New insights on WOX gene evolution in land plant development from characterizing their expression and functions in the fern Ceratopteris richardii
Creators: name: Youngstrom, Christopher E.
Publisher: name: University of Iowa
Year: 2024
Subjects: Ceratopteris richardii
ferns
WOX
wuschel
public abstract
Evolution
Stem Cells

description: Plants have the lifelong ability to generate new organs partly due to the persistent functioning ofstem cells. In seed plants, groups of stem cells are housed in the shoot apical meristem (SAM), root apical meristem (RAM), and vascular cambium (VC). In contrast, seed-free plants often have only a single stem cell in the SAM and RAM, and no VC. How land plants evolve and maintain multiple stem cells in apical meristems is an unanswered question in the evolution of plant development. In seed plants, a key regulator of stem cell maintenance are the WUSCHEL-related homeobox (WOX) transcription factors. In the angiosperm Arabidopsis thaliana, AtWUS, AtWOX5, and AtWOX4, maintain the number and identity of stem cells in the SAM, RAM, and VC, respectively. As the extant seed-free sister taxa of seed plants, ferns offer an opportunity to investigate if orthologs of AtWUS, AtWOX5 or AtWOX4 are required in stem cell maintenance of single-stem-cell type meristems and vascular tissues.
description: The model fern Ceratopteris richardii contains five WOX genes (CrWOX13A, CrWOX13B, CrWOXA, CrWOXB and CrWUL), one of which, CrWUL is likely orthologous to AtWUS, AtWOX5 and AtWOX4. I examined the expression domains and measured levels of all five CrWOX transcripts via RT- qPCR and in situ hybridization, then I used an RNAi knockdown approach to determine the functions of CrWOXA, CrWOXB, and CrWUL. I found that CrWOX genes affect lateral meristem activity in gametophytes. Specifically, CrWOXA and CrWOXB promote cell divisions, while CrWUL inhibits them. In sporophytes, CrWOXA, CrWOXB and CrWUL do not maintain the stem cell of the SAM and RAM, however in vascular tissues, CrWUL maintains the number of phloem cells. To determine conserved roles of CrWUL, I performed cross-species complementation assays in A. thaliana with atwox4 null mutants. I found that CrWUL maintains the stem cells in the VC of A. thaliana, which is consistent with the idea that ancestral WOX function arose in vascular tissues, before being co-opted into the SAM and RAM of seed plants. Furthermore, I examined the expression of the likely CrWUL ortholog in Selaginella kraussiana and showed that SkWOX11C is expressed in the vasculature but not in the shoot apical meristem, suggesting WOX genes were recruited to vascular tissues in the last common ancestor (LCA) of lycophytes and ferns.
description: Here I show how CrWOX expression and function in C. richardii can contribute to an evolutionary trajectory for the WOX transcription factor family in land plants. Given the success of cross- species complementation assays in the presented data, future studies could use a similar approach to examine the roles of WOX genes in specifying reproductive cells, embryogenesis, and response to chemical and peptide hormones. Understanding the ancestral roles of WOX genes in these processes allows us to deduce at what point along the land plant phylogeny did WOX networks integrate into developmental pathways.