MODIFIED MGLUR6 PROMOTER AND METHODS OF USE
The invention provides nucleic acids and nucleic acid expression vectors containing optimized mGluR6 promoters for expression of transgenes in the retina. The compositions and methods of the invention are useful for expression of gene products to preserve, improve, or restore phototransduction or vision.
1 . An adeno-associated virus (AAV) or a recombinant adeno-associated virus (rAAV) expression vector comprising a nucleic acid sequence encoding a capsid protein, a nucleic acid sequence of an mGluR6 promoter comprising SEQ ID NO: 6, or a sequence having at least 70% identity thereto, operably linked to a transgene encoding an opsin selected from the group consisting of a channelrhodopsin, a halorhodopsin, and a melanopsin.
2 . The expression vector of claim 1 , wherein the expression vector comprises an mGluR6 promoter comprising a nucleic acid sequence having at least 80% identity to SEQ ID NO: 6.
3 . The expression vector of claim 1 , wherein the expression vector comprises an mGluR6 promoter comprising a nucleic acid sequence having at least 90% identity to SEQ ID NO: 6.
4 . The expression vector of claim 1 , wherein the expression vector comprises an mGluR6 promoter comprising the nucleic acid sequence of SEQ ID NO: 6.
5 . The expression vector of claim 1 , wherein the vector further comprises an mGluR6 enhancer comprising the nucleic acid sequence of SEQ ID NO: 2, or a sequence having at least 70% identity thereto.
6 . The expression vector of claim 1 , wherein the vector further comprises an mGluR6 enhancer comprising the nucleic acid sequence of SEQ ID NO: 2, or a sequence having at least 80% identity thereto.
7 . The expression vector of claim 1 , wherein the vector further comprises an mGluR6 enhancer comprising the nucleic acid sequence of SEQ ID NO: 2, or a sequence having at least 90% identity thereto.
8 . The expression vector of claim 1 , wherein the channelrhodopsin is selected from the group consisting of Chop1 (SEQ ID NO: 11), Chop2 (SEQ ID NO: 12), Chop2 315 aa fragment (SEQ ID NO: 13), vChR1 (SEQ ID NO: 14), and vChR2 (SEQ ID NO: 15), or an amino acid sequence having at least 90% identity thereto.
9 . The expression vector of claim 1 , wherein the halorhodopsin is NpHR (SEQ ID NO: 16) or eNpHR.
10 . The expression vector of claim 1 , wherein the melanopsin is a human melanopsin (SEQ ID NO: 18), or an amino acid sequence having at least 90% identity thereto.
11 . The expression vector of claim 1 , wherein the vector is a recombinant AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, or AAV12 vector.
12 . The expression vector of claim 11 , wherein the vector is a recombinant AAV2 vector.
13 . The expression vector of claim 1 , wherein the amino acid sequence of the capsid protein is selected from the group consisting of SEQ ID Nos: 23-35.
14 . The expression vector of claim 12 , wherein the capsid protein comprises a peptide selected from the group consisting of SEQ ID NO: 35, SEQ ID NO: 36, and SEQ ID NO: 37.
15 . An adeno-associated virus (AAV) or a recombinant adeno-associated virus (rAAV) expression vector comprising a nucleic acid sequence encoding a capsid protein, a nucleic acid sequence of an mGluR6 promoter comprising SEQ ID NO: 6, or a sequence having at least 70% identity thereto, operably linked to a transgene encoding a human melanopsin (SEQ ID NO: 18), or an amino acid sequence having at least 90% identity thereto.
16 . The expression vector of claim 15 , wherein the vector further comprises an mGluR6 enhancer comprising the nucleic acid sequence of SEQ ID NO: 2, or a sequence having at least 70% identity thereto.
17 . A pharmaceutical composition comprising the expression vector of claim 1 , and a pharmaceutically acceptable carrier or excipient.
18 . The pharmaceutical composition of claim 17 , wherein the composition is formulated for intravitreal delivery.
19 . A pharmaceutical composition comprising the expression vector of claim 15 , and a pharmaceutically acceptable carrier or excipient.
20 . The pharmaceutical composition of claim 19 , wherein the composition is formulated for intravitreal delivery.