Transient expression of photosynthetic genes in transfected albinoid petunia protoplasts and correct processing of newly synthesized chloroplast‐destined polypeptides

Abstract

Protoplasts prepared from cultured albinoid cells of petunia do not express photosynthetic genes, such as those coding for chlorophyll a/b‐binding (Cab) proteins or ribulose‐1,5‐bisphosphate carboxylase/oxygenase (Rubisco). They therefore provide a convenient system for expressing recombinant photosynthetic genes, without background interference. Transfection of petunia protoplasts with vectors bearing the Lhcbl*1 Cab gene under the control of the 35S promoter of cauliflower mosaic virus (CaMV) resulted in the appearance of significant amounts of the specific transcripts, but not of the corresponding polypeptides, as inferred from northern and western blot analysis, respectively. The use of an expression vector carrying the translational enhancer Ω of tobacco mosaic virus (TMV) strongly enhanced the appearance of transfected gene products: western blot analysis of transfected protoplasts clearly revealed the appearance of Lemna gibba Lhcbl*1 and Lhcb2*1, tomato Lhcb2*1 and psaD, and pea rbcS gene products. Molecular weight estimations of the newly synthesized polypeptides indicated that each was promptly processed into its mature‐cleaved form within the transfected albinoid protoplasts. This occurred despite a lack of chlorophyll and the absence of a thylakoid network.