ERAD-mediated maturation of the regulatory protein of plant meristematic cells CLAVATA 3 emerged during evolution from algae to higher plants

The indefinite growth capacity of plants is guaranteed by meristems, where cells are in continuous division and differentiation. The activity of these stem cells is under control of the negative feedback loop between the genes WUSCHEL (WUS) and CLAVATA3 (CLV3) (Brand et al. 2000). CLV3 gene encodes a small secretory protein that undergoes proteolytic maturation to release an active dodecapeptide that binds to a receptor complex (CLV1/2), but the actual maturation mechanism is still unknown (Kondo et al. 2006). The current model predicts CLV3 secretion in the extracellular space and maturation mediated by specific, but not yet identified, proteases. Conversely, a recent study suggested that CLV3-GFP maturation in transgenic tobacco plants occurred in the endoplasmic reticulum (ER) via a process mediated by ER Associated Degradation (ERAD). Moreover, the processed CLV3 dodecapeptide was biologically active because it inhibited Arabidopsis thaliana root growth (De Marchis et al. 2018). Here we show, by using a physiological assay based on root growth inhibition, that the production of the active 12-amino acid peptide is absent in transgenic algae of the genus Chlamydomonas. Our idea is that the CLV3 maturation process emerged when the first photosynthetic organism evolved into vascular plants, therefore unicellular algae should lack this process. To prove our hypothesis, we transformed cells of C. reinardtii with a CLV3-YFP fusion gene. In contrast to what is observed in transgenic tobacco, we detected secretion of the intact CLV3-YFP protein in the algal growth medium, without the production of the active dodecapeptide, therefore corroborating our hypothesis. Our future goal is to identify the plant genes involved in CLV3 maturation.