hla-dqa1 and celiac disease
DESCRIPTION
HLA-DQA1 and Celiac Disease. Presented by Cassie Bac. Image: http://www.scientificamerican.com/article.cfm?id=celiac-disease-insights. Background on Celiac Disease (CD). Autoimmune disease that creates a toxic immune response to gluten-- damaging the lining of the small intestine - PowerPoint PPT PresentationTRANSCRIPT
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HLA-DQA1 and Celiac Disease
Presented by Cassie Bac
Image: http://www.scientificamerican.com/article.cfm?id=celiac-disease-insights
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Background on Celiac Disease (CD)
• Autoimmune disease that creates a toxic immune response to gluten-- damaging the lining of the small intestine
• Affects 1 in 133 healthy individuals in the United States
• Can appear at anytime throughout life!
• No known cure
Image: http://www.reallygreatfoods.com/
No wheat, rye, or barley allowed!!
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What are the classic symptoms of CD?
• Malnutrition
• Abdominal cramping
• Intestinal gas and stomach bloating
• Chronic diarrhea
• Weight loss or weight gain
• Dermatitis Herpetiformis
Images: http://healthhype.com; http://www.ehow.com/about_5141982_pediatric-celiac-disease-symptoms.html
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HLA-DQA1 protein structure
•cell-cell recognition•cell surface
receptors•immune system
•cellular interaction
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How is HLA-DQA1 associated with CD?
Peptide
Cell Surface
Images: http://en.wikipedia.org/wiki/Gluten_immunochemistry
T-Cell
T-cell Receptor
DQA1
Antigen Presenting
Cell
Peptide
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What is still unknown?
What mechanism causes the toxic immune system
response in Celiac Disease?
Image: http://jaymecarleton.com/secondnature/green-news/2011/04/toxic-toys-r-us/
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What do we know about gliadin?
• Gliadin= a protein derived from gluten and the toxic factor involved in Celiac Disease!
Question: How does gliadin act??
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Experimental Approach
1. Chemical Genetics How does gliadin affect HLA-DQA1 expression
levels?
2. PhenotypeHow does gliadin affect the phenotype of the
small intestine?
3. Interactions
How is gliadin acting at the molecular level?
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Experimental Approach
1. Chemical Genetics How does gliadin affect HLA-DQA1 expression
levels?
2. PhenotypeHow does gliadin affect the phenotype of the
small intestine?
3. Interactions
How is gliadin acting at the molecular level?
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Experimental ApproachHow does gliadin affect HLA-DQA1 expression
levels?
2. PhenotypeHow does gliadin affect the phenotype of the
small intestine?
3. Interactions
How is gliadin acting at the molecular level?
1. Chemical Genetics
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(1) How does gliadin affect HLA-DQA1 expression levels?
Method: Inject mouse intestinal cells with gliadin (+ other varieties) and measure HLA-DQA1 expression
levels with microarray
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Why use a mouse model?Mouse protein domains
H2-AaHuman protein domains
HLA-DQA1
71.8% conserved protein structure
Images: From SMART. HLA-DQA1 sequence search results. Retrieved from http://smart.embl-heidelberg.de/smart/show_motifs.pl; SMART. H2-Aa sequence search results. Retrieved from
http://smart.embl-heidelberg.de/smart/show_motifs.pl
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Microarray data
http://www.ncbi.nlm.nih.gov/geo/gds/profileGraph.cgi?&dataset=zAAqAB7R9Q&dataset=yLSxM0vrwr$&gmin=108.726000&gmax=3651.000000&absc=&uid=24204411&gds=2042&idref=213831_at&annot=HLA-DQA1
WT patient
expression levels
Celiac patientexpression
levels
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(1) Hypothesis
• The addition of gliadin to mouse intestinal cells should show decreased HLA-DQA1 expression.
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(1) Expected Data
Mouse Intestinal
Cells
HLA-DQA1 expression + +/-
WT WT + gliadin Celiac patient
Celiac patient + HLA-DQA1
Celiac patient + Gliadin
- - -
= High HLA-DQA1 expression (+)
= Low HLA-DQA1 expression (-)
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(2) How does gliadin affect small intestine phenotype?
Method: Observe phenotype of mouse intestinal cells
following gliadin treatment
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What do we know about intestinal cells?
Images: http://en.wikipedia.org/wiki/Small_intestine; http://en.wikipedia.org/wiki/Microvillus
Microvilli line intestinal cell membrane; involved in absorption.
Small intestine
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CD and the small intestine...
Image: Children’s Hospital Boston. 2009. http://www.youngmenshealthsite.org/gluten_free_diet.html
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(2) Hypothesis
The addition of gliadin to mouse intestinal cells should cause physical damage to the microvilli. In addition, the cells
that showed low HLA-DQA1 expression should show
increased intestinal damage.
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(2) Expected Data
Mouse intestinal
cells
WT WT + gliadin Celiac patient
Celiac patient + HLA-DQA1
Celiac patient + gliadin
Images: http://health.uml.edu/thc/HealthIssues/Meghan_Donovan/Meghans_Homepage.html
HLA-DQA1Expression
Levels
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(3) How is gliadin acting at the molecular level?
Method: TAP-tag Label gliadin and determine how it is
affecting HLA-DQA1
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Gliadin may function in this way...
Gliadin peptide?
Cell Surface
T-Cell
T-cell Receptor
DQA1
Antigen Presenting
Cell
Gliadin peptide?
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HLA-DQA1 protein interactions
Image: From STRING 8.3. HLA-DQA1 evidence map. Retrieved from: http://stringdb.org/newstring_cgi/show_network_section.pltaskId=8jlOaZSDtoSa&interactive=yes&advanced_menu=_unassigned&network_flavor=evidenc
Question: Is gliadin regulating/binding
HLA-DQA1?
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What role do the predicted functional partners play?
Question: Is gliadin regulating/binding any of these interacting proteins?
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(3) Hypothesis
Using Tandem Affinity Purification, gliadin should be found to regulate
HLA-DQA1 protein at the molecular
level.
Image: http://www.nature.com/nrm/journal/v4/n1/fig_tab/nrm1007_F1.html
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(3) Expected Data
Mass spectrometry to analyze the protein-protein
interactions
Gliadin binds HLA-DQA1
Gliadin binds interacting proteins
Image: http://www.fda.gov/Food/ScienceResearch/Laboratory
Methods/DrugChemicalResiduesMethodology/ucm113209.htm
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Future Directions• MUDPIT- Multidimensional Protein Identification Technology
•Hypothesis: HLA-DQA1 protein observed in WT patient tissue samples, not observed in celiac samples.
•Experiment: Take intestinal tissue samples from WT and celiac patients and observe specific protein identifications from the two samples.
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Future Directions• Microarray primary HLA-DQA1 interacting proteins- look at expression levels in WT vs. Celiac Disease patients. Are their levels expressed in similar patterns as HLA-DQA1?
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THE END
Image: http://www.herbalgranny.com/2009/09/06/herbal-treatment-for-celiac-disease/
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References
• University of Chicago Celiac Disease Center. Celiac Disease Facts and Figures. Print.
• SMART- http://smart.embl-heidelberg.de/
• GeneBee- http://www.genebee.msu.su/genebee.html
• http://knol.google.com/k/celiac‐disease#
• Bizzaro, N., R. Tozzoli, D. Villalta, M. Fabris, and E. Tonutti. "Cutting‐Edge Issues in Celiac Disease and in Gluten Intolerance." Clinical Reviews in Allergy and Immunology 15.1 (1997): n. pag. Web. 7 Feb 2011http://www.springerlink.com.ezproxy.library.wisc.edu/content/j17wu631711u1027/fulltext.pdf.
• http://www.ncbi.nlm.nih.gov/pubmedhealth/PMH0001280
• PubChem- http://pubchem.ncbi.nlm.nih.gov/
• STRING- http://string.embl.de/
• Homologene- http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=homologene&dopt=AlignmentScores&list_uids=123820
• Microarrays- http://www.ncbi.nlm.nih.gov/geo/gds/profileGraph.cgi?&dataset=zAAqAB7R9Q&dataset=yLSxM0vrwr$&gmin=108.726000&gmax=3651.000000&absc=&uid=24204411&gds=2042&idref=213831_at&annot=HLA-DQA1