Production of pluripotent cells through inhibition of bright/ARID3a function
The present invention involves the identification of Bright/ARID3a as involved in the regulation of pluripotency in cells, and the targeting of that function for the regulation of pluripotency. Thus, methods of de-differentiating cells into pluripotent cells are provided, as well as methods for re-differentiating such cells in a controlled fashion.
1. A method of rendering a differentiated cell multipotent comprising:
(a) providing a differentiated cell; and
(b) contacting said cell with an inhibitory nucleic acid that reduces Bright/ARID3a expression to induce de-differentiation in said cell; and
(c) culturing said cell in medium including leukemia inhibitory factor,
wherein de-differentiation renders said cell multipotent.
2. The method of claim 1 , wherein said cell of step (a) is a bone marrow cell, fibroblast cell or a spleen cell.
3. The method of claim 1 , wherein said cell of step (a) is a peripheral blood cell.
4. The method of claim 1 , wherein said inhibitory nucleic acid that reduces Bright/ARID3a expression is an interfering RNA.
5. The method of claim 4 , wherein interfering RNA is an shRNA.
6. The method of claim 5 , wherein said shRNA is expressed from an expression vector.
7. The method of claim 1 , wherein inhibition of Bright/ARID3a function is reversible.
8. A method of reprogramming a differentiated cell comprising:
(a) providing a differentiated cell;
(b) contacting said cell with a an inhibitory nucleic acid that reduces Bright/ARID3a expression to induce de-differentiation in said cell; and;
(c) contacting said cell, following de-differentiation, with a signal selected to produce a re-differentiated cell phenotype;
(d) culturing said cell with said signal for a period of time sufficient to produce said re-differentiated cell phenotype; and
(e) identifying one or more aspects of said re-differentiated cell phenotype in said cell.
9. The method of claim 8 , wherein said cell of step (a) is a bone marrow cell, a spleen cell, or a peripheral blood cell.
10. The method of claim 8 , further comprising restoring Bright/ARID3a function following step (d).
11. The method of claim 8 , wherein said signal is a chemokine.
12. The method of claim 8 , wherein said re-differentiated cell phenotype is a fat cell phenotype, a neuronal cell phenotype, a pancreatic cell phenotype, a hematopoietic cell phenotype, a muscle cell phenotype or an endothelial cell phenotype.
13. The method of claim 8 , wherein said inhibitory nucleic acid is an shRNA.
14. The method of claim 13 , wherein said shRNA is expressed from an expression vector.
15. The method of claim 14 , wherein said expression vector is a viral expression vector.
16. The method of claim 6 , wherein said expression vector is a viral expression vector.