Skip to main content

Identification of a cDNA that encodes a 1-acyl-sn-glycerol-3-phosphate acyltransferase from Limnanthes douglasii


Two different techniques were used to isolate potential cDNAs for acyl-CoA: 1-acyl-sn-glycerol-3-phosphate acyltransferase (LPA-AT) enzymes from Limnanthes douglasii. Both heterologous screening with the maize pMAT1 clone and in vivo complementation of the Escherchia coli mutant JC201 which is deficient in LPA-AT activity, were carried out. Clones identified by these procedures were different. Homology searches demonstrated that the clone isolated by heterologous probing, pLAT1, encodes a protein which is most similar to the maize (open reading frame in pMAT1) and yeast SLC1 proteins, which are putative LPA-AT sequences. This L. douglasii sequence shows much lower homology to the E. coli LPA-AT protein PlsC, which is the only LPA-AT sequence confirmed by over-expression studies. The clone isolated by complementation, pLAT2, encodes a protein with homology to both SLC1 and PlsC. It was not possible to over-express the complementing protein encoded by pLAT2 but further experimentation on membranes from complemented JC201 demonstrated that they possess a substrate specificity distinctly different from PlsC and similar to Limnanthes sp. microsome specificity. This data strongly supports the contention that pLAT2 is an LPA-AT clone. Northern blot analysis revealed different expression patterns for the two genes in pLAT1 and pLAT2. Transcription of the gene encoding the insert of pLAT2 occurred almost exclusively in developing seed tissue, whilst the cDNA of pLAT1 hybridised to poly(A)+ mRNA from seed, stem and leaf, demonstrating more widespread expression throughout the plant. Southern blot analysis indicated that the cDNA of pLAT2 was transcribed from a single-copy gene while that for pLAT1 was a member of a small gene family.

This is a preview of subscription content, access via your institution.


  1. 1.

    Bernerth R, Frentzen M: Utilisation of erucoyl-CoA by acyltransferases from developing seeds of Brassica napus (L.) involved in triacylglycerol biosynthesis. Plant Sci 67: 21–28 (1990).

    Google Scholar 

  2. 2.

    Brown AP, Coleman C, Tommey AM, Watson MD, Slabas AR: Isolation and characterisation of a maize cDNA that complements a 1-acyl-sn-glycerol-3-phosphate acyl-transferase mutant of Escherichia coli and encodes a protein which has similarities to other acyltransferases. Plant Mol Biol 26: 211–223 (1994).

    Google Scholar 

  3. 3.

    Browse J. Somerville C: Glycerolipid synthesis: biochemistry and regulation. Annu Rev Plant Physiol Plant Mol Biol 42: 467–506 (1991).

    Google Scholar 

  4. 4.

    Bullock WO, Fernandez JM, Short JM: XL1-Blue: a high efficiency plasmid transforming recA Escherichia coli strain with beta-galactosidase selection. Bio Techniques 5: 376–379 (1987).

    Google Scholar 

  5. 5.

    Cao Y-Z, Oo K-C, Huang AHC: Lysophosphatidate acyltransferase in microsomes from maturing seeds of meadowfoam (Limnanthes alba). Plant Physiol 94: 1199–1206 (1990).

    Google Scholar 

  6. 6.

    Coleman J: Characterization of the Escherichia coli gene for 1-acyl-sn-glycerol-3-phosphate acyltransferase (plsC). Mol Gen Genet 232: 295–303 (1992).

    Google Scholar 

  7. 7.

    Coleman J: Characterization of Escherichia coli cells deficient in 1-acyl-sn-glycerol-3-phosphate acyltransferase activity. J Biol Chem 265: 17215–17221 (1990).

    Google Scholar 

  8. 8.

    Delauney AJ, Verma DPS: Isolation of plant genes by heterologous complementation in Escherichia coli. In: Gelvin SB, Schilperoort RA, Verma DPS (eds) Plant Molecular Biology Manual A14: 1–23 Kluwer Academic Publishers. Dordrecht (1990).

    Google Scholar 

  9. 9.

    Draper J. Scott R: The isolation of plant nucleic acids. In: Draper J, Scott R, Armitage P, Walden R (eds) Plant Genetic Transformation and Gene Expression: A Laboratory Manual, pp. 201–236. Blackwell Scientific Publications, Oxford, UK (1988).

    Google Scholar 

  10. 10.

    Feinberg AP, Vogelstein B: A technique for radio-labelling DNA restriction fragments to high specific activity. Anal Biochem 132: 6–13 (1983).

    Google Scholar 

  11. 11.

    Hall TC, Ma Y, Buchbinder BU, Payne JW, Sun SN, Bliss FA: Messenger RNA for G1 protein of French bean seeds: cell free translation and product characterisation. Proc Natl Acad Sci USA 75: 3196 (1978).

    Google Scholar 

  12. 12.

    Higgins DG, Bleasby AJ, Fuchs R: Clustal-V: improved software for multiple sequence alignment. Comput Applic Biosci 8: 189–191 (1992).

    Google Scholar 

  13. 13.

    Icho T: Membrane-bound phosphatases in Escherichia coli: sequence of the pgpB gene and dual subcellular localization of the pgpB product. J Bact 170: 5117–5124 (1988).

    Google Scholar 

  14. 14.

    Kater MM, Koningstein GM, Nijkamp JJ, Stuitje AR: cDNA cloning and expression of Brassica napus enoylacyl carrier protein reductase in Escherichia coli. Plant Mol Biol 17: 895–909 (1991).

    Google Scholar 

  15. 15.

    Keegstra K, Olsen LJ, Theg SM: Chloroplastic precursors and their transport across the envelope membranes. Annu Rev Plant Physiol Plant Mol Biol 40: 471–501 (1989).

    Google Scholar 

  16. 16.

    Laurent P, Huang AHC: Organ- and development-specific acyl coenzyme A lysophosphatidate acyltransferases in palm and meadowfoam. Plant Physiol 99: 1711–1715 (1992).

    Google Scholar 

  17. 17.

    Lipman DJ, Pearson WR: Rapid and sensitive protein similarity searches. Science 227: 1435–1441 (1985).

    Google Scholar 

  18. 18.

    Löhden I, Frentzen M: Triacylclycerol biosynthesis in developing seeds of Tropaeolum majus L. and Limnanthes douglasii R. Br. Planta 188: 215–224 (1992).

    Google Scholar 

  19. 19.

    Marck C. DNA Strider a ‘C’ program for the fast analysis of DNA and protein sequences on the Apple Macintosh family of computers. Nucl Acids Res 16: 1829–1836 (1988).

    Google Scholar 

  20. 20.

    Marek Nagiec M, Wells GB, Lester RL, Dickson RC: A suppressor gene that enables Saccharomyces cerevisiae to grow without making sphingolipids encodes a protein that resembles an Escherichia coli fatty acyltransferase. J Biol Chem 268: 22156–22163 (1993).

    Google Scholar 

  21. 21.

    Nishida I, Tasaka Y, Shirashi H, Murata N: The gene and the RNA for the precursor to the plastid-located glycerol-3-phosphate acyltransferase of Arabidopsis thaliana. Plant Mol Biol 21: 267–277 (1993).

    Google Scholar 

  22. 22.

    Program manual for Wisconsin package, Version 8, Genetics computer group, 575 Science Drive, Madison, WI 3711 (1994).

  23. 23.

    Sambrook J, Fritsch EF. Maniatis T: Molecular Cloning: A Laboratory Manual (2nd ed.). Cold Spring Harbor Laboratory Press. Cold Spring Harbor, NY (1989).

    Google Scholar 

  24. 24.

    Smith T. Waterman MS: Identification of common molecular subsequences. J Mol Biol 147: 195–197 (1981).

    Google Scholar 

  25. 25.

    Taylor DC, MacKenzie SL. McCurdy AR, McVetty PBE, Giblin EM, Pass EW, Stone SJ, Scarth R, Rimmer SR, Pickard MD: Stereospecific analyses of seed triacylglycerols from high-erucic acid Brassicaceae: detection of erucic acid at the sn-2 position in Brassica oleracea L. genotypes. J Am Oil Chem Soc. 71: 163–167 (1994).

    Google Scholar 

  26. 26.

    von Heijne G: Signals for protein import into organelles. In: Herrmann RG. Larkins B (eds) Plant Molecular Biology vol. 2, pp. 583–593. Plenum Press. New York (1991).

    Google Scholar 

Download references

Author information



Rights and permissions

Reprints and Permissions

About this article

Cite this article

Brown, A.P., Brough, C.L., Kroon, J.T.M. et al. Identification of a cDNA that encodes a 1-acyl-sn-glycerol-3-phosphate acyltransferase from Limnanthes douglasii . Plant Mol Biol 29, 267–278 (1995).

Download citation

Key words

  • 1-acyl-sn-glycerol-3-phosphate acyltransferase
  • complementation cloning
  • Limnanthes douglasii