Presentation on theme: "The Effects of PUGNAc on O-GlcNAC Levels and Dauer Formation in C"— Presentation transcript:
1The Effects of PUGNAc on O-GlcNAC Levels and Dauer Formation in C The Effects of PUGNAc on O-GlcNAC Levels and Dauer Formation in C. elegansBy Nicole Kiess
2NeedPicture 1: Shows the rate of Type 2 Diabetes in relationship to obesity over 10 years.
3NeedEstimated prevalence of diagnosed and undiagnosed diabetes in people aged 20 years or older, by age group, United States, 2007Picture 2: Shows percent of people in different age groups with Type 2 Diabetes.
4Knowledge BasePicture 3: Is a model of what happends in the body of a Type 2 Diabetic
5Knowledge BasePicture 4a: Shows the pathway that regulate aging in C. Elegans.Picture4 b: Shows the model for the regulation of life span and sensory perception.
6Knowledge Base Picture 5: (Top) Insulin-like signaling pathway. (Bottom) C. Elegnas life cycle.
7Literature ReviewParadis, et al,(1998) showed that mg144 does not activate AGE-1 P13K and that Akt/PBK is not dependent on AGE-1/P13KPicture 6a: Shows normal growth conditionsPicture 6b: Shows dauer formation conditions
8Literature ReviewArias, et al, 2004 showed that PUGNAc increases O-GlcNAc levels in rat skeletal muscle
9Literature ReviewHanover, et al, 2005, findings show that OGT modulates macronutrient storage and dauer formation in C. ElegansPicture 7a: Insulin like signaling pathwayPicture 7b: Shows percentages of dauers in different strains
10Literature ReviewForsythe, et al, 2006 demonstrated that enzymes of O-GlcNAc cycling perfect insulin-like signaling in response to nutrient fluxPicture 8a: Insulin like signaling pathwayPicture 8b: Dauer formation in different strains with temperature as a variable.
11PurposeTherefore the purpose of this experiment is to observe the O-GlcNAc levels, using densinometry, and dauer formations with PUGNAc as a variable.
12HypothesisNull- There will be no change in O-GlcNAc levels and no change in dauer formationAlternative- The levels of O-GlcNAc will rise and dauer formation will rise
13MethodThe effects of PUGNAc on O-GlcNAC levels and dauer fomation in the insulin-like signaling pathway of C. elegansControlWild Type N2Ogt-1 (ok430)Oga-1 (ok1207)Daf-2 (e1370)50um100um50um100um50um100um50um100umC. Elegans will be cultured at 16 degrees CO-GlcNAC levels will be observed by treating C. elegans with RL1 and RL2 antibodiesPictures will be taken with camera under immunofluorescent microscope and photoshop will measure percentagesAdults of each strain lay embryos overnight at 15 degrees C. Cultures shifted to 22.5 degrees C and are scored 3 days later as either dauer or non dauerStatistical Analysis will be run with SPSS and an ANOVA
14ProtocolC. elegans will be cultured in petri dishes with the Nematode Growth Medium (NGM) and will be fed Escherichia coli (E. coli)Picture 9: How the C. elegans will be culturedDrawn by Victoria Wei
15Protocol RL1 and RL2 dilated 1:400 and given to C. Elegans Picture 10: Life cycle of C. elegans
16Protocol Fluorescent microscope and camera used for picture Photoshop used for percentagesPicture 11: A) Fluorescent picture to show O-GlcNAc levels with RL-2B) Fluorescent picture to show O-GlcNAc levels with RL-2Hanover, et al, 2005
17Do-Ability Need to Purchase: Already Obtained RL-1 RL-2 C. elegans strainsE. coliPUGNAcAlready ObtainedFluorescent microscopeCameraPhotoshopPetri Dishes
19BibliographyBiggs, Alton, Whitney Crispen Hagins, Chris Kapicka, Linda Lundgren, Peter Rillero, and Kathleen G. Tallman. Biology The Dinamics of Life. Columbus, Ohio: The McGraw-Hill Companies Inc., 2004Cauter, Eve V., Rachel Leproult, Esra Tasali, and Karine Spiegel. "Effects of poor and short sleep on glucose metabolism and obesity risk." Nature Reviews Endocrinology 5 (2009):Chang, Annette M., and Jeffrey B. Halter. "Aging and Insulin secretion." AJP-Endocrinol Metab 284 (2003).Clevenger, Christopher M., Pamela Parker Jones, Hirofumi Tanaka, Douglas R. Seals, and Christopher A. DeSouza. "Decline in insulin action with age in endurance-trained humans." Journal of APplied Physiology 93 (2002):"Introduction to C. Elegans." Genes, Genomes, and Human. 07 May 2009 <http://avery.rutgers.edu/WSSP/StudentScholars/project/introduction/worms.html>.Kao, G., C. Nordenson, M. Still, A. Ronnlund, S. Tuck, and P. Naredi. "ASNA-1 positively regulates insulin secretion in C. elegans and mammalian cells." Cell 128 (2007):Kimura K.D., Tissenbaum H.A., Liu Y., Ruvkun G. (1997) daf-2, an insulin receptor-like gene that regulates longevity and diapause in Caenorhabditis elegans.Science 277:942–946.Kodama, Eiji, Atsushi Kuhara, Akiko Mohri-Shiomi, Koutarou D. Kimura, Masatoshi Okumura, Masahiro Tomioka, Yuichi Iino, and Ikue Mori. "Insulin-like signaling and the neural circuit for integrative behavior in C. Elgans." Genes and Development 20 (2006):Manzella, Debra. "Obesity and Type 2 Diabetes - Why Being Overweight Can Damage Health." Diabetes - Diabetes Management - Signs of Diabetes - Symptoms of Diabetes. 04 May 2009 <http://diabetes.about.com/od/symptomsdiagnosis/a/obesity.htm>.Muzumadar, Radhika, Xiaohui Ma, Gil Atzmon, Patricia Vuguin, Xiaoman Yang, and Nir Barzilai. "Decrease in Glucose-Stimulated Insulin Secretion With Agine Is Independent of Insulin Action." Diabetes 53 (2004):Norman, James. "Introduction to Insulin: insulin discovery and actions." Endocrine Diseases: thyroid, parathyroid adrenal and diabetes. 07 May 2009 <http://www.endocrineweb.com/diabetes/2insulin.html>.Paradis, Suzanne, and Gary Ruvkun. "Caenorhabditis elegans Akt/PKB transduces insulin receptor-like signals from AGE-1 P13 kinase to the DAF-16 transcription factor." Genes and Development 12 (1998):"Type 2 Diabetes: Conditions, Treatments, Resources - American Diabetes Association." American Diabetes Association Home Page. 04 May 2009 <http://www.diabetes.org/type-2-diabetes.jsp>.Twyman, Richard. "Model organisms: The nematode worm." The Human Genome. 07 May 2009 <http://genome.wellcome.ac.uk/doc_WTD html>.Waksman Student Scholars. "Introduction to C. Elegans." Genes, Genomes, and Human Genetics. 12 May 2009 <http://avery.rutgers.edu/WSSP/StudentScholars/project/introduction/worms.html>.