, February 23, 2008

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The Department of Biological Sciences' Annual Research Day will be held on Saturday, February 23rd in the Owens Banquet Room(s) on campus. Graduate students and undergraduate research students are invited to submit posters for an all day poster session. There will also be oral presentations by grad students and a talk by a former departmental graduate student. Stay tuned for more specific information!

2008 Research Day Committee

 

Submitted Abstracts:

 

The Bacillus anthracis YaaH Protein is an N-Acetylglucosaminidase Involved in Spore Cortex Depolymerization

E.L. Lambert and D.L. Popham
Department of Biological Sciences, Virginia Tech, Blacksburg, VA

Bacillus anthracis spores, the infectious agent of anthrax, are notoriously difficult to remove from contaminated areas because they are resistant to many eradication methods. These resistance properties are partially due to multiple protective layers surrounding the spore core, one of which is the cortex. In order for B. anthracis spores to germinate and resume growth, the cortex peptidoglycan must be depolymerized. This study reports mutagenesis of yaaH, which encodes a cortex lytic enzyme, using temperature-dependent plasmid insertion mutagenesis and markerless allelic exchange. Inactivating yaaH does not affect spore viability or vegetative growth but negatively affects germination. The initial stages of germination including dipicolinic acid release do not appear to be compromised, but mutant spores exhibit a slight delay in loss of optical density when compared to wild type spores. Amino acid and hexosamine analyses indicate that mutants also retain more diaminopimelic acid and N-acetylmuramic acid than wild type spores suggesting that the cortex peptidoglycan is not being hydrolyzed as rapidly. This finding is supported by HPLC analysis of peptidoglycan structure used to identify YaaH as an N-acetylglucosaminidase. When yaaH is mutated the cortex peptidoglycan is not depolymerized into small fragments but instead is retained within the spore as large fragments. Complementation of ΔyaaH restores N-acetylglucosaminidase activity, and cortex hydrolysis becomes similar to that of wild type B. anthracis. Together these results indicate that spores germinating in the absence of the yaaH-encoded N-acetylglucosaminidase must be relying on other cortex lytic enzymes to break down the cortex peptidoglycan.

 

Death investigation: Computational analysis of the intrinsic apoptotic pathway

Tongli Zhang, Paul Brazhnik and John J. Tyson
Department of Biological Science, Virginia Polytechnic Institute & State University, Blacksburg VA 24061 USA

Programmed cell death, or apoptosis, is a mechanism by which multicellular organisms shape development and remove aberrant cells. Understanding of apoptosis is important from a theoretical biology standpoint as well as from the perspective of medical research of cancer, cardiovascular, neurodegenerative and autoimmune diseases. Apoptosis can be triggered by external signals (via death receptors) or induced by various stimuli (hypoxia, DNA damage etc.), via extrinsic and intrinsic pathways respectively. Molecular biologists have collected considerable amount of information about molecular components of these pathways and their interactions, and theoreticians have introduced several quantitative mathematical models describing possible dynamics resulting from these interactions in the extrinsic pathway and in some fragments of the intrinsic pathway. Here we propose a comprehensive computational mathematical model of the intrinsic pathway apoptosis-controlling machinery based on currently known molecular interactions. The model provides a global picture of the pathway dynamics, its interactions with upstream regulators, and modifications in specific cell lines. It captures such observed experimentally features of apoptosis as threshold, time delay, and commitment, and indicates that these features are generated by the interplay between two controlling switches. We compare our model with other published models and suggest experiments which could help to refine, distinguish and validate or disapprove the models and thus advance our conceptual understanding of apoptosis.

 

Integrated dynamics of temporal controls in the cell division cycle of Caulobacter crescentus

Shenghua Li1, Paul Brazhnik1, Bruno Sobral2 and John J. Tyson1,2; 1Department of Biological Sciences and 2Virginia Bioinformatics Institute, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, USA


Caulobacter crescentus is an important model organism for studying the regulation of cell division cycle and cellular differentiation in prokaryotes. Caulobacter undergoes asymmetric division producing two progeny cells with identical genome but different developmental programs: the "swarmer" cell is flagellated and motile, and the "stalked" cell is sessile. Only stalked cells undergo chromosome replication and cell division. Swarmer cells must shed their flagellum and grow stalks before they can enter the replication-division cycle. Based on published experimental evidence, we propose a molecular mechanism for cell cycle control in this bacterium. Our quantitative model predicts detailed temporal dynamics of regulatory gene expression during the cell cycle and differentiation process of wild-type cells (both stalked cells and swarmer cells) as well as several mutant strains. The simulation presents a unified view of temporal and spatial regulation of protein activities during the asymmetric cell division cycle of Caulobacter. It helps to interpret phenotypes of known mutants and predict novel ones. The model can serve as a starting point for investigating the regulation of cell division and differentiation in other genera of alpha-proteobacteria, such as Brucella and Rhizobium, because recent experimental data suggest that these alpha-proteobacteria share similar genetic mechanisms for cell cycle control.

 

The novel Alternaria brassicicola endoplasmic reticulum membrane protein, ATM1 is a plant pathogenicity factor

Kwang-Hyung Kim1, Robert A. Cramer Jr.2, Yangrae Cho1, and Christopher B. Lawrence1; 1 Virginia Bioinformatics Institute and Department of Biological Sciences, Virginia Polytechnic Institute and State University; 2Department of Veterinary Molecular Biology, Montana State University

The necrotrophic fungus Alternaria brassicicola is the causal agent of black spot disease of many economically important Brassica species. In this report we demonstrate that A. brassicicola ATM1, a novel fungal endoplasmic reticulum (ER) membrane protein, is necessary for normal conidiogenesis and plant pathogenesis. Based on predicted amino acid sequences and annotation of functional protein domains, ATM1 encodes a hybrid membrane protein containing a single adenylation, six putative transmembrane, and FAD and NAD(P)-binding domains, and shows high sequence similarity to proteins found in several filamentous fungi. GFP localization and gene expression analyses indicated that ATM1 is localized to the ER membrane and strongly expressed during conidiation and during initial invasive growth in planta. A. brassicicola ATM1-deficient mutants were generated with a targeted gene replacement strategy by homologous recombination. The ATM1 deletion resulted in mutants exhibiting abnormal conidiogenesis including increased conidial chain branching, less pigmentation and thinner conidial cell walls compared with the wild-type strain. Virulence assays on green cabbage and Arabidopsis plants showed dramatically reduced virulence of Δatm1 mutants. During plant infection the Δatm1 mutants elicited rapid and strong plant defense responses including callose deposition and defense gene expression. Microscopic studies revealed that attempted infection by Δatm1 mutants was arrested after formation of appressoria and penetration hyphae into the plant cuticle. In addition, the mutants caused decreased and delayed cell death during infection of plants compared with the wild-type. Interestingly, mutants were capable of partial colonization of wounded or very senescent plant tissues. Collectively, these results suggest that ATM1 is likely to be involved in the production or modification of a molecule(s) required for normal conidiation and circumvention of plant defense responses.

 

Quantitative Analysis Reveals Robust and Sensitive Control of a Quorum Sensing Circuit by Two Interlocked Feedback Loops

 Joshua W. Williams1, Xiaohui Cui1, Andre Levchenko2, and Ann M. Stevens11Dept. of Biological Sciences, Virginia Tech, Blacksburg, VA 240612Dept. of Biomedical Engineering, Johns Hopkins University, Baltimore, MD 21218

Induction of the quorum sensing response of Vibrio fischeri involves a rapid switch from a state of low to high induction over a narrow concentration range of the autoinducer (AI) 3-oxo-hexanoyl-L-homoserine lactone. In this system, LuxR is an AI-dependent positive regulator of the lux operon, and LuxI produces the AI molecule. This creates a positive feedback loop common in many bacterial species that exhibit quorum-sensing-controlled gene expression. Our mathematical model of the response indicates that the system should exhibit hysteresis. This behavior is caused by a LuxR autoregulatory feedback loop, which allows LuxR to increase its concentration in the cell during the switch to full lux activation. The buildup of LuxR provides more sensitivity to the increase in AI, and accelerates the induction process. Once full induction is achieved, LuxR buffers against spurious variations in AI levels ensuring the robustness of the response. In order to test the predictions of our model, lux gene fragments with or without the ability to synthesize AI, carrying a Gfp reporter under control of the luxI promoter, were stably inserted into the att site of the Escherichia coli MG1655 chromosome. AI was removed from fully induced cultures by a variety of dilution methods for precise determination of when the quorum sensing response switched from high to low induction. Results indicate that our mathematical predictions are correct. The quorum sensing response is capable of exhibiting hysteresis when undergoing loss of AI and LuxR autoregulation is important for hysteresis to occur.

 

The resiliency of stream ecosystem function as indicated by leaf processing following cattle exclusion

April H. Hughes1, E.F. Benfield1, J.R. Webster1, W.M. Aust2;1Department of Biological Sciences, Virginia Tech, Blacksburg, VA 24061, 2Department of Forestry, Virginia Tech, Blacksburg, VA 24061

Stream ecosystems are negatively influenced by cattle grazing in the riparian zone due to increased sedimentation and nutrient loading which lead to alterations to macroinvertebrate and microbial communities. The conservation reserve enhancement program (CREP) is a relatively new program developed by the United States Department of Agriculture-Natural Resources Conservation Service (USDA-NRCS). The goal of CREP is to encourage farmers, through the use of monetary incentives to halt agricultural production on environmentally sensitive lands. Our objectives for this study are 1) to provide an understanding of whether CREP and other cattle exclusion initiatives help to restore functional integrity to streams 2) and if they do, to estimate the time it takes for integrity to be restored. We hypothesize that leaf processing (a fundamental ecosystem level function) in streams will be influenced by cattle grazing in the riparian zone because of changes in nutrient availability and sediment abundance. We predict bacterial and fungal colonization will be most influential in leaf breakdown in areas with cattle present or recently excluded due to the presence of excessive nutrients resulting from cattle urine and feces. We further predict that leaf shredding macroinvertebrates will be most influential in leaf processing in areas where cattle have been excluded for longer periods and excessive sediment is not present. This project involves monitoring leaf processing mechanisms in several headwater streams in Floyd County, VA. Included in the study are sites with current cattle grazing, rotational grazing, and sites which have had cattle excluded for less than 1 year, 5 years, and 15 years respectively. We are also studying a forested site to represent complete and infinite cattle exclusion.

 

Seasonality of territorial aggression in equatorial rufous-collared sparrows Zonotrichia capensis

Alexandra M. Class and Ignacio T. Moore
Department of Biological Sciences, Virginia Tech, Blacksburg, VA, 24061

Our understanding of the seasonality of territorial aggression in birds is mainly based on studies of temperate species. Most temperate species are territorial primarily during their breeding season. However the majority of bird species are tropical. In tropical species territories are often occupied and defended year-round. We investigated seasonality of territorial behavior in a year-round resident rufous-collared sparrows (Zonotrichia capensis) in the humid forest of northeastern Ecuador (0°35-37'S, 77°53-54'W; elevation 2100-2250m). We did simulated territorial intrusions (STIs) on resident male Z. capensis using a decoy and song playback to test if territorial aggression differs significantly with annual stage. We tested birds during pre-breeding, breeding, feeding fledglings, molting and non-breeding periods. STI’s were divided into three ten minute periods: 1) pre- STI (decoy covered, song playback off), 2) STI (decoy uncovered, song playing back) and 3) post-STI (decoy covered, song playback off). The following aggressive behaviors were measured: time within 5 m of the intruder, closest approach to the intruder, number of songs, number of song-switches and number of flights. Using principle components analysis we then determined a single composite aggression score from the variables measured. We found significant differences in territorial aggression during different stages of the annual cycle. Males were significantly more aggressive during reproductive periods and least aggressive while feeding fledglings and molting. Year-round resident equatorial Z. capensis are seasonally aggressive similar to temperate congeners despite relative population asynchrony.

 

 

Last Update February 7, 2008