Non-human animals comprising a humanized ACE2 locus
Non-human animal cells and non-human animals comprising a humanized ACE2 locus and methods of using such non-human animal cells and non-human animals are provided. Non-human animal cells or non-human animals comprising a humanized ACE2 locus express a human ACE2 protein or a chimeric ACE2 protein, fragments of which are from human ACE2. Methods are also provided for using such non-human animals comprising a humanized ACE2 locus to assess in vivo ACE2 activity, e.g., coronavirus infection and/or the treatment or prevention thereof.
1 . A rodent comprising a genetically modified endogenous ACE2 locus encoding a recombinant ACE2 protein that comprises in operable linkage:
(i) an ACE2 signal sequence of an endogenous ACE2 protein;
(ii) an extracellular domain of a human ACE2 protein;
(iii) a transmembrane domain of the endogenous ACE2 protein; and
(iv) a cytoplasmic domain of the endogenous ACE2 protein;
wherein the rodent expresses the recombinant ACE2 protein.
2 . The rodent of claim 1 , wherein the genetically modified endogenous ACE2 locus comprises a replacement of a nucleotide sequence encoding the extracellular domain of the endogenous ACE2 protein with a nucleotide sequence encoding the extracellular domain of the human ACE2 protein such that the nucleotide sequence encoding the extracellular domain of the human ACE2 protein is operably linked to an endogenous nucleotide sequence encoding (iii) the transmembrane domain of the endogenous ACE2 protein, and (iv) the cytoplasmic domain of the endogenous ACE2 protein.
3 . The rodent of claim 1 , wherein the genetically modified endogenous ACE2 locus comprises the nucleotide sequence as set forth in SEQ ID NO: 5 or as set forth in SEQ ID NO:22.
4 . The rodent of claim 1 , further comprising the sequence as set forth in SEQ ID NO:5, the sequence as set forth in SEQ ID NO:22, or the sequence as set forth in SEQ ID NO:25.
5 . The rodent of claim 1 , wherein the amino acid sequence of (ii) the extracellular domain of the human ACE2 protein comprises the amino acid sequence as set forth in SEQ ID NO:27.
6 . The rodent of claim 1 , wherein the rodent is heterozygous or homozygous for the genetically modified endogenous ACE2 locus.
7 . The rodent of claim 1 , wherein the rodent is a rat or a mouse.
8 . The rodent of claim 7 ,
wherein the rodent is a mouse, and
wherein the recombinant ACE2 protein comprises in operable linkage:
(i) an ACE2 signal sequence of an endogenous mouse ACE2 protein;
(ii) an extracellular domain of a human ACE2 protein;
(iii) a transmembrane domain of the endogenous mouse ACE2 protein; and
(iv) a cytoplasmic domain of the endogenous mouse ACE2 protein.
9 . The rodent of claim 8 , wherein the amino acid sequence of the recombinant ACE2 protein comprises the amino acid sequence as set forth in SEQ ID NO:24.
10 . A rodent cell comprising:
a genetically modified endogenous ACE2 locus encoding a recombinant ACE2 protein that comprises in operable linkage:
(i) an ACE2 signal sequence of an endogenous ACE2 protein;
(ii) an extracellular domain of a human ACE2 protein;
(iii) a transmembrane domain of the endogenous ACE2; and
(iv) a cytoplasmic domain of the endogenous ACE2 protein;
wherein the rodent cell is:
(a) an embryonic stem(ES) cell, a pluripotent cell, or a germ cell; or
(b) a somatic cell that expresses the recombinant ACE2 protein.
11 . A rodent tissue comprising the rodent cell of claim 10 .
12 . The rodent of claim 1 , wherein the rodent comprises in its genome a genetic modification of the endogenous ACE2 locus encoding the recombinant ACE2 protein that comprises in operable linkage:
(i) an ACE2 signal sequence of a rodent ACE2 protein set forth in SEQ ID NO: 26;
(ii) an extracellular domain of a human ACE2 protein set forth in SEQ ID NO: 27;
(iii) a transmembrane domain of a rodent ACE2 protein set forth in SEQ ID NO: 28; and
(iv) a cytoplasmic domain of a rodent ACE2 protein set forth in SEQ ID NO: 29.
13 . The rodent cell of claim 10 , comprising a genetically modified endogenous ACE2 locus encoding a recombinant ACE2 protein that comprises in operable linkage:
(i) an ACE2 signal sequence of a rodent ACE2 protein set forth in SEQ ID NO: 26;
(ii) an extracellular domain of a human ACE2 protein set forth in SEQ ID NO: 27;
(iii) a transmembrane domain of a rodent ACE2 protein set forth in SEQ ID NO: 28; and
(iv) a cytoplasmic domain of a rodent ACE2 protein set forth in SEQ ID NO: 29.
14 . The rodent cell of claim 10 , wherein the rodent cell is an alveolar cell of the lung, an esophagus upper and stratified epithelial cell, an absorptive enterocyte from the ileum or colon, a cholangiocyte, a myocardial cell, a kidney proximal tubule cell, or a bladder urothelial cell.