The Four Arabidopsis Choline/Ethanolamine Kinase Isozymes Play Distinct Roles in Metabolism and Development.

Ying-Chen Lin,Anh H Ngo,Artik Elisa Angkawijaya,Galileo Estopare Araguirang,Yuki Nakamura,Kui-Ting Lin

doi:10.1104/pp.19.01399

Abstract

Phosphatidylcholine and phosphatidylethanolamine are two major phospholipid classes in eukaryotes. Each biosynthesis pathway starts with the phosphorylation of choline (Cho) or ethanolamine (Etn) catalyzed by either choline or ethanolamine kinase (CEK). Arabidopsis contains four CEK isoforms, but their isozyme-specific roles in metabolism and development are poorly described. Here, we showed that these four CEKs have distinct substrate specificities in vitro. While CEK1 and CEK2 showed substrate preference for Cho over Etn, CEK3 and CEK4 had clear substrate specificity for Cho and Etn, respectively. In vivo, CEK1, CEK2, and CEK3 exhibited kinase activity for Cho but not Etn, although the latter two isoforms showed rather minor contributions to total Cho kinase activity in both shoots and roots. The knockout mutants of CEK2 and CEK3 both affected root growth, and these isoforms had nonoverlapping cell-type-specific expression patterns in the root meristematic zone. In-depth phenotype analysis, as well as chemical and genetic complementation, revealed that CEK3, a Cho-specific kinase, is involved in cell elongation during root development. Phylogenetic analysis of CEK orthologs in Brassicaceae species showed evolutionary divergence between Etn kinases and Cho kinases. Collectively, our results demonstrate the distinct roles of the four CEK isoforms in Cho/Etn metabolism and plant development.

Highlights

Phospholipids are the conserved lipid component of cellular membranes from bacteria to plants and animals
We investigated the transcript levels of 4 Arabidopsis choline/ethanolamine kinase (CEK) in various tissues by reverse transcription quantitative PCR (RT-qPCR) analysis using the cDNA template prepared from 6 different tissues (Fig. 1)
Since CEK2 and CEK3 function as the Cho kinases in vitro (Fig. 2) and in vivo (Fig. 3), we examined whether the expression patterns are overlapping with that of CEK1, which is preferentially expressed in vegetative tissues in root (Lin et al, 2019)

Summary

Introduction

Phospholipids are the conserved lipid component of cellular membranes from bacteria to plants and animals. Phosphatidylcholine (PC) and phosphatidylethanolamine (PE) are the most abundant phospholipid classes in eukaryotes. An initial reaction step for the synthesis of PC and PE is the phosphorylation of choline (Cho) or ethanolamine (Etn), which is catalyzed by choline/ethanolamine kinase (CEK) activity. CEK activity plays an important role in the biosynthesis of PC and PE. The CEK homologs have been identified and characterized in many organisms (Arlauckas et al, 2016; Glunde et al, 2011; Wu and Vance, 2010), their substrate specificity and tissue-specific roles have been a primary issue for decades (Ishidate et al, 1985a; Aoyama et al, 2004). Back in the 1950s, the first purified CEK from Brewer’s yeast demonstrated a dual substrate specificity that phosphorylates both

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Conclusion