Canonical Signal Recognition Particle Components Can Be Bypassed for Posttranslational Protein Targeting in Chloroplasts

Abstract

The chloroplast signal recognition particle (cpSRP) and its receptor (cpFtsY) target proteins both cotranslationally and posttranslationally to the thylakoids. This dual function enables cpSRP to utilize its posttranslational activities for targeting a family of nucleus-encoded light-harvesting chlorophyll binding proteins (LHCPs), the most abundant membrane proteins in plants. Previous in vitro experiments indicated an absolute requirement for all cpSRP pathway soluble components. In agreement, a cpFtsY mutant in Arabidopsis thaliana exhibits a severe chlorotic phenotype resulting from a massive loss of LHCPs. Surprisingly, a double mutant, cpftsy cpsrp54, recovers to a great extent from the chlorotic cpftsy phenotype. This establishes that in plants, a new alternative pathway exists that can bypass cpSRP posttranslational targeting activities. Using a mutant form of cpSRP43 that is unable to assemble with cpSRP54, we complemented the cpSRP43-deficient mutant and found that this subunit is required for the alternative pathway. Along with the ability of cpSRP43 alone to bind the ALBINO3 translocase required for LHCP integration, our results indicate that cpSRP43 has developed features to function independently of cpSRP54/cpFtsY in targeting LHCPs to the thylakoid membranes.

LTD is a protein required for sorting light-harvesting chlorophyll-binding proteins to the chloroplast SRP pathway

“…cpSRP54 was distributed almost equally between the stroma and thylakoids, and cpFtsY was mainly associated with the thylakoid membrane (Fig. 2b), which is consistent with previous reports 16,23,34,35 .…”

“…To elucidate the LTD function, we compared the effects of LTD inactivation on LHCP biogenesis with Arabidopsis mutants known to have defects in LHCP biogenesis (chaos, which lacks cpSRP43; ffc, which lacks cpSRP54; and cpftsY, which lacks cpFtsY) 16,23,28 (Fig. 1d).…”

“…Input, 4% of the prey protein. pigment deficiency phenotypes 16,21,23,28,29,34 . The ltd mutant did not develop any fertile flowers (Fig.…”

“…Homologues of prokaryotic SRP54 and its receptor FtsY have been identified in chloroplasts 15,20 . However, LTD and cpSRP43 homologues are found in higher plants, moss and green algae but not in cyanobacteria and red algae 23 ( Supplementary Fig. S1).…”

“…The T-DNA insertion lines ltd (SALK_126967C), cpftsY (SALK_049077) 34 and ffc mutant (CS850421) were obtained from the Arabidopsis Biological Resources Centers, and the homozygous mutants and precise T-DNA insertion sites were confirmed by PCR and sequencing with primers A, C and LB1 (all primer sequences are listed in Supplementary Table S1) for ltd; primers LP54, RP54 and LB2 for ffc; and primers FtsYF, FtsYR and LB1 for cpftsY. The chaos mutant and chaos/ffc double mutant were kindly provided by Dr Laurent Nussaume 23,29 . Wild-type and homozygous mutant plants were grown on Murashige and Skoog (MS) media supplemented with 3% sucrose under short-day conditions (10 h of light/14 h of dark) with a photon flux density of 80 µmol m − 2 s − 1 in a growth chamber at 22 °C.…”