J Integr Plant Biol ›› 2011, Vol. 53 ›› Issue (11): 858-868.DOI: 10.1111/j.1744-7909.2011.01077.x

• Metabolism and Biochemistry • Previous Articles     Next Articles

Localisation of Seed Oil Body Proteins in Tobacco Protoplasts Reveals Specific Mechanisms of Protein Targeting to Leaf Lipid Droplets

Stefania De Domenico1, Stefania Bonsegna1, Marcello Salvatore Lenucci2, Palmiro Poltronieri1, Gian Pietro Di Sansebastiano2 and Angelo Santino1*   

  1. 1Institute of Sciences of Food Production C.N.R. Unit of Lecce, via Monteroni, Lecce 73100, Italy
    2Dipartimento di Scienze e Tecnologie Biologiche ed Ambientali, Università del Salento, via Monteroni, Lecce 73100, Italy
  • Received:2011-08-11 Accepted:2011-09-15 Published:2011-09-26
  • About author:*Corresponding author Tel: +39 08 3242 2606; Fax: +39 08 3242 2620; E-mail: angelo.santino@ispa.cnr.it

Abstract:

Oleosin, caleosin and steroleosin are normally expressed in developing seed cells and are targeted to oil bodies. In the present work, the cDNA of each gene tagged with fluorescent proteins was transiently expressed into tobacco protoplasts and the fluorescent patterns observed by confocal laser scanning microscopy. Our results indicated clear differences in the endocellular localization of the three proteins. Oleosin and caleosin both share a common structure consisting of a central hydrophobic domain flanked by two hydrophilic domains and were correctly targeted to lipid droplets (LD), whereas steroleosin, characterized by an N-terminal oil body anchoring domain, was mainly retained in the endoplasmic reticulum (ER). Protoplast fractionation on sucrose gradients indicated that both oleosin and caleosin-green fluorescent protein (GFP) peaked at different fractions than where steroleosin-GFP or the ER marker binding immunoglobulin protein (BiP), were recovered. Chemical analysis confirmed the presence of triacylglycerols in one of the fractions where oleosin-GFP was recovered. Finally, only oleosin- and caleosin-GFP were able to reconstitute artificial oil bodies in the presence of triacylglycerols and phospholipids. Taken together, our results pointed out for the first time that leaf LDs can be separated by the ER and both oleosin or caleosin are selectively targeted due to the existence of selective mechanisms controlling protein association with these organelles.

De Domenico S, Bonsegna S, Lenucci MS, Poltronieri P, Di Sansebastiano GP, Santino A (2011) Localization of seed oil body proteins in tobacco protoplasts reveals specific mechanisms of protein targeting to leaf lipid droplets. J. Integr. Plant Biol. 53(11), 858–868.

Key words: artificial oil bodies, caleosin, endoplasmic reticulum, lipid droplets, oleosin, protein sorting, steroleosin

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