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Expression and evolutionary features of the hexokinase gene family in Arabidopsis

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Abstract

Arabidopsis hexokinase1 (HXK1) is a moonlighting protein that has separable functions in glucose signaling and in glucose metabolism. In this study, we have characterized expression features and glucose phosphorylation activities of the six HXK gene family members in Arabidopsis thaliana. Three of the genes encode catalytically active proteins, including a stromal-localized HXK3 protein that is expressed mostly in sink organs. We also show that three of the genes encode hexokinase-like (HKL) proteins, which are about 50% identical to AtHXK1, but do not phosphorylate glucose or fructose. Expression studies indicate that both HKL1 and HKL2 transcripts occur in most, if not all, plant tissues and that both proteins are targeted within cells to mitochondria. The HKL1 and HKL2 proteins have 6–10 amino acid insertions/deletions (indels) at the adenosine binding domain. In contrast, HKL3 transcript was detected only in flowers, the protein lacks the noted indels, and the protein has many other amino acid changes that might compromise its ability even to bind glucose or ATP. Activity measurements of HXKs modified by site-directed mutagenesis suggest that the lack of catalytic activities in the HKL proteins might be attributed to any of numerous existing changes. Sliding windows analyses of coding sequences in A. thaliana and A. lyrata ssp. lyrata revealed a differential accumulation of nonsynonymous changes within exon 8 of both HKL1 and HXK3 orthologs. We further discuss the possibility that the non-catalytic HKL proteins have regulatory functions instead of catalytic functions.

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Abbreviations

GFP:

Green fluorescent protein

HA:

Hemagglutinin

HKL:

Hexokinase-like

HXK:

Hexokinase

Indels:

Insertions/deletions

K A :

Number of nonsynonymous substitutions per nonsynonymous site

K S :

Number of synonymous substitutions per synonymous site

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Acknowledgments

We appreciate technical assistance from Ms. Betsy Metters. We thank also Dr. Amy Lawton-Rauh for consultation on nucleotide sequence analyses and for supplying A. lyrata tissue and DNA. This work was supported by the U.S. Department of Agriculture-National Research Initiative Plant Biochemistry Program (grant no. 2001-035318) and the South Carolina Agricultural Experiment Station (technical contribution No. 5389 of the Clemson University Experiment Station). This material is based upon work supported by CSREES/USDA, under project number SC-170090.

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Authors and Affiliations

  1. Department of Genetics and Biochemistry, Clemson University, Clemson, SC, 29634, USA

    Abhijit Karve, Bradley L. Rauh, Xiaoxia Xia & Brandon d. Moore

  2. Genetics Department, University of Georgia, Athens, GA, 30602, USA

    Muthugapatti Kandasamy & Richard B. Meagher

  3. Department of Molecular Biology, Massachusetts General Hospital, Boston, MA, 02114, USA

    Jen Sheen

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  1. Abhijit Karve
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Correspondence to Brandon d. Moore.

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425_2008_746_MOESM1_ESM.tif

Supplemental Fig. 1 Sequence analyses of HXK family genes from A. lyrata. a Illustrations of gene structures were determined from genomic sequence data. Exons are indicated by rectangles, introns by lines. The lengths of both are proportional to the number of nucleotides that are present. b Phylogenetic relationships of predicted HXK family proteins from A. thaliana and A. lyrata. Phylogenetic reconstruction was done using MEGA4 (Tamura et al. 2007). The scale bar = the frequency of amino acid substitutions per unit length (TIFF 3321 kb)

425_2008_746_MOESM2_ESM.tif

Supplemental Fig. 2 Amino acid sequence alignment for predicted HXK family proteins from A. thaliana and A. lyrata. Sequences were aligned using ClustalW in BioEdit (Hall 1999). (TIFF 5546 kb)

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Karve, A., Rauh, B.L., Xia, X. et al. Expression and evolutionary features of the hexokinase gene family in Arabidopsis . Planta 228, 411–425 (2008). https://doi.org/10.1007/s00425-008-0746-9

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