Chimeric Retinoid X Receptors and Their Use in a Novel Ecdysone Receptor-Based Inducible Gene Expression System

Patent metadata

Field	Value
Publication number	US-2017096673-A1
Application number	US-201615293602-A
Country	US
Kind code	A1
Filing date	Oct 14, 2016
Priority date	Feb 20, 2001
Publication date	Apr 6, 2017
Grant date	—

Abstract

This invention relates to the field of biotechnology or genetic engineering. Specifically, this invention relates to the field of gene expression. More specifically, this invention relates to a novel ecdysone receptor/chimeric retinoid X receptor-based inducible gene expression system and methods of modulating gene expression in a host cell for applications such as gene therapy, large-scale production of proteins and antibodies, cell-based high throughput screening assays, functional genomics and regulation of traits in transgenic organisms.

First claim

1 .- 35 . (canceled)

36 . An isolated polynucleotide selected from the group consisting of:
(a) a polynucleotide encoding a truncated chimeric retinoid X receptor ligand binding domain comprising a truncation mutation, wherein the truncation mutation reduces ligand binding activity of the truncated chimeric retinoid X receptor ligand binding domain; (b) a polynucleotide encoding a truncated chimeric retinoid X receptor ligand binding domain comprising a truncation mutation, wherein the truncation mutation reduces steroid binding activity of the truncated chimeric retinoid X receptor ligand binding domain; (c) a polynucleotide encoding a truncated chimeric retinoid X receptor ligand binding domain comprising a truncation mutation, wherein the truncation mutation reduces non-steroid binding activity of the truncated chimeric retinoid X receptor ligand binding domain; (d) a polynucleotide encoding a truncated chimeric retinoid X receptor ligand binding domain comprising a truncation mutation, wherein the truncation mutation enhances ligand binding activity of the truncated chimeric retinoid X receptor ligand binding domain; (e) a polynucleotide encoding a truncated chimeric retinoid X receptor ligand binding domain comprising a truncation mutation, wherein the truncation mutation enhances steroid binding activity of the truncated chimeric retinoid X receptor ligand binding domain; (f) a polynucleotide encoding a truncated chimeric retinoid X receptor ligand binding domain comprising truncation mutation, wherein the truncation mutation enhances non-steroid binding activity of the truncated chimeric retinoid X receptor ligand binding domain; (g) a polynucleotide encoding a truncated chimeric retinoid X receptor ligand binding domain comprising a truncation mutation, wherein the truncation mutation increases ligand sensitivity of the truncated chimeric retinoid X receptor ligand binding domain; and (h) a polynucleotide encoding a truncated chimeric retinoid X receptor ligand binding domain comprising a truncation mutation, wherein the truncation mutation increases ligand sensitivity of a heterodimer wherein the heterodimer comprises the truncated chimeric retinoid X receptor ligand binding domain and a dimerization partner.

37 .- 44 . (canceled)

45 . An isolated polynucleotide encoding a chimeric retinoid X receptor ligand binding domain, wherein the polynucleotide comprises a nucleic acid sequence selected from the group consisting of
(a) nucleotides 1-348 of SEQ ID NO: 13 and nucleotides 268-630 of SEQ ID NO: 21, (b) nucleotides 1-408 of SEQ ID NO: 13 and nucleotides 337-630 of SEQ ID NO: 21, (c) nucleotides 1-465 of SEQ ID NO: 13 and nucleotides 403-630 of SEQ ID NO: 21, (d) nucleotides 1-555 of SEQ ID NO: 13 and nucleotides 490-630 of SEQ ID NO 21, (e) nucleotides 1-624 of SEQ ID NO 13 and nucleotides 547-630 of SEQ ID NO 21, (f) nucleotides 1-645 of SEQ ID NO: 13 and nucleotides 601-630 of SEQ ID NO: 21, and (g) nucleotides 1-717 of SEQ ID NO: 13 and nucleotides 613-630 of SEQ ID NO: 21.

46 . (canceled)

47 . An isolated polynucleotide encoding a chimeric retinoid X receptor ligand binding domain comprising an amino acid sequence selected from the group consisting of
(a) amino acids 1-116 of SEQ ID NO: 19 and amino acids 90-210 of SEQ ID NO: 27, (b) amino acids 1-136 of SEQ ID NO: 19 and amino acids 113-210 of SEQ ID NO: 27, (c) amino acids 1-155 of SEQ ID NO: 19 and amino acids 135-210 of SEQ ID NO: 27, (d) amino acids 1-185 of SEQ ID NO: 19 and amino acids 164-210 of SEQ ID NO: 27, (e) amino acids 1-208 of SEQ ID NO: 19 and amino acids 183-210 of SEQ ID NO: 27, (f) amino acids 1-215 of SEQ ID NO: 19 and amino acids 201-210 of SEQ ID NO: 27, and (g) amino acids 1-239 of SEQ ID NO 19 and amino acids 205-210 of SEQ ID NO 27.

48 - 65 . (canceled)

66 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-116 of SEQ ID NO 19 and amino acids 90-210 of SEQ ID NO 27.

67 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-136 of SEQ ID NO: 19 and amino acids 113-210 of SEQ ID NO: 27.

68 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-155 of SEQ ID NO 19 and amino acids 135-210 of SEQ ID NO: 27.

69 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-185 of SEQ ID NO: 19 and amino acids 164-210 of SEQ ID NO: 27.

70 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-208 of SEQ ID NO: 19 and amino acids 183-210 of SEQ ID NO: 27.

71 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-215 of SEQ ID NO: 19 and amino acids 201-210 of SEQ ID NO: 27.

72 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-208 of SEQ ID NO: 19 and amino acids 183-210 of SEQ ID NO: 27.

73 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-215 of SEQ ID NO: 19 and amino acids 201-210 of SEQ ID NO: 27.

74 . The isolated polynucleotide of claim 47 , wherein the polynucleotide encodes a chimeric retinoid X receptor ligand binding domain comprising amino acids 1-239 of SEQ ID NO: 19 and amino acids 205-210 of SEQ ID NO: 27.

Assignees

• Intrexon Corp

Inventors

• Kapitskaya Marianna Zinovievna
• Palli Subba Reddy

Classifications

• C07K14/70567

  Nuclear receptors, e.g. retinoic acid receptor [RAR], RXR, nuclear orphan receptors · CPC title

• C07K14/721

  Steroid/thyroid hormone superfamily, e.g. GR, EcR, androgen receptor, oestrogen receptor · CPC title

• C12N15/635 Primary

  Externally inducible repressor mediated regulation of gene expression, e.g. tetR inducible by tetracyline · CPC title

• C07K2319/70

  containing domain for protein-protein interaction · CPC title

• C12N15/1055

  Protein x Protein interaction, e.g. two hybrid selection · CPC title