Early events in phloem formation: Exploring the molecular network of SMXL3/4/5

Abstract

Growth and body shape of complex multicellular organisms is largely determined by a functional long-distance transport of energy metabolites that fuels stem cell activity. In vascular plants, sugars are photosynthetically produced in source tissues and delivered via the phloem to sink tissues for allocation into storage organs or to sustain distinct stem cell niches, called meristems. In the root apical meristem (RAM), which drives longitudinal root growth, sugar supply is ensured by a tight interplay between proto- and metaphloem. Formation of proto- and metaphloem starts with a single stem cell whose daughter cells divide and differentiate in a controlled spatio-temporal manner. Protophloem differentiates first within the RAM to enable sugar unloading close to the stem cell niche. Impaired or delayed protophloem formation has detrimental consequences for plant growth and vitality. Understanding the regulatory mechanisms behind (proto-)phloem formation is an important hub to enhance sink strength and thereby crop yield in the near or further future. In this study I report novel key-components in phloem regulation called SUPPRESSOR OF MAX2 1-LIKE3 (SMXL3), SMXL4 and SMXL5. Unlike most SMXL family members, SMXL3/4/5 act independently from strigolactone (SL) or karrikin (KAR) signaling as positive regulators of phloem formation. They are the first described phloem-specific genes that show promoter activity already in provascular tissues of the embryo, the first phloem stem cell in the RAM and along the whole phloem tissue in adult plants. SMXL3/4/5 promote protophloem initiation and differentiation in a dose-dependent manner. Deficiency of all three gene functions results in complete absence of phloem tissue and seedling lethality. In comparison, double mu-tants show reduced phloem-dependent transport and sugar accumulation in leaves. Moreover, SMXL3/5 play an additional and SMXL4-independent role in radial root growth by promoting procambial cell divisions. Interestingly, SMXL5 activity is sufficient to induce secondary phloem formation at the stem base, but acts redundantly with SMXL4 in suppressing radial stem thickening. This functional specialisation suggests that SMXL3/4/5 play distinct roles in molecular networks of phloem and/or (pro-)cambium formation. To integrate SMXL3/4/5 into such molecular networks, I characterized protein-protein interaction partners of SMXL5. The plant homeodomain (PHD)-finger protein OBERON 3 (OBE3) is the first interaction partner that genetically interacts with SMXL3/4/5 in protophloem formation. Previous studies reported that OBEs are important components in meristem maintenance and, potentially, chromatin remodelling. SMXL3/4/5 are nuclear localized, chap-eron-like proteins with conserved AAA ATPase and ETHYLENE-RESPONSE FACTOR Amphiphilic Repression (EAR) domain, which makes them perfect candidates to act in transcriptional regulation of downstream targets. This study and the characterization of SMXL3/4/5 and OBE3 as novel and fundamental phloem regulators enabled a deeper understanding of phloem development and sugar allocation in plants.