Engineered strains of the oleaginous yeast Yarrowia lipolytica capable of producing greater than 50 weight percent of eicosapentaenoic acid ["EPA"], an .omega.-3 polyunsaturated fatty acid, in the total oil fraction are described. These strains over-express heterologous .DELTA.9 elongases, .DELTA.8 desaturases, .DELTA.5 desaturases, .DELTA.17 desaturases, .DELTA.12 desaturases and C.sub.16/18 elongases, and optionally over-express diacylglycerol cholinephosphotransferases. Preferred gene knockouts are also described. Production host cells, methods for producing EPA within said host cells, and products comprising EPA from the optimized Yarrowia lipolytica strains are claimed.