1 / 26

Role of CR-1 in Cellular Heterogeneity and Drug Resistance

Explore the emergence of cellular heterogeneity in CR-1 expression and its implications in cancer, stem cells, and drug resistance through signaling pathways and transcriptional circuits.

danielburris
Download Presentation

Role of CR-1 in Cellular Heterogeneity and Drug Resistance

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. Emergence of Cellular Heterogeneity in Expression of an Oncofetal Protein Biplab Bose Department of Biosciences & Bioengineering IIT Guwahati

  2. Heterogeneity in Cancer Genetic heterogeneity within a tumor Mol Oncol. 2010, 4(3):267-83

  3. CFP YFP cfp yfp P P Development, 2009,136:3525-30 Science, 2002, 297(5584):1183-6 Population Heterogeneity in Gene Expression

  4. Stochasticity in Gene Expression

  5. Biological Implication of Heterogeneity Emergence of drug resistance Nat Rev Genet. 2009, 10(5):336-42

  6. Human Cripto-1 (CR-1): an Oncofetal Protein • Member of EGF-CFC family. • Expressed as membrane bound molecule • Over-expressed in various cancers • Functional in membrane-bound and soluble form During Development: • Patterning of the Anterior/Posterior axis • Specification of mesoderm and endoderm during gastrulation Intracellular Signaling Pathways: • Nodal/ALK4/Smad-2 • Glypican-1/c-Src/MAPK • Glypican-1/c-Src/AKT In Cancer: • Triggers proliferation • Induction of cellular migration and invasion • promotion of angiogenesis • Stimulates EMT

  7. CR-1  Heterogeneous Expression of CR-1 CR-1 expression is heterogeneous: two sub-populations exists Human melanoma cell lines Human embryonal carcinoma cell lines CR-1 (+)ve : ~50% CR-1 (+)ve : 2% - 5% Stem Cells. 2010, 28(8):1303-14 Cell Cycle. 2013 May 1;12(9):1450-6.

  8. Transcriptional Circuit Affects Expression Heterogeneity Linear Circuit Positive feedback

  9. A Possible Feedback Circuit for CR-1 Current Information Hypothesis

  10. CR-1 Induces Its Own Expression U-87 MG cells, treated with recombinant CR-1, 24 hr Real-time PCR Western Blot n = 2.3713; a = 10.85 K=156.99; R2 = 0.9747

  11. Treatment with CR-1 does not effects on mRNA stability Treatment condition: 200 ng/ml CR1-GST, 5 µg/ml Act-D Real Time PCR: No significance difference between treated and untreated (Rank sum test, P =0.127) 11

  12. The Pathway of Induction Western Blot PCR Treatment condition: Cells: U-87 MG; SB431542: 10 µM; CR1-GST or GST: 200 ng/ml; 24 hr

  13. The Pathway of Induction MCF-7: Expresses SMAD4 HT-29: Does not express SMAD4

  14. Transcriptional Circuit Affects Expression Heterogeneity Linear Circuit Positive feedback

  15. Induction of CR-1 Expression is Heterogeneous Treatment: U-87 MG cells treated with different conc. of recombinant CR-1, 24 hrs Induction of CR-1 expression happens only in a minority subpopulation

  16. Dose-dependent Behavior of CR-1 Positive Cells Dose-dependent Behavior of CR-1 Positive Cells Treatment: U-87 MG cells treated with different conc. of recombinant CR1, 24 hrs Overton histogram subtraction used to find CR-1 (+)ve population

  17. CR-1(-)ve CR-1(+)ve Differential Expression of Pathway Molecules RT-PCR Two subpopulations are sorted by FACS and gene expression was checked by RT-PCR

  18. Bimodality in Expression May Be Obscured

  19. Signature of Bimodality in Expression Noise Noise in CR-1 expression When protein expression, is unimodal and increases by increase in rate of transcription CR-1 expression was induced with different doses of recombinant CR-1 Mol Biosyst, 2012, 8(11):3068-76

  20. Simulated for three conditions: Signature of Bimodality in Expression Noise Simulated using MATLAB “Low cells” has lower mean and variance “High cells” have high mean and variance % of “High cells” varied: 0% to 100%

  21. Signature of Bimodality in Expression Noise Simulation Flow Cytometry Data

  22. Biological Implication of CR-1 Positive Subpopulation Stochasticity in gene expression  Spontaneous emergence of heterogeneity  Cells in different states  Some cells having drug resistance and/ or Some cells having Stemness Ref: Cell, 2011, 146: 633–644

  23. CR-1(-)ve CR-1(+)ve CR-1 Positive Cells May Not be Cancer Stem Cells RT-PCR: Two subpopulations are sorted by FACS and gene expression was checked by RT-PCR

  24. Isotype Control Untreated CR1-GST (400 ng/ml) CR-1 Positive Cells are MDR-1 Positive • MDR-1 is an ABC-transporter involved in multidrug resistance • CSC like cells isolated from U-87 MG have higher MDR-1 expression CR-1 and MDR-1 are co-induced in a small subpopulation: indicates co-regulation

  25. Cells Do Play Dice Signal PLoS ONE 10(2): e0116748

  26. FlowPy biplabbose@iitg.ernet.in @bose_biplab http://flowpy.wikidot.com/ Research student: Dr. Pojul Loying Collaborator: Dr. Janvie Manhas Dr. Sudip Sen, AIIMS Financial Support: DBT

More Related