Molecular and genetic approaches to elucidate the role of cis-phenylpropanoid acid during plant growth and development

Phenylpropanoid acid / cinnamic acid (CA) is the first product of the phenylpropanoic pathway. Naturally occurring CA has both cis- and trans-isoforms which has been successfully isolated from sunlight grown Arabidopsis, Brassica and rice. Conversion between cis and trans isomers is mediated by the UV light. The ratio of cis-/ trans- in plant can reach 1/1. Physiological experiment found that cis-CA has biological activity and can cause several plant physiological responses, while trans- form is relatively inactive.To study CA’s functions, two strategies have been employed. The first is a proteomic approach, by which LC-MS/MS is used to find the proteins responsive to CA treatment. Two-week light grown seedlings with or without 50 M mixture of CA (1/1 cis-/ trans-) treatment were collected, and total protein was extracted. Following a multidimensional separation of proteins, the trypsin-digested peptides were analyzed by LC-MS/MS. More than a thousand of proteins were detected form both samples. Total 32 CA-enhanced and 13 CA-repressible proteins were obtained. Transcriptional level of these genes was examined using RT-PCR. Two genes encoding Major Latex Protein (MLP) like proteins were CA-inducible, especially cis-CA induced. They were also up-regulated by other trans- or cis-phenylpropanoids but not as high as cis-CA. In some Arabidopsis mutants, gene induction by cis-CA still could be detected although the folds varied. One of the MLP like proteins—At2g01520 exists in cytosol. Knock out the MLP like protein genes makes plant relatively insensitive under 50 M cis-CA treatment. Secondly, a genetic screening for Arabidopsis mutants resistant to cis-CA was performed using root growth inhibition assay. Four Arabidopsis mutants, named cpar (c̱is-p̱henylpropanoid a̱cid-ṟesponse) 1, 2, 3 and 4 were successfully isolated. All cpar mutants showed larger seeds. Besides, cpar1 showed delayed senescence under CA treatment when grown in soil. Map-based cloning of cpar1 indicated that this mutation locates on chromosome 4