The peptides RALF4 and RALF19 are secreted by the pollen tube and bind to a plasma membrane receptor complex composed of BUDDHA’S PAPER SEAL 1/2 and ANXUR 1/2. This interaction is required for pollen tube integrity during growth through the transmitting tract. Once arrived, ovule-expressed RALF34 displaces RALF4/19 from their receptors, causing pollen tube rupture and sperm cell release. Hence, RALF peptides regulate both integrity and rupture of pollen tubes through intracellular signalling in the respective environments. Mechanistically, however, what maintains or disrupts cell wall integrity remains poorly understood. Extensins are a family of hydroxyproline-rich glycoproteins that can crosslink and thus form a network in the cell wall. Therefore, the previously reported interaction of RALF4 with LRX8 containing an extensin domain posed the question of whether the RALF4 peptide may attain a structural role through this interaction.
The researchers found that the interaction of RALF4 with LRX8 leads to the exposure of basic lysin and arginin residues of RALF4. Interestingly, basic residues of other extensins involved in pectin interactions are reduced in LRX8, suggesting that RALF4 may take over this role. Indeed, the RALF4–LRX8 complex is stably integrated into the cell wall and can bind to pectin with low levels of methylesterification. Mutation of the basic residues in RALF4 specifically impairs pectin interaction. Several phenotypes of mutants with reduced levels of RALF4 (and RALF19), such as decreased fertility, higher growth velocity and premature bursting rate, and callose accumulation, could not be fully complemented with RALF4 when basic residues were mutated. Interaction of RALF4–LRX8 with pectin leads to condensation and patterning into a reticulate network, suggesting that a pectin–RALF4–LRX8 complex may be required for maintenance of cell wall integrity during pollen tube growth.
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