Upstream Open Reading Frame Mediated Translation of WNK8 Is Required for ABA Response in Arabidopsis.

Zhiyong Li,Jinyu Shen,Jiansheng Liang,Yajuan Fu

doi:10.3390/ijms221910683

Abstract

With no lysine (K) (WNK) kinases comprise a family of serine/threonine kinases belonging to an evolutionary branch of the eukaryotic kinome. These special kinases contain a unique active site and are found in a wide range of eukaryotes. The model plant Arabidopsis has been reported to have 11 WNK members, of which WNK8 functions as a negative regulator of abscisic acid (ABA) signaling. Here, we found that the expression of WNK8 is post-transcriptionally regulated through an upstream open reading frame (uORF) found in its 5′ untranslated region (5′-UTR). This uORF has been predicted to encode a conserved peptide named CPuORF58 in both monocotyledons and dicotyledons. The analysis of the published ribosome footprinting studies and the study of the frameshift CPuORF58 peptide with altered repression capability suggested that this uORF causes ribosome stalling. Plants transformed with the native WNK8 promoter driving WNK8 expression were comparable with wild-type plants, whereas the plants transformed with a similar construct with mutated CPuORF58 start codon were less sensitive to ABA. In addition, WNK8 and its downstream target RACK1 were found to synergistically coordinate ABA signaling rather than antagonistically modulating glucose response and flowering in plants. Collectively, these results suggest that the WNK8 expression must be tightly regulated to fulfill the demands of ABA response in plants.

Highlights

Protein kinases comprise a superfamily of enzymes that regulate a wide range of cellular processes through induced phosphorylation of downstream protein substrates in all living cells
We showed that the translation of WNK8 was repressed by CPuORF58 in vivo and that this element may act by a ribosome stalling mechanism, independently of the main open reading frame downstream of the upstream open reading frame (uORF)
The results showed that weak GUS expression was detected in the plants transformed with pWNK8:GUS, whereas a strong GUS color was detected in all selected tissues and organs of the plants transformed with pWNK8m:GUS (Figure 3A)

Summary

Introduction

Protein kinases comprise a superfamily of enzymes that regulate a wide range of cellular processes through induced phosphorylation of downstream protein substrates in all living cells. WITH NO LYSINE (WNK) kinases belong to a superfamily of serine/threonine protein kinases uniquely characterized by the lack of a catalytic lysine residue in the kinase subdomain II, which is crucial for coordinating ATP and catalyzing phosphoryl transfers within other protein kinase superfamily and downstream targets [1]. WNK kinases have been identified in various eukaryotes. They were first discovered in mammals while screening for new members of the mitogen-activated protein, serine/threonine kinase (MAPK) family. A total of 11 and 9 WNK members have been identified in the model plant Arabidopsis and Oryza sativa, respectively [13,14]

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Conclusion