CLV3 Is Localized to the Extracellular Space, Where It Activates the Arabidopsis CLAVATA Stem Cell Signaling Pathway

Abstract

Plant growth and development depends on the activity of a continuously replenished pool of stem cells within the shoot apical meristem to supply cells for organogenesis. In Arabidopsis, the stem cell-specific protein CLAVATA3 (CLV3) acts cell nonautonomously to restrict the size of the stem cell population, but the hypothesis that CLV3 acts as an extracellular signaling molecule has not been tested. We used genetic and immunological assays to show that CLV3 localizes to the apoplast and that export to the extracellular space is required for its function in activating the CLV1/ CLV2 receptor complex. Apoplastic localization allows CLV3 to signal from the stem cell population to the organizing center in the underlying cells. INTRODUCTIONPlants, unlike animals, continuously produce organs from their growing tips, which are called apical meristems. In the shoot apical meristem (SAM), which generates all of the aboveground structures of the plant, a few cells at the apex are maintained as a pluripotent stem cell population. As the stem cells divide, their progeny are displaced toward the flanks of the meristem, where they become incorporated into organ primordia. The maintenance of a functional SAM requires precise coordination between the loss of stem cells from the meristem through organogenesis and their replacement through cell division.The Arabidopsis CLAVATA1 ( CLV1 ), CLV2 , and CLV3 genes are required to restrict the amount of stem cell accumulation in both shoot and floral meristems. Plants with loss-of-function mutations in any of the CLV genes form greatly enlarged shoot and floral meristems, causing stem overgrowth and the production of extra flowers and floral organs (Clark et al., 1993(Clark et al., , 1995Kayes and Clark, 1998). The CLV1 gene encodes a Leu-rich repeat receptor Ser/Thr kinase (Clark et al., 1997), a member of a large class of proteins found in both plants and animals, many of which are involved in cell signaling. CLV2 encodes a Leu-rich repeat receptor-like transmembrane protein with a short cytoplasmic tail (Jeong et al., 1999). Together, CLV1 and CLV2 form components of a membrane-bound receptor signal transduction complex (Trotochaud et al., 2000). CLV3 encodes a 96-amino acid predicted extracellular protein (Fletcher et al., 1999) that can act in a cell nonautonomous manner (Fletcher et al., 1999;Brand et al., 2000). CLV3 binds to CLV1 and CLV2 and is required for the formation of the active receptor complex, indicating that CLV3 acts as a ligand that signals through CLV1 and CLV2 (Trotochaud et al., 2000).CLV3 and CLV1 expression is restricted to subsets of shoot and floral meristem cells. CLV3 mRNA is expressed in stem cells, which are found primarily in the epidermal and subepidermal cell layers of shoot and floral meristems (Fletcher et al., 1999). CLV1 is expressed in the underlying cells, in a domain partially overlapping that of CLV3 (Clark et al., 1997). It has been hypothesized that CLV3 is made in the overlying cell layers and moves to the underlying cells to activate the C...

ArabidopsisRETINOBLASTOMA-RELATED Is Required for Stem Cell Maintenance, Cell Differentiation, and Lateral Organ Production

“…At present, our results cannot distinguish if downregulation of RBR or the resulting loss of cell-cell contact between L1 and L2 disrupts the CLV-WUS feedback loop, which maintains the SAM stem cell niche that contributes to meristem activity and primordia formation (Brand et al, 2000(Brand et al, , 2002Rojo et al, 2002;Tucker and Laux, 2007). Loss of CLV3 activity leads to gradual conversion of bordering cells in the SAM peripheral zone to cells with central zone characteristics , suggesting that CLV3 is involved in a signaling pathway required for the switch from stemness to differentiated cells.…”

CLAVATA1 Dominant-Negative Alleles Reveal Functional Overlap between Multiple Receptor Kinases That Regulate Meristem and Organ Development

“…Thus, CLV3 retains some function in clv1 mutants, especially in clv1 null mutants. Because CLV3 is secreted and required to activate CLV1 Rojo et al, 2002), CLV3 must activate another target in clv1 null mutants, presumably a receptor kinase. Furthermore, the hypothesis that the clv1 dominant-negative isoforms sequester the putative CLV3 ligand is not consistent with studies on a likely feedback loop between WUS and CLV3 (Brand et al, 2000;Schoof et al, 2000;Lenhard et al, 2002).…”

“…CLV2 belongs to the receptor-like protein family and contains LRR motifs in its predicted extracellular domain but no intracellular kinase domain (Jeong et al, 1999). CLV3 encodes a small polypeptide that has been shown to be secreted (Fletcher et al, 1999;Rojo et al, 2002). The CLV1 , CLV2 , and CLV3 genes function in the same pathway, the formation of active CLV1 complexes requires CLV3, CLV3 overexpression phenotypes require CLV1 and CLV2, and CLV3 is secreted from cells adjacent to those expressing CLV1 -all suggesting that CLV3 acts as the ligand for CLV1 (Clark et al, 1995(Clark et al, , 1997Fletcher et al, 1999;Brand et al, 2000;Rojo et al, 2002).…”

Enlarged meristems and delayed growth in plp mutants result from lack of CaaX prenyltransferases

“…Plants with mutations in any of three loci, CLV1, CLV2, or CLV3, show a progressive increase in meristem size beginning in the embryo and continuing throughout life, indicating a loss of cell division restriction (5)(6)(7)(8). Available evidence indicates that CLV3 encodes a small secreted peptide expressed in outer cell layers (9,10) and likely binds to the leucine-rich repeat receptor kinase CLV1 and its putative dimerization partner CLV2, which are expressed in inner cell layers (11)(12)(13). Activation of the CLV complex results in negative regulation of cell proliferation (14).…”