Identification of ZmRBOHD1 interacting with ZmCDPK32 in maize

Both respiratory burst oxidase homologue (RBOH) and calcium dependent protein kinase (CDPK) regulate pollen tube polar growth, but whether these two proteins interact remains unclear． To explore the mechanism of ZmRBOH and ZmCDPK synergistic regulation of pollen tube polar growth, 14 ZmRBOH family genes we identified from the maize genome, temporal and spatial expression patterns of ZmRBOH genes were examined from transcriptome data． Expression analyses found that ZmRBOHD1 and ZmRBOHD2 were specifically expressed in mature maize pollen． The interaction between pollen specifically expressed ZmRBOHD1 and a known pollen tube polar growth regulator, ZmCDPK32, was investigated． Transient expression of ZmRBOHD1-GFP fusion protein demonstrated that ZmRBOHD1 was localized to cell membranes． Yeast two-hybrid and bimolecular fluorescence complementation techniques confirmed that ZmRBOHD1 interacted with ZmCDPK32． Bioinformatics analysis revealed potential CDPK phosphorylation motifs in ZmRBOHD1． The above data together suggest that ZmCDPK32 synergically regulates pollen tube development in maize by interacting with ZmRBOHD1． Further study of the molecular mechanism of pollen tube polar growth may be needed．