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David Shintani

Photo of David Shintani


Associate Dean of Resident Instruction
Department of CABNR/NAES - Administration
University of Nevada/Mail Stop 221
1664 North Virginia Street
Reno,  Nevada   89557

 

Office: (775) 784-1095
Lab: 327-5139

Fax: 784-1419

Cell: 240-1707

Email: shintani@unr.edu
Building: Max Fleischmann Agriculture,  Office 207
Personal Web: http://www.ag.unr.edu/shintani/
 
 
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EDUCATION

B.Sc. Biochemistry, 1985 University of California, Davis
Ph.D. Biology, 1996 Michigan State University

 

ACADEMIC & RESEARCH INTEREST

Rubber Biosynthesis

The goal of this project is to determine how natural rubber is synthesized in plants. Natural rubber is required for the manufacture of thousands of products needed in daily life. Due to its superior performance properties, natural rubber is an irreplaceable material in the manufacture of many products, such as automobile and aircraft tires. Surprisingly, even with its high economic and strategic importance, the biosynthesis of rubber has been poorly characterized. Move than fifty years of biochemical experimentation has so far failed to identify the proteins required for rubber biosynthesis in plants. This is primarily due to the fact that the membrane associated rubber biosynthetic machinery is resistant to purification by classic biochemical methods. To circumvent this problem, proteomics, genomics and reverse genetic analyses will be used to functionally identify the genes/proteins required for rubber biosynthesis from two hyper-producing rubber species, guayule (Parthenium argentatum) and Russian dandelion (Taraxacum kok-saghyz). The novel approach used here represents the most rapid means of advancing our knowledge of rubber biosynthesis in plants and will lead to identification of genes/proteins that regulate the quantity and quality of natural rubber.

The gene-based resources generated from this research will be used for the improvement of current rubber producing crops and the development of alternative rubber producing domestic crops through genetic engineering and molecular breeding approaches. The development of domestic rubber producing crops will provide a number of benefits to the American public including: 1) decreased dependence on imported natural rubber, 2) the creation of a new high value commodity crops for the American farmer, 3) the generation of a hypoallergenic alternatives to Hevea derived rubber for persons with latex allergies and 4) decreased dependence on petroleum for the synthesis of synthetic polymers.

Vitamin B1 (Thiamin) Biosynthesis In Plants

Thiamin (Vitamin B1) deficiencies in humans can lead to a condition known as Beriberi that is manifested by severe neurological disorders and a general wasting phenomenon. This disease is primarily associated with poverty-stricken populations of developing countries whose diets subsist primarily of polished grain products such as polished rice or bleached wheat flour. A sustainable solution to thiamin deficiencies in humans would be to increase the nutritional content of staple food crops that endogenous populations of the world commonly consume. By genetic engineering crops for increased thiamin, it should be possible to positively impact the nutritional needs of the global population. Unfortunately, the major impediment to this effort is a current lack of knowledge pertaining to the biosynthesis of thiamin in plants. We are using a combination of biochemical, molecular, and genomic-based approaches to dissect the regulatory mechanisms controlling thiamin biosynthesis in plants. The increased biosynthetic knowledge obtained through our research will be important for the rational design of crops engineered for elevated thiamin levels for improved human and animal nutrition.

 
COURSES & TEACHING RESPONSIBILITIES

BCH 400/600 Introductory Biochemistry

 
REPRESENTATIVE PUBLICATIONS

 

Journals
Henry, O., Lopez-Gallego, F., Agger, S., Schmidt-Dannert, C., Sen, S., Shintani, D. K., Katrina, C., Distefano, M. D. 2009, A versatile photoactivatable probe designed to label the diphosphate binding site of farnesyl diphosphate utilizing enzymes.., Bioorganic & Medicinal Chemistry, 17(13), 4797-4805.  
Abstract:

Farnesyl diphosphate (FPP) is a substrate for a diverse number of enzymes found in nature. Photoactive analogues of isoprenoid diphosphates containing either benzophenone, diazotrifluoropropionate or azide groups have been useful for studying both the enzymes that synthesize FPP as well as those that employ FPP as a substrate. Here we describe the synthesis and properties of a new class of FPP analogues that links an unmodified farnesyl group to a diphosphate mimic containing a photoactive benzophenone moiety; thus, importantly, these compounds are photoactive FPP analogues that contain no modifications of the isoprenoid portion of the molecule that may interfere with substrate binding in the active site of an FPP utilizing enzyme. Two isomeric compounds containing meta- and para-substituted benzophenones were prepared. These two analogues inhibit Saccharomyces cerevisiae protein farnesyltransferase (ScPFTase) with IC(50) values of 5.8 (meta isomer) and 3.0 microM (para isomer); the more potent analogue, the para isomer, was shown to be a competitive inhibitor of ScPFTase with respect to FPP with a K(I) of 0.46 microM. Radiolabeled forms of both analogues selectively labeled the beta-subunit of ScPFTase. The para isomer was also shown to label Escherichia coli farnesyl diphosphate synthase and Drosophila melanogaster farnesyl diphosphate synthase. Finally, the para isomer was shown to be an alternative substrate for a sesquiterpene synthase from Nostoc sp. strain PCC7120, a cyanobacterial source; the compound also labeled the purified enzyme upon photolysis. Taken together, these results using a number of enzymes demonstrate that this new class of probes should be useful for a plethora of studies of FPP-utilizing enzymes.

Tunc-Ozdemir, M., Miller, G., Lua, S., Kim, J., Sodek, A., Koussevitzky, S., Misra, A. N., Mittler, R., Shintani, D. K. 2009, Thiamin confers enhanced tolerance to oxidative stress in Arabidopsis.., Plant Phsyiology, 151(1), 421-432.  
Abstract:

Thiamin and thiamin pyrophosphate (TPP) are well known for their important roles in human nutrition and enzyme catalysis. In this work, we present new evidence for an additional role of these compounds in the protection of cells against oxidative damage. Arabidopsis (Arabidopsis thaliana) plants subjected to abiotic stress conditions, such as high light, cold, osmotic, salinity, and oxidative treatments, accumulated thiamin and TPP. Moreover, the accumulation of these compounds in plants subjected to oxidative stress was accompanied by enhanced expression of transcripts encoding thiamin biosynthetic enzymes. When supplemented with exogenous thiamin, wild-type plants displayed enhanced tolerance to oxidative stress induced by paraquat. Thiamin application was also found to protect the reactive oxygen species-sensitive ascorbate peroxidase1 mutant from oxidative stress. Thiamin-induced tolerance to oxidative stress was accompanied by decreased production of reactive oxygen species in plants, as evidenced from decreased protein carbonylation and hydrogen peroxide accumulation. Because thiamin could protect the salicylic acid induction-deficient1 mutant against oxidative stress, thiamin-induced oxidative protection is likely independent of salicylic acid signaling or accumulation. Taken together, our studies suggest that thiamin and TPP function as important stress-response molecules that alleviate oxidative stress during different abiotic stress conditions.

Xie, W., McMahan, C. M., McGraw, A. J., Distefano, M. D., Cornish, K., Whalen, M. C., Shintani, D. K. 2008, Initiation of rubber biosynthesis: In vitro comparisons of benzophenone-modified diphosphate analogues in three rubber-producing species., Phytochemistry, 69(14), 2539-2545.  
Older Publications

Ajjawi I, Shintani D. Engineered plants with elevated Vitamin E: a nutriceutical success story. Trends in Biotechnology 22, 104-107.

Branen, J.K., Shintani, D. and Engeseth, N.J. Expression of Antisense Acyl Carrier Protein-4 (LMI-ACP) Reduces Lipid Content in Arabidopsis Leaf Tissue. (2003) Plant Physiology 132:748-756.

Tsegaye, Y., Shintani, D.K., DellaPenna, D. (2002) Overexpression of the enzyme p-hydroxyphenylpyruvate dioxygenase in Arabidopsis and its relation to tocopherol biosynthesis. Plant Physiology and Biochemistry. (2002) 40: 913-920.

Tang, S.; Yu, J.-K.; Slabaugh, M. B.; Shintani, D. K.; Knapp, S. J. (2002) Simple sequence repeat map of the sunflower genome.    Theoretical and Applied Genetics 105(8): 1124-1136. 

Shintani DK, DellaPenna D. (2002) The role of 2-methyl-6-phytylbenzoquinone methytransferase in determining tocopherol composition in Synechocystis sp. PCC6803. FEBS 511: 1-5

Shintani DK, DellaPenna D (1998) Elevating the vitamin E content of plants through metabolic engineering. Science 282: 2098-2100.

Shintani DK, Roesler K, Shorrosh BS, Savage L, Ohlrogge JB (1997) Antisense expression and overexpression of biotin carboxylase in tobacco leaves. Plant Physiology 114: 881-886

Wada H, Shintani DK, Ohlrogge JB (1997) Why do mitochondria synthesize fatty acids? Evidence for involvement in lipoic acid production. Proc Natl Acad Sci USA 94: 1591‑1596.

Roesler K, Shintani D, Savage L, Boddupalli S, Ohlrogge J (1997) Targeting of the Arabidopsis homomeric acetyl-Coenzyme A carboxylase to plastids of rapeseeds. Plant Physiology 113: 75 – 81

Roesler K, Savage L, Shintani DK, Shorrosh BS, Ohlrogge JB (1996) Co-purification, co-immunoprecipitation, and coordinate expression during oilseed development of acetyl-CoA carboxylase activity, biotin carboxylase protein, and biotin carboxyl carrier protein from higher plants. Planta 198:517-525.

Shintani DK and Ohlrogge JB (1995) Feedback regulation of fatty acid synthesis in tobacco cell cultures. The Plant Journal 7: 577-587

Shorrosh BS, Roesler K, Shintani DK, Van De Loo F, Savage L, Ohlrogge JB (1995) Characterization of the biotin carboxylase subunit of the plastid localized acetyl-CoA carboxylase found in dicots. Plant Physiology 108: 805-812.

Shintani DK and Ohlrogge JB (1994) Characterization of a mitochondrial acyl carrier protein isoform isolated from Arabidopsis thaliana.  Plant Physiology 104: 1221-1229.

Houck CM, Shintani DK, Knauf VC (1993) The use of Agrobacterium as a gene transfer agent for plants.  In KG Mukerji, VP Singh eds, Frontiers in Applied Microbiology, vol. 4, Rastogi Publishing Company, pp 1-25.

Thompson GA, Scherer DE, Foxall-Van Aken S, Kenny JW, Young HL, Shintani DK, Kridl JC, Knauf VC (1991) Primary structure of the precursor and mature forms of stearoyl-acyl carrier protein desaturase from safflower embryos and requirement of ferredoxin for enzyme activity. Proc Natl Acad Sci USA 88: 2578-2582.