Increasing Cellular Lipid Production by Increasing the Activity of Diacylglycerol Acyltransferase and Decreasing the Activity of Triacylglycerol Lipase

Abstract

Disclosed are methods and compositions for increasing the triacylglycerol content of a cell by up-regulating diacylglycerol acyltransferase and down-regulating triacylglycerol lipase. In some embodiments, a DGA1 protein is expressed and a native TGL3 gene is knocked out, thereby increasing the synthesis of triacylglycerol and decreasing its consumption, respectively.

1 .- 13 . (canceled) 
     
     
         14 . A method of increasing the lipid content of a cell, comprising transforming a parent cell with a first nucleic acid and a second nucleic acid; wherein said first nucleic acid increases the activity of a native diacylglycerol acyltransferase or comprises a diacylglycerol acyltransferase gene; and said second nucleic acid decreases the activity of a triacylglycerol lipase. 
     
     
         15 . The method of any one of  claim 14 , wherein said first nucleic acid comprises a diacylglycerol acyltransferase gene. 
     
     
         16 . The method of  claim 15 , wherein said diacylglycerol acyltransferase gene encodes a type I diacylglycerol acyltransferase polypeptide, a type II diacylglycerol acyltransferase polypeptide, or a type III diacylglycerol acyltransferase polypeptide. 
     
     
         17 . The method of  claim 15 , wherein said diacylglycerol acyltransferase gene encodes an amino acid sequence selected from the group consisting of: (a) an amino acid sequence set forth in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 41, SEQ ID NO: 43, SEQ ID NO: 45, SEQ ID NO: 47, SEQ ID NO: 49, SEQ ID NO: 51, SEQ ID NO: 53, SEQ ID NO: 55, SEQ ID NO: 57, SEQ ID NO: 59, SEQ ID NO: 61; SEQ ID NO: 29, SEQ ID NO: 31, SEQ ID NO: 33, SEQ ID NO: 35, SEQ ID NO: 37, SEQ ID NO: 39, SEQ ID NO: 63, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 69, SEQ ID NO: 71, SEQ ID NO: 73, SEQ ID NO: 75, SEQ ID NO: 83, or SEQ ID NO: 85, or a biologically-active portion of any one of them; and (b) an amino acid sequence having at least 80% sequence identity to an amino acid sequence forth in SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 5, SEQ ID NO: 7, SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 17, SEQ ID NO: 41, SEQ ID NO: 43, SEQ ID NO: 45, SEQ ID NO: 47, SEQ ID NO: 49, SEQ ID NO: 51, SEQ ID NO: 53, SEQ ID NO: 55, SEQ ID NO: 57, SEQ ID NO: 59, SEQ ID NO: 61; SEQ ID NO: 29, SEQ ID NO: 31, SEQ ID NO: 33, SEQ ID NO: 35, SEQ ID NO: 37, SEQ ID NO: 39, SEQ ID NO: 63, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 69, SEQ ID NO: 71, SEQ ID NO: 73, SEQ ID NO: 75, SEQ ID NO: 83, or SEQ ID NO: 85, or a biologically-active portion of any one of them. 
     
     
         18 . The method of  claim 14 , wherein said second nucleic acid is capable of recombining with a nucleic acid sequence in a triacylglycerol lipase gene and/or a nucleic acid sequence in the regulatory region of a triacylglycerol lipase gene. 
     
     
         19 . The method of  claim 14 , wherein said second nucleic acid is capable of recombining with a nucleic acid sequence in a TGL3, TGL3/4, or TGL4 gene and/or a nucleic acid sequence in the regulatory region of a TGL3, TGL3/4, or TGL4 gene. 
     
     
         20 . The method of  claim 14 , wherein said cell is  Yarrowia lipolytica  or  Arxula adeninivorans.