Deubiquitinase <scp>ZmUBP5</scp> is essential for maize kernel development

Ubiquitination and deubiquitination are critical post-translational modifications in eukaryotes, playing essential roles in various aspects of plant growth and development. However, their involvement in maize kernel development remains largely unexplored. In this study, we characterized a novel maize kernel mutant (ubp5-ems#1) induced by ethyl methanesulfonate, which exhibited a smaller size, arrested embryo, smaller endosperm, and pigment deficiencies. Through bulked segregant analysis, we identified a stop-gained mutation in the ZmUBP5 gene as the cause of these phenotypic abnormalities. This was further validated by allelic confirmation and CRISPR/Cas9 knockout. ZmUBP5, identified as a ubiquitin-specific protease with deubiquitinase (DUB) activity, was localized to the nucleus, and the stop-gained mutation disrupted these functions. Protein interaction and degradation assays revealed that ZmUBP5 interacts with and may deubiquitinate ZmEMB140. The stop-gained mutations in ZmEMB140 resulted in smaller kernels with reduced embryo and endosperm size, as well as lethal seedlings. ZmEMB140 is likely a spliceosome-associated factor, interacting with six other proteins involved in pre-mRNA processing. Overall, this study underscores the critical roles of the DUB ZmUBP5 and its potential substrate ZmEMB140 in maize kernel development.