Diversification of CLE expression patterns and nonmeristematic roles for CLAVATA receptor-like kinases in a moss

The CLAVATA pathway controls meristematic cell proliferation and multiple nonmeristematic processes in Arabidopsis development. While CLAVATA ancestrally regulates meristematic proliferation in nonseed plant gametophytes, ancestral sporophytic and nonmeristematic functions in land plants are unknown. Here, we analysed the promoter activities of all peptide (PpCLE) and receptor-encoding (PpCLV1a, PpCLV1b and PpRPK2) genes throughout the moss (Physcomitrium patens) life cycle and validated our expression analyses using mutant phenotype data. In gametophore apices, PpCLE3 expression marked apical cells, and PpCLV1b and PpRPK2 overlapped. In nonmeristematic tissues, gametophytes showed highly focal PpCLE but broader receptor-encoding gene expression, and many genes were co-expressed. Mutant phenotype analysis revealed roles for PpCLV1a, PpCLV1b and PpRPK2 in fertility and male and female reproductive development. In sporophytes, no PpCLE expression specifically marked the apical cells, and PpCLV1b and PpRPK2 expression initially marked distinct apical and basal domains, but later overlapped at the intercalary meristem. Overall, fewer genes were co-expressed in sporophytes than in gametophytes, but all genes were co-expressed in guard cells. Our data indicate that nonmeristematic CLAVATA functions in gametangium development and stomatal development may be ancestral within land plants. Peptide encoding (CLE) gene copy numbers amplified in mosses, and promoter evolution was a likely driver of cell type diversification during moss evolution.