Dr. Illimar Altosaar

Professor

Degrees

B.Sc. (McGill), Ph.D. (U.B.C.),
NATO Fellow (Imperial College of Science, Technology and Medicine, London, UK)

Contact info:

RGN, Rm. 4232
Phone: 562-5800 x 6374
Email: altosaar@uottawa.ca

Research Interests:

Innate immunity of mammary and gastrointestinal epithelia

Nutritional and immunological modulation of neonatal gut health intrigues us. We endeavour to shed more light on the ‘black box’ between breast milk and baby’s first bacterial invaders. Several basic and translational projects focus on how mother’s milk and/or infant formula and enteric microflora interact with gastric associated lymphoid tissue to impart gut health. Molecular dissection of the immune signals may help to stimulate intestinal adaptation, maintain function and prevent disease in premature infants, such as necrotizing enterocolitis (NEC). The group has established a unique model of collaboration among neonatal intensive care, pediatric gastroenterology and innate immunity. Fate and function of breast milk proteins with immuno-modulatory and anti-microbial properties are monitored both in the newborn gastrointestinal tract and mammary gland: passage, uptake, localization and distribution of breast milk proteins in newborn animal GI tracts; and, immune responses of these breast milk proteins against bacterial infections on human epithelial cell lines.

Solaromics: Proteomics of solid fuel cells

Solar energy and nanotechnology converge in biofuels research. Carbohydrate polymers are the most abundant organic substances on our planet yet we still do not understand how they are generated, nor how polymer lengths are controlled, be they in the form of wheat starch granules or liver hepatic cell glycogen granules. This information could allow better harvest of biomass carbon for sustainable bioenergy. For bioethanol, starch represents a viable option, giving rise to short-medium term commercialization opportunities, and a long-term complement to cellulosic feedstocks as even switchgrass yield can be ‘grown’ via molecular traits. Lignocellulosics should be the long-term answer, but starch proteomics will be a big piece of the biofuels picture for many years to come.

Photosynthesis is a process by which plants and green algae use energy from the sun to store and transform carbon dioxide from the atmosphere into a glucose reserve called starch granules. Therefore, starch represents microscopic units of trapped solar energy that can be converted into other useful forms of energy as required. For example, plants use their own starch to continue growing during night time while animals, including humans, use starch as food to sustain life. Starch can also be transformed into other forms of energy, such as ethanol, that powers our vehicles and industries. This collective process of solar energy storage in plants, in essence, is what makes biofuel possible.


*Click to enlarge the above images* (by www.apostrophe-design.be/)

Given starch's immediate importance for biofuels, the processes that control starch formation and transformation are not well understood. We know that biochemical processes are driven by protein "machinery" during starch biosynthesis. However, we do not know exactly which proteins are involved, or how they coordinate this process.

The Altosaar Lab has been “stuck on starch” for some time. We’ve been studying the surface of the starch granule since 1984. The ‘protein community network’ and the proteomic “Systems” landscape at the solid granule surface is becoming a hot spot for bioenergy research. We view the amyloplast as a bio-battery, solid fuel cells storing solar energy as fixed carbon. We collaborate with photosynthesis physiologists, starch biochemists, mass spectrometry proteomics experts, bioinformaticians, plant scientists and statisticians not only here at SITE and the Ottawa Institute of Systems Biology but also with experts at Montana State University, Carleton University, York U, UNB, and U of Calgary. The goal of our collaborative Starch Regulomics Project is to understand how photosynthetic organisms harness energy from starch and to use those discoveries to improve the means of biofuel production. Long term goals include increasing the yield of starch per hectare (i.e. improved energy yield) and improving the amount of extractable energy from the same amount of biomass (i.e. improved recovery yield).

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Starch Granule Associated Proteins (SGAPs) were first reported by our lab in 1984. LCA-treated GMA sections of starch granules from wheat showed growth rings (arrowhead) in the large type A starch granules. Lectin binding also occurs in the smaller type B granules (arrow) but no growth rings are visible. "Halos" are discernible around both types of granules (Fig. 7) and we have been in hot pursuit of this proteomic-calyx ever since, using advanced biochemical and molecular genetic strategies.

proteins

These are not ‘random’ proteins stuck to the surface of the bio-battery. When expressed in transgenic corn (Fig. F), they can have dramatic regulatory effects that need to be urgently dissected. From wheat the starch granule associated protein sporting a unique tryptophan-rich binding domain, puroindoline (Pin), regulates a 25% increase in oil! – implications to biomass and biodiesel outcomes are obvious. Representative publications include:

Miller Yiu Fulcher Altosaar 1984 Food Microstructure 3:133-9 (Figs. 5-10 panel)
Zhang J, Martin JM, Beecher B, Lu CF, Hannah LC, Wall ML, Altosaar I, Giroux MJ. 2010. The wheat puroindoline genes increase germ size and seed oil content in transgenic corn. Plant Molecular Biology 74:353–365 (Fig. F)
Wall ML, Wheeler HL, Smith JC, Figeys D, Altosaar I. 2010. Mass spectrometric analysis reveals remnants of host-pathogen molecular interactions at the starch granule surface in wheat endosperm. Phytopathology 100: 848-854
Wall ML, Wheeler H, Huebsch MP, Smith JC, Figeys D. Altosaar I. 2010. The tryptophan rich domain of puroindoline is directly associated with the starch granule surface as judged by tryptic shaving and mass spectrometry. Journal of Cereal Science 52: 115-120
Feiz L, Wanjugi HW, Melnyk CW, Altosaar I, Martin JM, Giroux MJ. 2009. Puroindolines co-localize to the starch granule surface and increase seed bound polar lipid content. J Cereal Science 50: 91–98

“The starch grain . . . . opens the door to the establishment of a new discipline, … the molecular mechanics of organized bodies.” Carl Nägeli, 1858.

Molecular Bio-Pharming:

Production of Human Pharmaceutical Proteins in Transgenic Plants Pharmaceutical polypeptides are not only free of viral contamination but may also enjoy improved recovery economics when produced in a plant background. cDNA's coding for medically-important polypeptides (MIPs) are being cloned into a series of suitable plant transformation vectors. Human breast milk protein soluble CD14 (sCD14), for example, is an integral member of the Innate Immune System and we are studying not only its biology and fate in various deployment scenarios but its cheap, safe and large-scale biological production in Plant-Made Pharmaceutical Production Platforms such as transgenic seed compartments. Candidate proteins (CP) include bifunctional "humanized" mouse monoclonal antibodies active against breast cancer oncogenes like anti-HER2, growth factors, serum albumin and potential HIV-family vaccine antigens. Various promoters and signal peptides are being studied to see which ones target the CP into various deposition sites (cytoplasm, organelles, vacuoles, protein bodies) in the endosperm cell environment. Short-term goals include maximal synthesis of CP in the seed and maximal recovery of the CP from flour while long-term goals pursue maximal biological activity of CP in oral delivery animal trials. Transformation of tobacco plants, as well as rice and oat cereal plants use the Biolistic (gene-gun) and Agrobacterium-based methodologies, both are ongoing in our lab. Transgenic plant regeneration (tissue culture, antibiotic selection), growth and maintenance, PCR, Southern analyses follows proven protocols. Western blot studies and/or ELISA's with anti-CP antibodies are used to quantitate protein levels. Immunoaffinity chromatography is used to purify plant-derived (pd) CP by conventional biochemical protein sciences including FPLC and HPLC. Immunolocalization in seeds correlates deposition sites with particular signal peptides, sorting signals, and glycosylation patterns. Glycosylation status is monitored by electro-spray mass spectrometry and hydrazinolysis. With transgenic seeds yielding high CP recoveries, chemico-physical methods of protein flour fractionation are also being explored. Throughout our research, DNA synthesis technologies are incorporated for codon-bias optimization and site-directed mutations, with and without amino acid modifications, for studying both biological activity and potential glycosylation.

Clearly the potential impacts on human health from climate change are of significant concern. The top three Green House Gases are carbon dioxide (CO 2), methane (CH 4) and nitrous oxide (N 2O). Although very little N 2O is released to the atmosphere, it is the most potent GHG and accounts for approximately 7% of yearly GHG potentials. It has 310 times the warming potential of CO 2 per molecule and an atmospheric half-life of 120 years. Besides its heat absorbing capacity, N 2O helps deplete stratospheric ozone. Atmospheric concentrations of N 2O have grown by 17% since 1750 and at present, N 2O increases 0.3% per year. Food production contributes a small proportion of GHGs but is responsible for 80% of N 2O emissions nationally, primarily through use of nitrogen fertilizers. Dissimilative denitrification is the reduction of nitrate (NO 3 -) to dinitrogen gas (N 2) through the anaerobic respiration of N. We are amplifying N 2O breakdown, engineered masterfully by Pseudomonas over eons of evolution, by transferring the nos gene into crop plants using rhizospheres signal peptides to amass a stable soil ameliorant into food production cycles. Call it Atmospheric Medicine via Genetically Engineered soils!

Selected Publications:

Fladung M, Altosaar I, Bartsch D, Baucher M, Boscaleri F, Gallardo F, Häggman H, Hoenicka H, Nielsen K, Paffetti D, Séguin A, Stotzky G, Vettori C. 2012. European discussion forum on transgenic tree biosafety. Nature Biotechnology 30(1): 37-38.
Koziol AG, Marquez BK, Huebsch MP, Smith JC, Altosaar I. The starch granule associated proteomes of commercially purified starch reference materials from rice and maize. J Proteomics In press, ms #JPROT-D-11-00373R2, accepted Oct 21, 2011.
Wan S, Johnson A, Altosaar I. 2012. Expression of nitrous oxide reductase from Pseudomonas stutzeri in transgenic tobacco roots using the root-specific rolD promoter from Agrobacterium rhizogenes. Ecology and Evolution 2(2):286-297.
Wan S, Mottiar Y, Johnson AM, Goto K, Altosaar I. 2011. Phytoremediation of nitrous oxide: expression of the nos operon proteins from Pseudomonas stutzeri in transgenic plants to assemble nitrous oxide reductase. Transgenic Research in press (accepted Sept 2, 2011 ms# TRAG1456R1).
Mehrotra M, Singh AK, Sanyal I, Altosaar I, Amla DV. 2011. Pyramiding of modified cry1Ab and cry1Ac genes of Bacillus thuringiensis in transgenic chickpea (Cicer arietinum L.) for improved resistance to pod borer insect Helicoverpa armigera. Euphytica 182(1): 87-102, DOI 10.1007/s10681-011-0501-3
Mottiar Y, Altosaar I. 2011. Iodine sequestration by amylose to combat iodine deficiency disorders. Trends Food Sci & Technology 22(6): 335-340, doi:10.1016/j.tifs.2011.02.007.
Davis LDR, Spencer WJ, Mack DR, Altosaar I. 2011. Maternal separation and gastrointestinal transit time in neonate rats. Laboratory Animals 45: 280-282, doi: 10.1258/la.2011.010167.
Davis LDR, Spencer WJ, Pham VT, Ward TL, Blais DR, Mack DR, Kaplan H, Altosaar I. 2011. 14C-Radiolabeling of proteins to monitor biodistribution of ingested proteins. Analytical Biochemistry 410: 57–61.
Zhang J, Martin JM, Beecher B, Lu C, Hannah LC, Wall ML, Altosaar I, Giroux MJ 2010. The ectopic expression of the wheat puroindoline genes increases germ size and seed oil content in transgenic corn. Plant Molecular Biology 74:353–365.
Shen XJ, Ye GY, Cheng XY, Yu CY, Altosaar I, Hua C. 2010. Characterization of an abaecin-like antimicrobial peptide identified from a Pteromalus puparum cDNA clone. Journal of Invertebrate Pathology 105(1): 24-29.
Spencer WJ, Binette A, Ward TL, Davis LDR, Blais DR, Harrold J, Mack DR, Altosaar I. 2010. Alpha-lactalbumin in human milk alters the proteolytic degradation of soluble CD14 by forming a complex. Pediatric Research 68(6):490-493
De Guglielmo-Cróquer Z, Altosaar I, Zaidi M, Menéndez-Yuffá A. 2010. Transformation of coffee (Coffea arabica L. cv. Catimor) with the cry1Ac gene by biolistic without the use of markers. Braz. J. Biol. 70: 387-393
Wall ML, Wheeler HL, Smith JC, Figeys D, Altosaar I. 2010. Mass spectrometric analysis reveals remnants of host-pathogen molecular interactions at the starch granule surface in wheat endosperm. Phytopathology. 100(9): 848-854. doi: 10.1094/PHYTO-100-9-0848
Wall ML, Wheeler H, Huebsch MP, Smith JC, Figeys D. Altosaar I. 2010. The tryptophan rich domain of puroindoline is directly associated with the starch granule surface as judged by tryptic shaving and mass spectrometry. Journal of Cereal Science 52(2): 115-120
Loit E, Hincke M, Altosaar I. 2010. Synthetic antimicrobial peptide “L8”, MHLHKTSRVTLYLL, has membrane permeabilization and bacterial aggregation activity. International Journal of Antimicrobial Agents 35(4): 410-411
Zaidi MA, Ye GY, Yao HW, You TH, Dean DH, Riazuddin S, Altosaar I. 2009. Transgenic rice plants expressing a modified cry1Ca gene are resistant to Spodoptera litura and Chilo suppressalis. Molec Biotechnol 43: 232-242, DOI 10.1007/s12033-009-9201-9
Loit E, Melnyk CW, MacFarlane AJ, Scott FW, Altosaar I. 2009. Identification of three wheat globulin genes by screening a T. aestivum BAC genomic library with cDNA from a diabetes-associated globulin. BMC Plant Biology 9: 93 (11 pp) DOI:10.1186/1471-2229-9-93
Cheng XY, Liu GZ, Ye GY, Wang HJ, Shen XJ, Wu KC, Xie JH, Altosaar I. 2009. Screening and cloning of antimicrobial DNA sequences using a vital staining method. Gene 430: 132-139
Feiz L, Wanjugi HW, Melnyk CW, Altosaar I, Martin JM, Giroux MJ. 2009. Puroindolines co-localize to the starch granule surface and increase seed bound polar lipid content. J Cereal Science 50: 91–98
Taga I, Agbor GA, Djountsop S, Moshin ZA, Soh Oumbe VA, Altosaar I, Ngogang JY. 2009. Iodine deficiency in children 7-19 years old in Eastern Province of Cameroon. Tropical Medicine & International Health 14: 311-315
Gulbitti–Onarici S, Zaidi MA, Taga I, Ozcan S, Altosaar I. 2009. Expression of Cry1Ac in transgenic tobacco plants under the control of a wound-inducible promoter (AoPR1) isolated from Asparagus officinalis to control Heliothis virescens and Manduca sexta. Molecular Biotechnology 42:341–349. doi:10.1007/s12033-009-9168-6
Pham VT, Altosaar I, Duhig MN, Kaplan H. 2009. Glass-immobilized glycated trypsin: A novel modified trypsin that is remarkably thermostable. J Molecular Catalysis B: Enzymatic 58:48–53. DOI:10.1016/j.molcatb.2008.11.007
Taga I, Oumbe VAS, Johns R, Zaidi MA, Yonkeu JN, Altosaar I. 2008. Youth of west-Cameroon are at high risk of developing IDD due to low dietary iodine and high dietary thiocyanate. African Health Sciences 8: 180-185. http://www.ajol.info/viewarticle.php?jid=45&id=43888
Loit E, Wu K, Cheng X, Hincke M, Altosaar I. 2008. Functional whole-colony screening method to identify antimicrobial peptides. J Microbiological Methods 75:425-431. DOI:10.1016/j.mimet.2008.07.023
Tackaberry ES, Prior FA, Rowlandson K, Tocchi M, Mehic J, Porter S, Walsh M, Ganz PR, Sardana RK, Altosaar I, Dudani AK. 2008. Sustained expression of human cytomegalovirus glycoprotein B (UL55) in the seeds of homozygous rice plants. Molecular Biotechnology 40(1): 1-12. DOI 10.1007/s12033-007-9029-0
Kim S, Kim C, Li W, Kim T, Li Y, Zaidi MA, Altosaar I. 2008. Inheritance and field performance of transgenic Korean Bt rice lines resistant to rice yellow stem borer. Euphytica 164: 829–839. DOI 10.1007/s10681-008-9739-9
US Patent 7,214,862 Production of Human Granulocyte Macrophage-Colony Stimulating Factor in Plants. May 8, 2007. "The present invention discloses a method of producing granulocyte-macrophage colony stimulating factor (GM-CSF) in a plant comprising, transforming the plant with a genetic construct comprising a regulatory region functional in the plant, operably associated with a GM-CSF coding sequence, or a fragment or a derivative thereof, operably associated with a transcriptional terminator, and expressing the GM-CSF. Also disclosed are transgenic plants, seeds and cells comprising GM-CSF coding sequences and plant optimized GM-CSF coding sequences."
Homrich MS, Passaglia LMP, Pereira JF, Bertagnolli PF, Pasquali G, Zaidi MA, Altosaar I, Bodanese-Zanettini MH. 2008. Resistance to Anticarsia gemmatalis Hübner (Lepidoptera, Noctuidae) in transgenic soybean (Glycine max (L.) Merrill Fabales, Fabaceae) cultivar IAS5 expressing a modified Cry1Ac endotoxin. Genetics and Molecular Biology 31(2): 522-531 DOI link
Chen M, Liu ZC, Ye GY, Shen ZC, Hu C, Peng YF, Altosaar I, Shelton AM. 2007. Impacts of transgenic cry1Ab rice on non-target planthoppers and their main predator Cyrtorhinus lividipennis (Hemiptera : Miridae) - A case study of the compatibility of Bt rice with biological control. Biological Control 42: 242-250.
Blais DR and Altosaar I. 2007. Humanizing infant milk formula to decrease postnatal HIV transmission. Trends in Biotechnology 25: 376-384. DOI link
Schroder M, Poulsen M, et al., Altosaar, I, Knudsen I. 2007. A 90-day safety study of genetically modified rice expressing Cry1AB protein (Bacillus thuringiensis toxin) in Wistar rats. Food Chemical Toxicololy 45: 339-349

Recent Supervised Theses

Wall ML, PhD 2011 The Starch Granule Surface: technological and biological implications of puroindoline and host-pathogen interactions.
Davis LDR, MSc 2010 The biodistribution of 14-C in the digestive organs of rats fed [14-C]CD14 protein using in vacuo radiolabeling..
Loit, E. Improving food quality and quantity: Diabetes-associated Glo-3 genes in wheat; Synthetic antimicrobial peptides from a random oligonucleotide library. PhD 2009.

Pham, V. T. Application of in vacuo chemical modification for protein characterization and enhancement of the physico-chemical properties of proteins. PhD 2008.
McNulty, M. Preliminary characterization of wheat, Triticum aestivum, embryo globulins. M.Sc. 2006.
Staebler JM. Clearing the air: Expression of Nitrous Oxide Reductase from Pseudomonas stutzeri in Transgenic Plants. M.Sc. 2006.
Mao, JQ. Improved resistance to insects in maize (Zea mays L.) and cowpea (Vigna unguiculata L.). M. Sc. 2005.
Blais DR, Fate and function of soluble CD14 at ocular and gastrointestinal surfaces and in transgenic tobacco seeds. PhD 2005.
Wu K, Solubility and manipulation of disulfides in puroindoline-b, recombinant pin-b shows antifungal activity. M.Sc 2005.
Alli Z, Novel vaccines for Hepatitis B Virus: plant foods to produce and deliver edible vaccines against common infectious diseases. PhD 2004.
Girard L, Expression of recombinant proteins in plant cell cultures, University of Rennes, Ecole Doctorale Vie Agronomie Santé, Rennes, France.
Co-supervisor Didier Courtois, PhD 2004.
Panahi, M. Ph.D. 2002. Biological activity of recombinant prohormone precursor insulin like growth factor-1B (1GF-1B) and the mature form of IGF-1 expressed in transgenic plants.
Alli, Z. M.Sc. 2001. Expression of biologically active human granulocyte macrophage colony stimulating factor in the seeds of transgenic tobacco.
Callaghan, M. M.Sc. 2001. Chimeric orthohepadnavirus core particles for oral delivery of vaccines. Part I. Transformation of tobacco plants with a gene encoding a C-terminus truncated hepatitis B virus core protein. Part II. Construction of a Woodchuck Hepatitis virus core protein-based universal epitope carrier and test expression in E. coli

Current Research Group

Whitney Faiella – UROP scholar; Hwan-Hee 'Connie' Son - Undergraduate Research Scholar; Pamela Hammoud - BScHon Thesis; Trevor Greenham - MSc student; Adam Koziol, Shen Wan, Tonya Ward, George Styles - PhD students; Ibrahim Taga - Postdoctoral Fellow; Mohsin Zaidi - Visiting Scientist.

Last modifications: 2011.11.30

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