RBMP2 shapes specialized membranes for CO(2) delivery in the pyrenoid condensate.
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RBMP2 is in the reticulated region
"The predicted transmembrane protein RBMP2 localizes to the reticulated region27"
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Loss of RBMP2 removes the reticulated region, mislocalizes CAH3 and impairs growth at very low CO2
"We show that RBMP2 loss eliminates this central reticulated membrane network, mislocalizes CAH3 and impairs growth under very low CO2."
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Growth defect is rescued by RBMP2-Venus
"We conclude that RBMP2 is required for normal CAH3 localization and for maintaining pyrenoid function under strongly CO2-limiting conditions."
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TEM confirms the reticulated region is absent and restored by complementation
"Together, these results indicate that RBMP2 is required for reticulated-region formation."
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Overexpression enlarges the reticulated region
"Thus, RBMP2 is not only required for reticulated-region formation; overexpressing the RBMP2 protein is sufficient to promote the expansion of the reticulated region in wild-type cells."
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Cylindrical tubules still enter the matrix without RBMP2 but do not form a reticulated network
"In the rbmp2 mutant, cylindrical tubules still extended from the pyrenoid periphery toward the center, but we never observed them forming a reticulated network"
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Domain architecture, including a non-catalytic rhodanese domain flanked by transmembrane helices and six C-terminal RBMs
"RBMP2 contains an N-terminal region with predicted membrane-contact probability (MCP)33, a central non-catalytic rhodanese domain (RHO), predicted transmembrane helices (TM) on both sides of the RHO domain and six C-terminal Rubisco-binding motifs (RBMs)27"
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An aspartate replaces the catalytic cysteine in the RBMP2 rhodanese domain
"The catalytic cysteine residue is marked in green, and the non-catalytic aspartic acid residue in RBMP2 is marked in grey."
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The rhodanese domain is needed for tubule extension
"These results suggest that the RHO domain performs an essential and upstream function in reticulated-region biogenesis, likely promoting extension of the tubules."
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The MCP and TM regions are needed to convert tubules into reticulated-region tubules
"MCP and TM domains are not required for cylindrical tubule elongation. Instead, they are required to convert elongated cylindrical tubules into reticulated-region tubules."
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The six RBMs are dispensable for reticulated-region formation
"These results suggest that the RBMs are dispensable for detectable reticulated-region formation, although they may still contribute to RBMP2 function in ways not resolved by these assays."
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Tubule and minitubule fusion use other factors
"Tubule and minitubule fusion are likely mediated by factors other than RBMP2"
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LCI16 interacts with RBMP2 but is not needed for the reticulated region
"However, the presence of a normal reticulated region in the lci16 mutant38 suggests that LCI16 is not essential for reticulated-region biogenesis."