Classification of Receptor-Like Cytoplasmic Kinases in Maize and Functional Analysis of ZmBLK1

Receptor-like cytoplasmic kinases (RLCKs) form a large family of proteins in plants. RLCKs have been found in different plant species, regulating plant immunity to different bacterial and fungal pathogens. Previous studies implicated <i>Arabidopsis</i> <i>botrytis induced kinase1 (BIK1)</i> and <i>tomato protein kinase 1b (TPK1b)</i> in plant resistance to <i>Pseudomonas syringae</i> and <i>Botrytis cinerea</i>. In this study, we classified 195 putative maize RLCKs into ten subfamilies. Based on the amino acid sequence similarity to BIK1 and TPK1b, a novel maize RLCK,<i> zea mays bik1-like kinase 1 (ZmBLK1)</i> was identified. Enzyme assays with cloned <i>ZmBLK1</i> revealed a functional kinase when expressed in planta. The recombinant protein located to the plasma membrane. Expression of <i>ZmBLK1</i> is highest in maize leaves compared to other structures at silking stage. Expression of the recombinant <i>ZmBLK1</i> significantly reduced the rate of lesion spread in maize leaves inoculated with the Goss’s wilt pathogen. In maize kernels, expression of <i>ZmBLK1</i> increases during kernel maturation. Kernels from transgenic maize overexpressing <i>ZmBLK1</i> were not resistant to <i>Aspergillus flavus</i> or to aflatoxin contamination. In addition, mutations were made in <i>ZmBLK1</i> that were hypothesized to create a constitutively active kinase. However, resulting proteins had similar activity to the wild-type ZmBLK1 and transgenic plants showed similar responses to the Goss’s wilt and Aspergillus ear rot pathogens. Overall, this research established the first characterization of RLCKs in maize and described a potential contribution of ZmBLK1 to maize immune responses.