As above, so below: CLE peptide signaling in shoot and root apical meristems

The shoot apical meristem (SAM) and the root apical meristem (RAM) contain stem cells that give rise to plant organs above and below the ground. Maintenance of meristems is essential for the development and growth and hence regulated by signal transduction systems that contain multiple components that function redundantly. The CLAVATA3 (CLV3)/EMBRYO SURROUNDING REGION-related (CLE) peptides are major regulators of meristem homeostasis that are perceived by leucine-rich repeat receptor-like protein kinases (LRR-RLKs) together with their co-receptors in the plasma membrane. However, the intracellular signal transduction events that follow CLE peptide perception are largely unknown.

In the SAM, the peptide CLV3 is recognized by a receptor complex composed of the LRR-RLK CLV1 and a CLV3-INSENSITIVE RECEPTOR KINASE (CIK) (Hu et al., 2018; Song et al., 2021). In the RAM, the CLE peptides 25 and 45 are similarly recognized by a receptor complex comprising the LRR-RLK BARELY ANY MERISTEM (BAM) and a CIK (Zhu et al., 2020; Hu et al., 2022). In the current issue of The Plant Cell, Wenping Wang, Chong Hu and co-workers (Wang et al., 2022) show that the receptor-like cytoplasmic kinases (RLCKs) PBS1-like 34, 35, and 36 (PBL34/35/36) act as intracellular signaling components downstream of CLE peptide sensing by the plasma membrane receptor complexes both in the SAM and the RAM.

As PBS1-like proteins (PBLs) from the RLCK subfamily VII were previously associated with peptide signal transduction, the authors tested if they were also involved in CLE peptide signaling in the SAM and the RAM. They identified a redundant role for PBL34/35/36. Triple mutants lacking all respective PBLs had higher SAMs and more carpels per flower, and were less sensitive to suppression of root length triggered by exogenous CLE25 or CLE45 peptides (see Figure). These findings suggested that PBL34/35/36 are involved in CLE peptide-mediated regulation of meristem homeostasis. In line with this possibility, the pbl34 pbl35 pbl36 triple mutant was less sensitive to exogenous CLV3 in addition to CLE25/45. In shoots, CLV3 treatment led to termination of the SAM in wild-type plants, whereas 60% of the triple mutants retained the SAM and could generate inflorescence stems. In roots, exogenous CLE45 suppressed thickening of root cell walls associated with protophloem differentiation in wild-type plants, but not in pbl34 pbl35 pbl36 triple mutants.

Figure.

CLE peptide-mediated suppression of root growth is reduced in pbl34 pbl35 pbl36 triple mutants. Exogenous CLE25 and CLE45 suppress root growth in Col wild-type but not in pbl34 pbl35 pbl36 triple mutants. Adapted from Wang et al. (2022), Figure 1.

Combining triple or double mutations in PBL34/35/36 with impairments in various CLE peptide receptors indicated that these PBLs function in the same pathway as CLV1 in shoots and as BAM3 and CIK2 in roots. Yeast two-hybrid assays and bimolecular fluorescence complementation experiments showed that PBL34/35/36 interact with CLV1, supporting the hypothesis that CLV1 transmits the CLV3 peptide signal to these PBLs. For PBL35/36, the interaction with CLV1 was further confirmed by fluorescence resonance energy transfer (FRET) analyses and co-immunoprecipitation experiments. Similarly, interactions of each of the three PBLs with BAM3 and CIK2 were identified, suggesting that these interactions are important for CLE peptide signaling in roots. In vitro kinase assays showed that PBL34 and PBL35 were phosphorylated by CLV1 and BAM1, indicating that these RLCKs are direct substrates of CLV1 and BAM1 in pathways that control SAM and RAM homeostasis.

The work by Wang et al. (2022) identifies a missing link between CLE peptide perception and downstream regulation of gene expression: RLCKs PBL34/35/36 are direct substrates of CLE peptide receptor complexes that contribute to CLE peptide signaling both in the SAM and the RAM. The partially impaired meristem regulation phenotypes of the pbl34 pbl35 pbl36 triple mutant suggest that other PBLs may also be involved in maintaining apical meristem homeostasis. The authors also point out that the pbl34 pbl35 pbl36 triple mutant is a relatively weak mutant with moderately increased SAM size that is not accompanied by severe detrimental effects on shoot development. Similar mutations could thus be potentially used to improve crop productivity.