Translocation of a Phycoerythrin α Subunit across Five Biological Membranes

Abstract

Cryptophytes, unicellular algae, evolved by secondary endosymbiosis and contain plastids surrounded by four membranes. In contrast to cyanobacteria and red algae, their phycobiliproteins do not assemble into phycobilisomes and are located within the thylakoid lumen instead of the stroma. We identified two gene families encoding phycoerythrin ␣ and light-harvesting complex proteins from an expressed sequence tag library of the cryptophyte Guillardia theta. The proteins bear a bipartite topogenic signal responsible for the transport of nuclear encoded proteins via the ER into the plastid. Analysis of the phycoerythrin ␣ sequences revealed that more than half of them carry an additional, third topogenic signal comprising a twin arginine motif, which is indicative of Tat (twin arginine transport)-specific targeting signals. We performed import studies with several derivatives of one member using a diatom transformation system, as well as intact chloroplasts and thylakoid vesicles isolated from pea. We demonstrated the different targeting properties of each individual part of the tripartite leader and show that phycoerythrin ␣ is transported across the thylakoid membrane into the thylakoid lumen and protease-protected. Furthermore, we showed that thylakoid transport of phycoerythrin ␣ takes place by the Tat pathway even if the 36 amino acid long bipartite topogenic signal precedes the actual twin arginine signal. This is the first experimental evidence of a protein being targeted across five biological membranes.

Chlorophyll-binding proteins revisited - a multigenic family of light-harvesting and stress proteins from a brown algal perspective

“…In the case of red algae, only the reduced genome of C. merolae was available for public use, and we added EST sequences from C. crispus (4,114 ESTs, 3 coding for CBPs; [51]), P. yezoensis (20,069 ESTs, 4 coding for CBPs; [52]), and G. changii (8,147 ESTs, 7 coding for CBPs; Teo et al, unpublished), all of which were obtained from the NCBI EST database (dbEST). Furthermore, as no genome sequences are available for cryptophytes, the same strategy was applied for an EST library of Guillardia theta (15,173 ESTs, 21 coding for CBPs; [53]), and we also took advantage of the availability of five CBP sequences obtained in a previous study by Broughton et al on Rhodomonas sp. CS24 [35].…”

Complementation of a phycocyanin-bilin lyase from Synechocystis sp. PCC 6803 with a nucleomorph-encoded open reading frame from the cryptophyte Guillardia theta

“…The β-subunit is encoded on the plastid genome [22], whereas the phycoerythrin α-subunits are encoded by a nuclear-located gene family. In the latter case, the genes encode preproteins containing a tripartite topogenic signal responsible for the translocation across five biological membranes [23]. Because a wide range of genomic data exists from this unicellular phototrophic organism, existing knowledge can be used to reconstruct the biochemistry of these organisms.…”

Light capture and pigment diversity in marine and freshwater cryptophytes

“…Cryptophytes are thus complex with respect to their genetic and cellular structure, and are distinctive in their pigmentation and the organization of these pigments within the plastids (Gould et al. ).…”