Modified form of oleosin that when expressed in plants leads to increased triacylglycerol (oil) accumulation

Patent metadata

Field	Value
Publication number	US-12415840-B2
Application number	US-202117180195-A
Country	US
Kind code	B2
Filing date	Feb 19, 2021
Priority date	Feb 21, 2020
Publication date	Sep 16, 2025
Grant date	Sep 16, 2025

Abstract

The invention provides mutant or variant oleosin polypeptides having one or more amino acid substitutions, particularly one or more arginine substitution for lysine, and having one or more amino acid deletions. The mutant oleosin polypeptides provide for higher triacylglycerol compared to wild type oleosin, including when the mutant oleosin is expressed in plants. Also provided are polynucleotides encoding the mutant oleosin(s), constructs and host cells comprising the polynucleotides, methods for producing oil bodies comprising the mutant oleosin(s) and for producing oil in host cells and plants. The invention also relates to plants, particularly transgenic or recombinantly engineered plants, expressing one or more of the mutant oleosin polypeptides, as well as seeds and oil bodies derived from the plants.

First claim

What is claimed is:

1. A mutant sesame oleosin (OLE) polypeptide comprising an arginine replacement at one or more amino acid residues selected from positions 27, 105, 117, 119 and 128 in SEQ ID NO: 1 or in SEQ ID NO:20, and an amino acid deletion at one or more amino acid residues selected from positions 3, 12, 23, 112, 123 and 136 in SEQ ID NO: 1 or in SEQ ID NO:20.

2. The polypeptide of claim 1 wherein the mutant sesame oleosin polypeptide comprises:
(a) an arginine replacement at one or more lysine residues 27, 105, 117, 119 and 128 in SEQ ID NO:1 or in SEQ ID NO:20 and further comprises an amino acid deletion at one or more of amino acid residue positions 3, 12, 23, 112, 123 and 136 in SEQ ID NO: 1 or in SEQ ID NO:20;
(b) an arginine replacement at lysine residues at positions 27, 105 and 117 in SEQ ID NO: 1 or in SEQ ID NO:20 and further comprises an amino acid deletion at amino acid residue positions 12 and 123 in SEQ ID NO: 1 or in SEQ ID NO:20; or
(c) an arginine replacement at lysine residue 27 in SEQ ID NO: 1 or in SEQ ID NO:20 and further comprises an amino acid deletion at amino acid residue positions 12 and 123 in SEQ ID NO: 1 or in SEQ ID NO:20.

3. The polypeptide of claim 1 wherein the mutant sesame oleosin has an amino acid sequence as set out in any one of SEQ ID NOs: 2 or 4-19.

4. The polypeptide of claim 3 wherein the mutant oleosin has an amino acid sequence as set out in SEQ ID NO: 6, 7 or 8.

5. The polypeptide of claim 1 wherein expression of the mutant oleosin in a host cell or host plant results in increased levels or accumulation of oleosin in the cell or plant.

6. The polypeptide of claim 1 wherein expression of the mutant oleosin in a host cell or host plant results in increased levels or accumulation of triacylglycerol (TAG) in the cell or plant.

7. An isolated nucleic acid encoding the polypeptide of claim 1 .

8. A recombinant vector comprising the nucleic acid of claim 7 .

9. A host cell comprising the vector of claim 8 .

10. A host cell recombinantly engineered or genetically modified to produce the mutant oleosin polypeptide of claim 1 .

11. A host cell recombinantly engineered to overproduce oleosin and/or TAG by introducing nucleic acid encoding the polypeptide of claim 1 , wherein the engineered host cell produces more oleosin and/or TAG than a wild type/native host cell.

12. The host cell of claim 11 which is further engineered to express a triacylglycerol (TAG) synthesizing enzyme.

13. The host cell of claim 11 which is further engineered to express one or more of diacylglycerol acyltransferase (DGAT), wrinkled1 (WRL1) or medium chain thioesterase (MCT or T).

14. The host cell of claim 10 wherein the cell is a plant cell.

15. The host cell of claim 10 that produces triacylglycerol.

16. The host cell of claim 10 , wherein the host cell is a yeast cell, fungal cell, an animal cell or a plant cell.

17. A host plant comprising a recombinant vector having a nucleic acid encoding a mutant sesame oleosin (OLE) polypeptide, wherein the mutant oleosin has an arginine replacement at one or more amino acid residues selected from positions 27, 105, 117, 119 and 128 in SEQ ID NO: 1 or in SEQ ID NO:20 and further has an amino acid deletion at one or more amino acid residues selected from positions 3, 12, 23, 112, 123 and 136 in SEQ ID NO: 1 or in SEQ ID NO:20.

18. A host plant recombinantly engineered to produce the polypeptide of claim 17 .

19. The host plant of claim 17 wherein the plant is a bioenergy crop that is sesame, sugar cane or sorghum.

20. A method for producing an oil body in a host cell, the method comprising: a) engineering the host cell to produce a mutant oleosin of claim 1 ; and b) culturing the host cell in order to express the mutant oleosin.

21. The method of claim 20 , wherein the host cell is engineered in (a) by introducing into the host cell at least one nucleic acid of claim 7 .

22. The method of claim 21 , further comprising introducing into the host cell in a) a nucleic acid molecule encoding a TAG synthesizing enzyme; and b) culturing the host cell in order to express the modified oleosin and the TAG synthesizing enzyme.

23. The host cell of claim 10 , wherein the mutant oleosin polypeptide has an amino acid sequence as set out in SEQ ID NO: 6, 7 or 8.

24. The host cell of claim 10 , recombinantly engineered or genetically modified to produce a combination of mutant oleosin polypeptides, wherein the mutant oleosin polypeptides have the amino acid sequence of SEQ ID NO: 6 and of SEQ ID NO: 7.

25. The host cell of claim 11 , wherein the mutant oleosin polypeptide has an amino acid sequence as set out in SEQ ID NO: 6, 7 or 8.

26. The host plant of claim 17 , wherein the mutant oleosin has an amino acid sequence as set out in SEQ ID NO: 6, 7 or 8.

27. The host plant of claim 17 comprising a recombinant vector encoding a combination of mutant sesame oleosin (OLE) polypeptides, wherein the mutant oleosin polypeptides have the amino acid sequence of SEQ ID NO: 6 and of SEQ ID NO: 7.

28. The method of claim 20 , wherein the mutant oleosin has an amino acid sequence as set out in SEQ ID NO: 6, 7 or 8.

29. The method of claim 20 , wherein the host cell is engineered to produce a combination of mutant oleosins of claim 1 and wherein the mutant oleosins have the amino acid sequence of SEQ ID NO: 6 and of SEQ ID NO: 7.

Assignees

• Brookhaven Science Ass Llc

Inventors

• Shanklin John
• Anaokar Sanket P

Classifications

• C12N15/8247

  involving modified lipid metabolism, e.g. seed oil composition · CPC title

• C07K14/415 Primary

  from plants · CPC title