Dual targeting of the protein disulfide isomerase RB60 to the chloroplast and the endoplasmic reticulum.
RB60 is an atypical protein disulfide isomerase (PDI) that functions as a member of a redox regulatory protein complex controlling translation in the chloroplast of Chlamydomonas reinhardtii, but also contains a C-terminal endoplasmic reticulum (ER) retention signal, -KDEL. Here, we show by fluorescence microscopy that RB60 resides in the chloroplast but also outside of the chloroplast colocalized with BiP, an ER marker protein. RB60 accumulates in microsomes that exhibit a typical ER magnesium-shift, and cotranslationally translocates into ER microsomes. The first 50-aa leader of RB60 is sufficient for both chloroplast and ER targeting. The leader is cleaved upon translocation into the ER, whereas it remains intact after import to the chloroplast. The leader sequence also contains an acidic domain that appears necessary for the protein's association with the thylakoid membranes. Based on these and additional results, we propose that the dual localization of RB60 occurs via the two conserved transport mechanisms, to the chloroplast and to the ER, that the chloroplast RB60 most likely carries an additional function in the ER, and that its mode of transport, including the differential cleavage of its N terminus, plays an important role in its suborganellar localization and organellar-specific function.
Levitan
Alexander
A
Department of Plant Sciences, Weizmann Institute of Science, Rehovot 76100, Israel.
Trebitsh
Tova
T
Kiss
Vladimir
V
Pereg
Yaron
Y
Dangoor
Inbal
I
Danon
Avihai
A
D000595:Amino Acid Sequence; D000818:Animals; D016825:Chlamydomonas reinhardtii / Q000201:enzymology* / Q000254:growth & development; D002736:Chloroplasts / Q000201:enzymology*; D004721:Endoplasmic Reticulum / Q000201:enzymology*; D006801:Humans; D008969:Molecular Sequence Data; D018532:Pisum sativum / Q000201:enzymology; D019704:Protein Disulfide-Isomerases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D019704:Protein Disulfide-Isomerases