Page 158 - Plant Canada 2024 Proceeding

PLANT CANADA 2024

like motif is necessary for targeting, where the transit peptide-like motif significantly improves targeting
fidelity. Additionally, the transit peptide-like motif exhibits a preferential interaction toward galactolipids
unique to the plastid membrane as shown by Langmuir-Blodgett trough experiments using galactolipid
monolayers and CD spectroscopy experiments using liposomes containing galactolipids. Together, these
experiments demonstrate higher maximum insertion pressures of the transit peptide-like motif in the
presence of galactolipid monolayers compared to monolayers composed of phosphatidylcholine (PC).
The transit peptide-like motif is unstructured in buffer and in the presence of PC liposomes, but form ⍺-
helical structures in the presence of liposomes containing galactolipids. This implies a role for lipid
composition in targeting. A better understanding of TOC159 receptor biogenesis is critical in describing
how TOC complexes are assembled and function in the chloroplast outer envelope, responsible for
regulating chloroplast biogenesis and plastid morphogenesis in plant cells.

[O111] THE REGULATORY FUNCTION OF PLASTID CHAPERONE HSP90C C-TERMINAL
1,2
1,2
EXTENSION. Bona Mu , Adheip Monakan Nair , and Rongmin Zhao . Departments of Biological
1,2 1
2
Sciences, University of Toronto Scarborough; and Cell & Systems Biology, University of Toronto, Toronto,
Canada
Correspondence to: rongmin.zhao@utoronto.ca

HSP90Cs are essential molecular chaperones localized in the plastid stroma that maintain protein
homeostasis and assist the import and thylakoid transport of chloroplast proteins. While HSP90C
contains all conserved domains as an HSP90 family protein, it also possesses a unique feature in its
variable C-terminal extension (CTE) region. This study elucidated the specific function of this HSP90C
CTE region. Our phylogenetic analyses revealed that this intrinsically disordered region contains a highly
conserved DPW motif in the green lineages. With biochemical assays, we showed that the CTE is
required for the chaperone to effectively interact with client proteins PsbO1 and LHCB2 to regulate ATP-
independent chaperone activity and to effectuate its ATP hydrolysis. While the CTE truncation mutants
could support plant growth and development indistinguishably from the wild-type protein under normal
conditions, higher HSP90C expression was observed to correlate with a stronger response to specific
photosystem II inhibitor DCMU, and CTE truncations dampened the response. Additionally, when treated
with lincomycin to inhibit chloroplast protein translation, CTE truncation mutants showed a delayed
response to PsbO1 expression repression, suggesting its role in chloroplast retrograde signalling. Our
study therefore provides insights into the mechanism of HSP90C in client protein binding and in the
regulation of green chloroplast maturation and function, especially under stress conditions.

*[O112] IDENTIFICATION AND CHARACTERIZATION OF OEP6 MOTIFS AND THEIR ROLE IN
1
1
TARGETING TO THE CHLOROPLAST OUTER MEMBRANE. Holly Ferguson , Matthew Smith , and
Simon Chuong . Wilfrid Laurier University, 75 University Ave W, Waterloo, ON, Canada, N2L 3C5; and
2 1
2 University of Waterloo, 200 University Ave W, Waterloo, ON, Canada, N2L 3G1
Correspondence to: ferg2170@mylaurier.ca

The chloroplast is a type of membrane-bound plastid commonly known for its role in photosynthesis.
Chloroplasts evolved from a cyanobacterium that was engulfed by a eukaryotic cell and through this
endosymbiotic process, the majority of the cyanobacterial genes were lost to the host nucleus. Therefore,
the vast majority of chloroplast proteins necessary for its biogenesis and function are encoded in the host
nucleus, translated in the cytoplasm, and then transported from the cytosol back to the chloroplast.
Proteins that are targeted to different compartments of the chloroplast require targeting signals to facilitate
their delivery and import. The import of these stromal peptides is facilitated by the activity of protein
complexes at the outer and inner envelope membranes, TOC and TIC, respectively. To date, chloroplast
outer envelope proteins (OEPs) have been reported to utilize one of five different targeting mechanisms:
(1) an N-terminal transit peptide; (2) a transmembrane domain located at either the N-terminus (signal-
anchored proteins) or (3) the C-terminus (tail-anchored proteins); (4) a signal included within a -barrel
structure that forms a channel; and (5) a reverse transit peptide-like signal at the C-terminus (Fish et al.
2022). My research focuses on OEP6, which is predicted to be a tail-anchored protein, but whose
targeting mechanism is not yet well-characterized. My study aims to identify and characterize features

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