CBLB endonuclease variants, compositions, and methods of use
The present disclosure provides improved genome editing compositions and methods for editing a CBLB gene. The disclosure further provides genome edited cells for the prevention, treatment, or amelioration of at least one symptom of, a cancer, an infectious disease, an autoimmune disease, an inflammatory disease, or an immunodeficiency.
1. A polynucleotide encoding a polypeptide comprising an I-OnuI homing endonuclease (HE) variant that comprises an amino acid sequence that is at least 90% identical to SEQ ID NO: 6 and that binds and cleaves a target site in the human casitas B-lineage (Cbl) lymphoma proto-oncogene B (CBLB) gene, wherein the target site is SEQ ID NO: 20.
2. The polynucleotide of claim 1 , wherein the I-OnuI HE variant comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 6.
3. The polynucleotide of claim 1 , wherein the I-OnuI HE variant comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 6.
4. The polynucleotide of claim 1 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 6.
5. The polynucleotide of claim 1 , wherein the polypeptide further comprises a TALE DNA binding domain, wherein the polypeptide binds and cleaves a target site in the human CBLB gene, wherein the target site is SEQ ID NO: 22.
6. The polynucleotide of claim 5 , wherein the TALE DNA binding domain comprises about 9.5 TALE repeat units to about 15.5 TALE repeat units.
7. The polynucleotide of claim 5 , wherein the TALE DNA binding domain comprises about 12.5 TALE repeat units.
8. The polynucleotide of claim 5 , wherein the TALE DNA binding domain comprises 12.5 TALE repeat units.
9. The polynucleotide of claim 5 , wherein the TALE DNA binding domain binds the polynucleotide sequence set forth in SEQ ID NO: 21.
10. The polynucleotide of claim 5 , wherein the I-OnuI HE variant comprises an amino acid sequence that is at least 95% identical to SEQ ID NO: 6.
11. The polynucleotide of claim 5 , wherein the I-OnuI HE variant comprises an amino acid sequence that is at least 99% identical to SEQ ID NO: 6.
12. The polynucleotide of claim 5 , wherein the I-OnuI HE variant comprises the amino acid sequence set forth in SEQ ID NO: 6.
13. The polynucleotide of claim 5 , wherein the polynucleotide is a cDNA.
14. The polynucleotide of claim 5 , wherein the polynucleotide is an mRNA.
15. The polynucleotide of claim 14 , wherein the mRNA the polynucleotide sequence set forth in SEQ ID NO 31.