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
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
• 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!!
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
HLA-DQA1 protein structure
•cell-cell recognition•cell surface
receptors•immune system
•cellular interaction
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
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/
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??
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?
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?
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
(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
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
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
(1) Hypothesis
• The addition of gliadin to mouse intestinal cells should show decreased HLA-DQA1 expression.
(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 (-)
(2) How does gliadin affect small intestine phenotype?
Method: Observe phenotype of mouse intestinal cells
following gliadin treatment
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
CD and the small intestine...
Image: Children’s Hospital Boston. 2009. http://www.youngmenshealthsite.org/gluten_free_diet.html
(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.
(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
(3) How is gliadin acting at the molecular level?
Method: TAP-tag Label gliadin and determine how it is
affecting HLA-DQA1
Gliadin may function in this way...
Gliadin peptide?
Cell Surface
T-Cell
T-cell Receptor
DQA1
Antigen Presenting
Cell
Gliadin peptide?
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?
What role do the predicted functional partners play?
Question: Is gliadin regulating/binding any of these interacting proteins?
(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
(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
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.
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?
THE END
Image: http://www.herbalgranny.com/2009/09/06/herbal-treatment-for-celiac-disease/
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