Co-evolution of Two GTPases Enables Efficient Protein Targeting in an RNA-less Chloroplast Signal Recognition Particle Pathway

Abstract

The signal recognition particle (SRP) is an essential ribonucleoprotein particle that mediates the co-translational targeting of newly synthesized proteins to cellular membranes. The SRP RNA is a universally conserved component of SRP that mediates key interactions between two GTPases in SRP and its receptor, thus enabling rapid delivery of cargo to the target membrane. Notably, this essential RNA is bypassed in the chloroplast (cp) SRP of green plants. Previously, we showed that the cpSRP and cpSRP receptor GTPases (cpSRP54 and cpFtsY, respectively) interact efficiently by themselves without the SRP RNA. Here, we explore the molecular mechanism by which this is accomplished. Fluorescence analyses showed that, in the absence of SRP RNA, the M-domain of cpSRP54 both accelerates and stabilizes complex assembly between cpSRP54 and cpFtsY. Cross-linking coupled with mass spectrometry and mutational analyses identified a new interaction between complementarily charged residues on the cpFtsY G-domain and the vicinity of the cpSRP54 M-domain. These residues are specifically conserved in plastids, and their evolution coincides with the loss of SRP RNA in green plants. These results provide an example of how proteins replace the functions of RNA during evolution.

PALE‐GREEN LEAF 1, a rice cpSRP54 protein, is essential for the assembly of the PSI‐LHCI supercomplex

“…The stroma‐located chloroplast signal recognition particle (cpSRP) consists of two subunits, the conserved 54‐kDa GTPase cpSRP54 and a chloroplast‐specific 43‐kDa protein, cpSRP43, and both are found to be essential for assisting the mature but unfolded light‐harvesting Chl a/b binding proteins (LHCP) integration into thylakoid membrane (Akopian et al, 2013 ; Ziehe et al, 2017 ). After binding the LHCP to the heterodimeric cpSRP43/cpSRP54 complex, they form a soluble transit complex and dock to the SRP receptor cpFtsY and the Alb3 translocase at the membrane followed by the release and integration of the LHCP into the thylakoid membrane in a GTP‐dependent manner (Chandrasekar et al, 2017 ). The cpSRP54 protein contains an N‐terminal N domain, a central G domain with GTPase activity, and a methionine‐rich M domain in the C‐terminus (Franklin & Hoffman, 1993 ).…”

Chloroplast SRP43 autonomously protects chlorophyll biosynthesis proteins against heat shock

“…cpSRP43 is an ATP-independent chaperone that is responsible for protecting LHCPs from aggregation 26 , while cpSRP54 serves to allosterically activates its LHCP-targeted chaperone activity 27,28 . cpSRP54 also mediates interaction of the cpSRP complex with the SRP receptor cpFtsY to deliver LHCPs to the translocase Albino 3 (Alb3) at the thylakoid membrane 29,30 . Alb3 then triggers the release of LHCP from cpSRP43 and mediates the integration of LHCP into the thylakoid membrane 31 , where assembly of Chls into the LHCs takes place.…”

Chloroplast SRP43 acts as a chaperone for glutamyl-tRNA reductase, the rate-limiting enzyme in tetrapyrrole biosynthesis

“…The chloroplast-specific cpSRP43 interacts with LHCPs and cpSRP54 to form a soluble "transit complex" (11). Interaction of the GTPase domains of cpSRP54 with the SRP receptor cpFtsY targets the transit complex to the ALB3 translocase at the thylakoid membrane, which mediates the in-sertion of LHCPs (12). Remarkably, cpSRP43 coevolved with the emergence of LHCPs in green land plants (13,14) and is suggested to be a dedicated chaperone that quantitatively prevents aggregation of the LHCP family in aqueous environments (15)(16)(17).…”