title: The BSD2 ortholog in Chlamydomonas reinhardtii is a polysome-associated chaperone that co-migrates on sucrose gradients with the rbcL transcript encoding the Rubisco large subunit.
abstract: The expression of the CO2 -fixation enzyme ribulose-bisphosphate carboxylase/oxygenase (Rubisco), which is affected by light, involves the cysteine-rich protein bundle-sheath defective-2 (BSD2) that was originally identified in maize bundle-sheath cells. We identified the BSD2 ortholog in Chlamydomonas reinhardtii as a small protein (17 kDa) localized to the chloroplast. The algal BSD2-ortholog contains four CXXCXGXG DnaJ-like elements, but lacks the other conserved domains of DnaJ. BSD2 co-migrated with the rbcL transcript on heavy polysomes, and both BSD2 and rbcL mRNA shifted to the lighter fractions under oxidizing conditions that repress the translation of the Rubisco large subunit (RbcL). This profile of co-migration supports the possibility that BSD2 is required for the de novo synthesis of RbcL. Furthermore, BSD2 co-migrated with the rbcL transcript in a C. reinhardtii premature-termination mutant that encodes the first 60 amino acids of RbcL. In both strains, BSD2 shared its migration profile with the rbcL transcript but not with psbA mRNA. The chaperone activity of BSD2 was exemplified by its ability to prevent the aggregation of both citrate synthase (CS) and RbcL in vitro following their chemical denaturation. This activity did not depend on the presence of the thiol groups on BSD2. In contrast, the activity of BSD2 in preventing the precipitation of reduced β-chains in vitro in the insulin turbidity assay was thiol-dependent. We conclude that BSD2 combines a chaperone 'holdase' function with the ability to interact with free thiols, with both activities being required to protect newly synthesized RbcL chains.
authors.affiliation: Department of Life Sciences, Ben-Gurion University of the Negev, POB 653, Beer Sheva, 84105, Israel.
authors.forename: Lior
authors.initials: L
authors.lastname: Doron
authors.forename: Na'ama
authors.initials: N
authors.lastname: Segal
authors.forename: Hadas
authors.initials: H
authors.lastname: Gibori
authors.forename: Michal
authors.initials: M
authors.lastname: Shapira
mesh_terms: D000595:Amino Acid Sequence; D016825:Chlamydomonas reinhardtii / Q000378:metabolism*; D018832:Molecular Chaperones / Q000378:metabolism*; D008969:Molecular Sequence Data; D018384:Oxidative Stress; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011132:Polyribosomes / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics*; D012273:Ribulose-Bisphosphate Carboxylase / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D013395:Sucrose
chemical_list: D018832:Molecular Chaperones; D010940:Plant Proteins; D012333:RNA, Messenger; D013395:Sucrose; D012273:Ribulose-Bisphosphate Carboxylase
keywords: BSD-2; Chlamydomonas; Rubisco; chaperone; oxidative stress; translation