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Alterations in Hsp27

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Jon Zonum

on 28 April 2010

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Transcript of Alterations in Hsp27

Alterations in Hsp27 By Jonathan Munoz
Advised by Dr. Jody Martin What's Hsp27? Thermal
Ultraviolet
Oxidative (Hydrogen Peroxide)
Apoptosis
Free Radicals Answer: A Protective Protein that maintains homeostasis Focusing Question:

How will changing the 137th amino acid, cytosine, affect the localization, olgimerization, and shape of Hsp27? Hsp27 Responds Hsp27 Prevents

Phosphorilating -Membrane Damage
Oligmerating -Denaturization
Localizing -Necrosis
-Apoptosis Environmental Stressors The Mutation Polar amino acid- cystine

Non-Polar Amino Acid Replaced with Video Imaging

Stressed native/mutant a7r5 cells
Microscopic Images
Each minute for sixty minutes
150 micromolar Hydrogen Peroxide dillution
DTNB (Dithio-bis-2-nitrobenzoic acid)
Cell types

a7r5 and H9c2 myoblasts
Applicable to Human studies
Increased amounts of Hsp27
Easily Transfected
Readily available

Cancer

-Stops Apoptosis
-Malignant cells survive Saves Structural Supports Materials and Methods Hsp27 with
cfp fusion C137A Hsp27
mutation Before Hydrogen
Peroxide dilution Cell bleaching
Lens Focusing

T147 Breast Cancer cells
DTNB (Dithio-bis-2-nitrobenzoic acid)

Obstacles After Hydrogen
Peroxide dilution We stressed six different H9c2 myoblasts and recorded the percentage of surviving cells.
Survivability Test
We tested six different cell lines:

1. one native species
2. one overexpressing hsp27
3. one with a green fluorescent protein attached
4. one with C137A Hsp27 mutation attached to a cfp
5. one infected with an empty virus
6. one overexpressing hsp27 attached to a cfp Each were subjugated to a 46° heat shock for a 1/2 hour and a 0, 250, 500, 750, 1000 M dilution of Hydrogen Peroxide for 4 hours. -Fluorescent proteins inhibit Hsp27
-Result in false positives
-Extra 27 kilodaltons
-Possibly test new theorem
Discussion Hsp27 with a cfp or gfp fusion had a significantly higher cell death. Conclusion Hypothesis: Inconclusive White blebing and cell convergance appeared in both the C137A mutation and unaltered a7r5 species. but... Results Video Imaging Hydrogen Peroxide No Treatment Both a7r5 strains, the native species and the c137a mutant experianced...

White blebbing
Cell shrinkage
Fading Away Anova Test
-P-value proves correlation Both a7r5 strains, the native species and the c137a mutant showed signs of...

Cell shrinkage
Fading Away Survivability General decreasing trend

Overexpression of hsp27 increased survivability
cfp and gfp fusions showed slight decrease

Blank Virus harmed cell survival C137A Hsp27 + cfp fusion Native + fluourescent protein Alterations in Hsp27 Acknowledgments

SIR Staff Dr. Martin Loyola University Medical Center IMSA Native + fluorescent protein C137A Hsp27 + cfp fusion

Arrigo, A., Simon, S., Gibert, B., Kretz-Remy, C., Nivon, M., Guillet, D., Moulin, M., Diaz-Latoud, C., & Vicart, P. (2007). Hsp27 (HspB1) and aB-crystallin (HspB5) as therapeutic targets.

Federation of European Biochemical Societies, 581, 3665-3674..Embryonic Rat Thoracic Aorta Cells (A7r5 Line). (n.d.). Retrieved October 21, 2009, from http://digitalimagingu.com/galleries/digitalimages/a7r5/a7r5cells.html http://digitalimagingu.com/galleries/digitalimages/a7r5/a7r5cel

Goldman, B. (n.d.). Heat Shock Proteins' Vaccine Potential. Retrieved October 21, 2009, from http://www.antigenics.com/whitepapers/hsp_potential.html

Heat shock proteins: basics. (n.d.). Retrieved September 30, 2009, from http://www.antigenics.com/products/tech/hsp/

Molecular Aspects of the Stress Response: Chaperones, Membranes and Networks (Advances in Experimental Medicine and Biology). (2006). New York: Springer

Prasad, M., & Goyal, R. (2004). Differential modulation of voltage-dependent K+ currents in colonic smooth muscle by oxidants. American Journal of Physiology, 286, C671-C682. References
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