Compositions for enhancing the pluripotency of stem cells
Described herein is the finding that increasing the frequency of Zscan4 activation in mouse ES cells not only enhances, but also maintains their developmental potency in long-term cell culture. Particularly disclosed herein is the finding that the constitutive presence of Zscan4-ERT2, even in the absence of its usual activator tamoxifen, can increase the frequency of endogenous Zscan4 activation in ES cells, resulting in the increase of developmental potency of the ES cells. Accordingly, provided herein are Zscan4-ERT2 fusion proteins and nucleic acid molecules and vectors encoding Zscan4-ERT2 fusion proteins. Further provided are methods of prolonging and/or enhancing stem cell plmipotency using the disclosed Zscan4-ERT2 nucleic acid molecules and fusion proteins.
1. An expression vector comprising a nucleic acid molecule encoding a Zscan4-ERT2 fusion protein operably linked to a promoter, wherein the Zscan4-ERT2 fusion protein consists of the amino acid sequence of SEQ ID NO: 22 or SEQ ID NO: 23.
2. An isolated cell comprising the expression vector of claim 1 .
3. The isolated cell of claim 2 , which is a stem cell.
4. The isolated cell of claim 3 , wherein the stem cell is an embryonic stem (ES) cell or an induced pluripotent stem (iPS) cell.
5. The isolated cell of claim 3 , wherein the stem cell is a mouse, rat, human or non-human primate stem cell.
6. A composition comprising the expression vector of claim 1 and a pharmaceutically acceptable carrier.
7. The expression vector of claim 1 , wherein the Zscan4-ERT2 fusion protein consists of the amino acid sequence of SEQ ID NO: 23.
8. The expression vector of claim 1 , wherein the Zscan4-ERT2 fusion protein consists of the amino acid sequence of SEQ ID NO: 22.
9. An expression vector comprising a nucleic acid molecule encoding a mutant Zscan4-AC protein operably linked to a promoter, wherein the mutant Zscan4-AC protein consists of the amino acid residues (i) 1-311 of SEQ ID NO:2, (ii) 1-339 of SEQ ID NO: 2, (iii) 1-367 of SEQ ID NO: 2, (iv) 1-395 of SEQ ID NO: 2, (v) 1-394 of SEQ ID NO: 8, (vi) 1-423 of SEQ ID NO: 8, (vii) 1-451 of SEQ ID NO: 8, or (viii) 1-479 of SEQ ID NO: 8.
10. An isolated cell comprising the expression vector of claim 9 .
11. The isolated cell of claim 10 , which is a stem cell.
12. A composition comprising the expression vector of claim 9 and a pharmaceutically acceptable carrier.
13. The expression vector of claim 9 , wherein the mutant Zscan-ΔC protein consists of the amino acid sequence of SEQ ID NO: 25.
14. The expression vector of claim 9 , wherein the mutant Zscan4-AC protein consists of the amino acid residues (i) 1-311 of SEQ ID NO:2, (ii) 1-339 of SEQ ID NO: 2, (iii) 1-367 of SEQ ID NO: 2, or (iv) 1-395 of SEQ ID NO: 2.
15. The expression vector of claim 9 , wherein the mutant Zscan4-AC protein consists of the amino acid residues (v) 1-394 of SEQ ID NO: 8, (vi) 1-423 of SEQ ID NO: 8, (vii) 1-451 of SEQ ID NO: 8, or (viii) 1-479 of SEQ ID NO: 8.