Functions Of Arabidopsis PERK4 In ABA-Regulated Root Growth

The plant hormone abscisic acid (ABA) affects several important physiological processes in plants such as seed germination, stomatal aperture and root development. Although many genes that function in ABA signal transduction pathways have been identified and play important role in the ABA signaling pathway(Li et al., 2000; Lemichez et al., 2001; McCourt, 2001; Sang et al., 2001; Finkelstein et al., 2002; Guo et al., 2002; Mustilli et al., 2002; Yoshida et al., 2002; Zheng et al., 2002; Coursol et al., 2003); rather little is know about how changes in gene expression are involved in the control of ABA related root growth. In Arabidopsis, ABA block lateral root formation and inhibit early lateral root development have been studied (Smet et al., 2003). But how ABA inhibits radicle elongation in Arabidopsis is still poorly understood. A link between the effect of ABA on restraining growth of primary roots and the signal transduction pathway remains to be established.In our previous studies of AtERF7 overexpression line, we found that the expression of one proline-rich extensin like receptor kinase family member, PERK4, was highly regulated (data not shown). Previous researches showed that The Arabidopsis PERK family shares sequence similarity with plant cell wall-associated proteins (Silva and Goring 2002), indicated that PERK-based signaling cascades might mainly function in perceiving and signaling changes to the cell wall occurring in plant growth and development (Nakhamchik et al., 2004). However, little is known about the function of the members of the PERK family in ABA signal transduction.We show here a detail analysis of Arabidopsis PERK4 (accession number: At2g18470). The expression of the AtPERK4 gene was investigated by RT-PCR analysis, and only ABA significantly induced AtPERK4's mRNA accumulated. In addition, strong GUS expression was detected in the roots, flowers and the juvenile leaves of GUS-AtPERK4 transgenic plants.We obtained T-DNA insertion line for the At2gl8470, Salk₀34666, from the Salk Institute Genomic Analysis laboratory Collection. Homozygous mutant plants were screened by PCR amplification according to the method provided by http://signal.salk.edu. After establishing the presence of homozygote—perk4-1, we backcrossed the knockout plants and analyzed the F2 progeny. Useing X² test, we confirmed At2g18470 was recessive and had a single insertion of T-DNA. perk4-l markedly decrease sensitivity specificity to ABA in primary root growth after treated with various plant hormone. To further test it is the knockout of AtPERK4 responsible for the phenotype, the wild-type AtPERK4 cDNA was inserted under the control of the superpromoter, and then transformed to the perk4-l plants. The transgenic plants recovered sensitivity of root growth to ABA, just as the wild-type. The expression pattern at the cellular level using GFP-AtPERK4 transgenic plants shows the localization of PERK4 on the plasma membrane, and the PERK4 protein expressed with yeast showed kinase activity phosphorylating itself and the substrate.The root tip morphologies of the WT and the perk4-1 showed dramatical difference in the root cell length, the perk4-l root cell is longer than that of the WT. QRT-PCR analysis showed that AtPERK4 regulate root cell elongation related genes in plant to restrain the root cell growth; the upregulated H₂O₂-induced gene AtOXII suggests H₂O₂ may participate in the ABA signaling transduction pathway in restraining root growth.Taken together, our data provide evidence that AtPERK4 encodes a receptor like kinase essential in...