Heterologous Expression and Functional Analysis of Rice GLUTAMATE RECEPTOR-LIKE Family Indicates its Role in Glutamate Triggered Calcium Flux in Rice Roots

Jun Ni, Zhiming Yu, Guankui Du, Yanyan Zhang, Jemma L. Taylor, Chenjia Shen, Jing Xu, Xunyan Liu, Yifeng Wang, Yunrong Wu

Tremendous progress has been made in understanding the functions of the GLUTAMATE RECEPTOR-LIKE (GLR) family in Arabidopsis. Still, the functions of OsGLRs in rice, especially the ion channel activities, are largely unknown. Using the aequorin-based luminescence imaging system, we screened the specificity of amino acids involved in the induction of Ca(2+) flux in rice roots. Of all the amino acids tested, glutamate (Glu) was the only one to trigger Ca(2+) flux significantly in rice roots. Detailed analysis showed a dose response of Ca(2+) increase to different concentrations of Glu. In addition, the Ca(2+) spike response to Glu was rapid, within 20 s after the application. A desensitization assay and pharmacological tests showed that the Glu-triggered Ca(2+) flux is mediated by OsGLRs. Whole genome analysis identified 13 OsGLR genes in rice, and these genes have various expression patterns in different tissues. Subcellular localization studies showed that all the OsGLRs examined are likely localized to the plasma membrane. Bacteria growth assays showed that at least OsGLR2.1 and OsGLR3.2 have the potential to mediate ion uptake in bacteria. Further analysis using Fura-2-based Ca(2+) imaging revealed a Glu-triggered Ca(2+) increase in OsGLR2.1-expressing human embryonic kidney (HEK) cells. Our work provides a molecular basis for investigating mechanisms of Glu-triggered Ca(2+) flux in rice.