Research: Biology / Pre-Professional Health Sciences

Environmental Planning and Management (Kelsey Gilcrease)

Kelsey Gilcrease's research involves pragmatic field work strategies for environmental planners in wildlife management. This involves evaluating and improving species action plans, assessing current studies, and informing wildlife planners to help improve the science that will be underpinned in the field. Limited population data are available for certain rabbits and hares in the United States, and current research involves the population status of these leporid species. Instructor Dahl has also had the opportunity to work with the planning and management of koalas and wallabies in Australia and, in the future, plans to work with some of our larger South Dakota fauna, such as mountain lions.

Extremophilic Bioprocessing (Dr. Rajesh Sani)

A continuous decrease in fossil fuel reserves warrants the urgent need for the development of alternative fuel sources. Lignocellulosic biomass has long been recognized as a potential sustainable source of biofuels such as bioethanol, biodiesel, biobutanol, and biogas. The Energy Independence and Security Act of 2007 and Biomass Multi-Year Program Plan (2009) declared the need for >36 billion gallons per year of biofuel production. To meet these demands, it is critical that a proportion of non-food biomass including lignocellulosic waste materials should be transformed into biofuels.

The literature suggests that current strategies of lignocellulose processing cannot be used to produce biofuels in a sustainable and economical way until we overcome:  high production costs of lignocellulose deconstruction enzymes; the very slow enzymatic hydrolysis kinetics of sugar-release from lignocellulose; and the low yields of sugars from lignocellulose (incomplete hydrolysis). These issues need a significant fundamental as well as applied research input and are currently being addressed by the research group at SD Mines. For example, one method to increase lignocellulose hydrolysis rates is to use enzymes and microorganisms naturally adapted to environments of extreme temperatures. High temperature processes offer faster, more effective and reliable conversion of substrates to commodity chemicals due to the increased solubility and hydrolysis of lignocelluloses as well as a lower risk of contamination.

This research group has been working on extremophiles isolated from the deepest mine in North America, the Homestake Gold Mine (7,800 feet deep, also known as Deep Underground Science and Engineering Laboratory DUSEL), a Local Compost Facility, and Hot Springs State Park in Thermopolis, WY, for lignocellulose conversion under thermophilic (≥60oC) conditions. The Homestake Mine is located in the heart of the Black Hills in South Dakota, within one-hour driving distance from SD Mines. The mine was active for 125 years until its closure in 2001. Microbial communities in the geographically isolated deep biosphere of the mine have survived under nutrient-limited, extreme environmental conditions, leading their evolutionary lineage to be distinct from the surface microorganisms. However, during active mining operations over 125 years, surface microbes and lignocellulosic substrates were introduced into the extreme environments of the mine. Interactions between introduced and existing microorganisms through horizontal genetic transfer might have resulted in alteration in the gene sequences, becoming “the novel genes”. We hypothesized that genetically distinct microbes including extremophiles with diverse, novel metabolic activities might be present in the biofilms, decaying timbers, and soils of the Homestake Mine. This hypothesis is well-supported and at present we have unique thermophiles producing lignocellulose-degrading enzymes growing in the presence of cellulose as the source of carbon and energy. (See Dr. Sani’s Home page for more details.)

We have been working on “Hydrogen Production from Lignocellulose using Extremophiles”.  Hydrogen, due to its ability to be converted into electrical energy in fuel cells, has gained much interest as a viable biofuel. We are developing an efficient and cost-effective consolidated biohydrogen production process which will include production of thermostable enzymes (cellulases and xylanases from thermophilic bacteria), hydrolysis, and fermentation of lignocellulosic materials in a single reactor at 60°C.

Human Telomere Biology and Cancer (Dr. David Gilley)

Our major research interests are in telomere biology and the causes of genomic instability in cancer. Our current research focuses on four study areas: 1) determining the extent and mechanisms of direct crosstalk via signaling between telomere maintenance and DNA repair; 2) epigenetic regulation of telomere maintenance; 3) exploring the hypothesis that telomere dysfunction is a key cause of genomic instability in human cancer; 4) identifying the basic composition and fundamental workings of mammalian telomere maintenance, telomerase activation and telomere-based DNA damage responses in human epithelial repopulating cells and how this is altered during tumorigenesis.

Our lab has developed innovative technologies to drive progress in the detection and analysis of telomere dysfunction in human breast tumor tissue and in the analysis of the telomere biology and telomere-based DNA damage responses of normal and tumor stem and progenitor cells from the human breast. We have moved these areas forward in fundamental ways by developing novel technologies and experimental approaches to study telomere dysfunction in human tumor tissues. We have recently discovered a dynamic level of telomere biological plasticity in normal human breast subpopulations isolated directly from normal human tissues. These experiments have revealed critical tumor suppressive pathways that are likely bypassed during breast tumorigenesis. The translational goals of our research are to develop genetic-based telomere dysfunction markers for early cancer detection and monitoring disease recurrence. My group has worked for several years to establish new methods and concepts in human-based telomere and cancer research, which we anticipate will drive substantial progress in these areas. Many of our studies have involved successful and sustained collaborations with researchers locally and at other institutions.

- Crosstalk between the telomere and the DNA damage response: Our work in this area was originally sparked by reports that the DNA repair protein Ku80 functioned at yeast telomeres.  Following up on these yeast studies, we discovered that mammalian DNA repair proteins (Ku and the DNA-PKcs) played protective roles at telomeres. Our work in this area has been highly cited and was, in part, responsible for the emergence of the expanding field connecting the telomere with the DNA damage response. Interestingly, within this collection of DNA damage repair proteins, with telomeric protective roles, were two important protein kinases, ataxia telangiectasia mutated protein kinase (ATM) and the DNA-dependent protein kinase catalytic subunit (DNA-PKcs). These findings led my group to study whether telomere-associated proteins were substrates for these kinases, and if so, precisely which function(s) were facilitated by these modifications.  We discovered that ATM phosphorylates the telomere-associated protein TRF2 at a specific residue upon DNA damage.  This was the first example of a DNA damage response-induced phosphorylation of a telomere-associated protein. We discovered that disruption of the phosphorylated residue completely disrupted the “fast” pathway of DNA DSB repair (dependent on DNA-PK or classic-nonhomologous end joining).  Recently we confirmed earlier results from another group that TRF2 localizes to sites of DNA damage. Recent reports indicated that other telomere associated proteins may also be phosphorylated upon DNA damage and migrate to DNA damage sites. 

- Epigentics and the telomere: Telomere length homeostasis, critical for chromosomal integrity and genome stability, is controlled by intricate systems of molecular regulatory machinery that includes epigenetic modifications. This research will examine site-specific and spatiotemporal alteration of the subtelomeric methylation of CpG islands using powerful optogenetic tools recently developed by our team to uncover the epigenetic regulatory mechanisms of telomere length maintenance and stability. The power of our approach comes from its sensitivity to explore telomere regulation with single molecule techniques in live single cells, to modify methylation marks specifically at subtelomeric regions, its modular design that promotes customization, and its applicability to a wide range of basic and applied research questions related to gene regulation, DNA replication and repair, and chromosome dynamics. 

- Telomere dysfunction in spontaneous human carcinomas: We are currently interested in the role of telomere maintenance and dysfunction in human development and tumorigenesis. Our work in this area began by developing novel methods to examine human tissue samples (i.e., telomere fusion PCR) and in forming collaborations with groups that possess defined areas of complimentary expertise. Initially, we determined the extent of telomere fusions in spontaneous human breast tumor tissues, which was the first report of this definitive marker in human solid tumorigenesis. We have recently discovered high levels of telomere fusions in prostate and ovarian tumor tissues, suggesting that telomere dysfunction may be a common gateway event in tumorigenesis.

- Telomere dynamics in human epithelial repopulating cells: We have also established the telomere biological base-line state of normal mammary epithelial subpopulations isolated directly from normal breast tissues. We found that telomeric states in human mammary epithelial subpopulation are surprisingly dynamic and change markedly with aging.  We discovered that the luminal-restricted progenitors of the normal human mammary gland acquire extremely short telomeres - sufficient to initiate a prevalent spontaneous telomere-associated DNA damage response in these cells. This “naturally” occurring and unique telomeric state of the luminal progenitors in the normal human mammary gland is associated with the induction in these cells of multiple senescence markers and an elevated telomerase activity that declines with age. These findings are of high clinical interest since mounting evidence suggests that luminal progenitors may be the “cell of origin” of many breast cancers, especially those with a poor prognosis. Current work in the area will study telomere dynamics in human epithelial repopulating cells outside the breast.

DNA Structure and Function/Neurodegenerative Disease (Dr. Richard Sinden)

- DNA structure and supercoiling: The Sinden lab studies DNA, alternative conformation of DNA, and DNA supercoiling. Alternative conformations of DNA include cruciforms, left-handed Z-DNA, intramolecular triplex DNA, unwound DNA structures, and G-quadruplex structures. In 1995 we discovered a new alternative DNA structure called slipped-strand DNA. We have shown that many of these structures exist in the chromosomes of living bacterial cells. We work to understand the biological roles and consequences of these alternative DNA conformations in mammalian cells.

- Molecular mechanisms of spontaneous and genotoxicant-induced mutation: A second area of research interest involves understanding the molecular mechanisms of mutagenesis. An exciting correlation exists between DNA sequences that form alternative structures and mutations that cause cancer and human genetic disease. That is – mutations do not really occur randomly, rather they are often templated by the DNA sequence itself. In other words, certain DNA sequences (DNA repeats) are their own worst enemy. These DNA sequences are prone to, or better perhaps, programmed for, self-directed mutation. We work to understand these molecular mechanisms of spontaneous mutagenesis that involve alternative DNA conformations. In addition, we have shown that many types of mutations occur preferentially on either the leading or lagging strand during replication. Currently, we are investigating the genetic instability of DNA sequences that form four-stranded, G-quadruplex structures. These sequences are associated with genetic instabilities associated with cancer.

- DNA repeat instability associated with human genetic neurodegenerative disease: A third area of interest involves understanding the molecular basis of certain human genetic diseases. This area of research integrates the above two focus areas: DNA structure and spontaneous mutagenesis. Currently, more than 40 human genetic neurodegenerative diseases are caused by the massive expansion of (CTG)n•(CAG)n, (CGG)n•(CCG)n, (GAA)n•(TTC)n, (CCTG)n•(CAGG)n, or (ATTCT)n•(AGAAT)n DNA repeats. All these DNA repeats form one or more alternative DNA conformations, including hairpins, slipped strand DNA, parallel DNA, triplex DNA, and unwound structures, which are likely involved in their genomic instability (i.e., expansion or deletion mutations). We have developed genetic assays for studying the deletion of DNA repeats in a model bacterial system. A goal of our laboratory is to understand the molecular basis for the expansion (and deletion) mutations and to find a therapeutic approach for reducing repeat length. We are currently investigating the role of tryptanthrins and coralyne derivatives as chemicals that may stabilize intramolecular triplex DNA and lead to increased rates of deletion of the Friedreich Ataxia  (GAA)n•(TTC)n repeats. With such an approach, one may be able to prevent or delay onset of repeat expansion diseases.