Glycome profiling and immunohistochemistry uncover changes in cell walls of Arabidopsis thaliana roots during spaceflight

Abstract

A large and diverse library of glycan-directed monoclonal antibodies (mAbs) was used to determine if plant cell walls are modified by low-gravity conditions encountered during spaceflight. This method called glycome profiling (glycomics) revealed global differences in non-cellulosic cell wall epitopes in Arabidopsis thaliana root extracts recovered from RNA purification columns between seedlings grown on the International Space Station-based Vegetable Production System and paired ground (1-g) controls. Immunohistochemistry on 11-day-old seedling primary root sections showed that ten of twenty-two mAbs that exhibited spaceflight-induced increases in binding through glycomics, labeled space-grown roots more intensely than those from the ground. The ten mAbs recognized xyloglucan, xylan, and arabinogalactan epitopes. Notably, three xylem-enriched unsubstituted xylan backbone epitopes were more intensely labeled in space-grown roots than in ground-grown roots, suggesting that the spaceflight environment accelerated root secondary cell wall formation. This study highlights the feasibility of glycomics for high-throughput evaluation of cell wall glycans using only root high alkaline extracts from RNA purification columns, and subsequent validation of these results by immunohistochemistry. This approach will benefit plant space biological studies because it extends the analyses possible from the limited amounts of samples returned from spaceflight and help uncover microgravity-induced tissue-specific changes in plant cell walls.

Conserved plant transcriptional responses to microgravity from two consecutive spaceflight experiments

“…Genes involved in remodeling of plant cell walls have been reported to be differentially expressed in previous spaceflight experiments ( Paul et al., 2013 ; Kwon et al., 2015 ; Johnson et al., 2017 ; Choi et al., 2019 ; Nakashima et al., 2023 ). Glycomic analysis of spaceflight plant samples have shown changes in cell wall glycans, consistent with spaceflight related modification of cell walls ( Johnson et al., 2017 ; Nakashima et al., 2023 ).…”

“…Genes involved in remodeling of plant cell walls have been reported to be differentially expressed in previous spaceflight experiments ( Paul et al., 2013 ; Kwon et al., 2015 ; Johnson et al., 2017 ; Choi et al., 2019 ; Nakashima et al., 2023 ). Glycomic analysis of spaceflight plant samples have shown changes in cell wall glycans, consistent with spaceflight related modification of cell walls ( Johnson et al., 2017 ; Nakashima et al., 2023 ). Similar to these previous reports, we also observed that genes involved in cell wall modification are overrepresented in our datasets; particularly we found approximately 100 genes associated with cell wall organization and modification downregulated in µ g .…”

Integrative transcriptomics and proteomics profiling of Arabidopsis thaliana elucidates novel mechanisms underlying spaceflight adaptation

“…(2017) also reported the reduced abundance of xylan and pectic epitopes in spaceflight-grown plants, which resonate with our findings, highlighting the potential changes in the biosynthesis of cell wall non-cellulosic polysaccharides in spaceflight conditions, particularly in the root. Nakashima et al. (2023) in their novel glycome profiling and immunohistochemistry of Arabidopsis in spaceflight also observed that roots from spaceflight Arabidopsis seedlings displayed a generally increased labeling intensity when targeted with xyloglucan monoclonal antibodies (mAbs) compared to ground controls.…”