Plant meristems require a constant supply of photoassimilates and hormones to the dividing meristematic cells. In the growing root, such supply is delivered by protophloem sieve elements. Due to its preeminent function for the root apical meristem, protophloem is the first tissue to differentiate. This process is regulated by a genetic circuit involving in one side the positive regulatorsDOFtranscription factors,OCTOPUS(OPS) andBREVIX RADIX(BRX), and in the other side the negative regulatorsCLAVATA3/EMBRYO SURROUNDING REGION RELATED(CLE) peptides and their cognate receptorsBARELY ANY MERISTEM(BAM) receptor-like kinases.brxandopsmutants harbor a discontinuous protophloem that can be fully rescued by mutation inBAM3, but is only partially rescued when all three known phloem-specificCLEgenes,CLE25/26/45are simultaneously mutated. Here we identify aCLEgene closely related toCLE45, namedCLE33. We show that double mutantcle33cle45fully suppressesbrxandopsprotophloem phenotype.CLE33orthologs are found in basal angiosperms, monocots, and eudicots, and the gene duplication which gave rise toCLE45in Arabidopsis and otherBrassicaceaeappears to be a recent event. We thus discovered previously unidentified ArabidopsisCLEgene that is an essential player in protophloem formation.