Transit Peptide Mutations That Impair in Vitro and in Vivo Chloroplast Protein Import Do Not Affect Accumulation of the -Subunit of Chloroplast ATPase¹

Karen L. Kindle^* and Susan D. Lawrence²

Plant Science Center, Biotechnology Program, 151 Biotechnology Building, Cornell University, Ithaca, New York 14853

We have begun to take a genetic approach to study chloroplast protein import in Chlamydomonas reinhardtii by creating deletions in the transit peptide of the -subunit of chloroplast ATPase-coupling factor 1 (CF₁-, encoded by AtpC) and testing their effects in vivo by transforming the altered genes into an atpC mutant, and in vitro by importing mutant precursors into isolated C. reinhardtii chloroplasts. Deletions that removed 20 or 23 amino acid residues from the center of the transit peptide reduced in vitro import to an undetectable level but did not affect CF₁- accumulation in vivo. The CF₁- transit peptide does have an in vivo stroma-targeting function, since chimeric genes in which the stroma-targeting domain of the plastocyanin transit peptide was replaced by the AtpC transit peptide-coding region allowed plastocyanin to accumulate in vivo. To determine whether the transit peptide deletions were impaired in in vivo stroma targeting, mutant and wild-type AtpC transit peptide-coding regions were fused to the bacterial ble gene, which confers bleomycin resistance. Although 25% of the wild-type fusion protein was associated with chloroplasts, proteins with transit peptide deletions remained almost entirely cytosolic. These results suggest that even severely impaired in vivo chloroplast protein import probably does not limit the accumulation of CF₁-.

Plant Physiol. (1998) 116: 1179-1190
Copyright Clearance Center:   0032-0889/98/116/1179/12
© 1998 American Society of Plant Physiologists

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