METHODS AND COMPOSITIONS FOR MODULATING GENE EXPRESSION
The present disclosure provides compositions with a modulating gene expression and methods for modulating transcription
1 .- 21 . (canceled)
22 . A method of decreasing expression of a gene within a cell, the gene being within an anchor sequence-mediated conjunction that comprises a first anchor sequence and a second anchor sequence, the method comprising a step of:
contacting the cell with a composition comprising a targeting element or a nucleic acid encoding the targeting element, wherein the targeting element comprises a TAL array which targets the first anchor sequence, the second anchor sequence, or both of the first anchor sequence and the second anchor sequence,
wherein the first and second anchor sequences each comprise a CTCF binding site.
23 . The method of claim 22 , wherein the CTCF binding site has the sequence of SEQ ID NO: 1 or SEQ ID NO: 2.
24 . The method of claim 22 , wherein the anchor sequence-mediated conjunction further comprises an internal enhancing sequence.
25 . The method of claim 22 , wherein neither the first nor second anchor sequences are located within an internal enhancing sequence or a promoter.
26 . The method of claim 22 , wherein the gene is FOXJ3, MYC, SHMT2, or CDK6.
27 . The method of claim 22 , wherein the cell is a human cell.
28 . The method of claim 22 , wherein contacting comprises transfecting the cell with the composition.
29 . The method of claim 22 , wherein the composition physically interferes with formation and/or maintenance of the anchor sequence-mediated conjunction.
30 . The method of claim 22 , wherein the gene is an oncogene, a tumor suppressor, or a gene associated with a nucleotide repeat.
31 . The method of claim 22 , wherein the targeting element is linked to an effector domain with an amino acid linker.
32 . The method of claim 31 , wherein the effector domain comprises a DNA methyltransferase, histone methyltransferase, acetyltransferase, or deacetylase.
33 . The method of claim 31 , wherein the effector domain comprises a KRAB domain, or wherein the effector domain promotes DNA methylation.
34 . The method of claim 22 , wherein the gene is separated from the first anchor sequence or the second anchor sequence by about 100 bp to about 500 Mb.
35 . The method of claim 22 , wherein the anchor sequences are between 20-80 nucleotides.
36 . The method of claim 23 , wherein the targeting element binds the sequence of SEQ ID NO: 1 or SEQ ID NO: 2.
37 . The method of claim 23 , wherein the targeting element binds a sequence proximal to SEQ ID NO: 1 or SEQ ID NO: 2.
38 . The method of claim 22 , which comprises contacting the cell with the nucleic acid encoding the targeting element.
39 . The method of claim 22 , wherein the nucleic acid comprises mRNA.
40 . A composition comprising a targeting element or a nucleic acid encoding the targeting element,
wherein the targeting element comprises a Cas protein and gRNA which bind specifically to a first anchor sequence, a second anchor sequence, or both of the first anchor sequence and the second anchor sequence, wherein the first and second anchor sequences each comprise a CTCF binding site, wherein the CTCF binding site has the sequence of SEQ ID NO: 1 or SEQ ID NO: 2, and
wherein the first anchor sequence, the second anchor sequence, and a gene are comprised in an anchor sequence-mediated conjunction.
41 . A composition comprising a targeting element or a nucleic acid encoding the targeting element,
wherein the targeting element comprises a zinc finger array which targets the first anchor sequence, the second anchor sequence, or both of the first anchor sequence and the second anchor sequence, wherein the first and second anchor sequences each comprise a CTCF binding site, and wherein the CTCF binding site has the sequence of SEQ ID NO: 1 or SEQ ID NO: 2,
wherein the first anchor sequence, the second anchor sequence, and a gene are comprised in an anchor sequence-mediated conjunction.