First published online April 19, 2002; 10.1104/pp.001776
Plant Physiol, May 2002, Vol. 129, pp. 363-373
Molecular and Biochemical Characterization of a
Cold-Regulated Phosphoethanolamine
N-Methyltransferase from Wheat1
Jean-Benoit Frenette
Charron,
Ghislain
Breton,
Jean
Danyluk,
Ingrid
Muzac,
Ragai K.
Ibrahim, and
Fathey
Sarhan*
Département des Sciences Biologiques, Université du
Québec à Montréal, Case Postale 8888, Succursale "Centre-Ville," Montréal, Québec,
Canada H3C 3P8 (J.-B.F.C., G.B., J.D., F.S.); and Plant Biochemistry
Laboratory, Department of Biology, Concordia University, 1455 De
Maisonneuve Boulevard West, Montréal, Québec, Canada H3G
1M8 (I.M., R.K.I.)
A cDNA that encodes a methyltransferase (MT) was cloned from
a cold-acclimated wheat (Triticum aestivum) cDNA
library. Molecular analysis indicated that the enzyme WPEAMT
(wheat phosphoethanolamine [P-EA] MT) is a bipartite protein with
two separate sets of
S-adenosyl-L-Met-binding domains, one close
to the N-terminal end and the second close to the C-terminal end. The
recombinant protein was found to catalyze the three sequential
methylations of P-EA to form phosphocholine, a key precursor for the
synthesis of phosphatidylcholine and glycine betaine in plants.
Deletion and mutation analyses of the two
S-adenosyl-L-Met-binding domains indicated
that the N-terminal domain could perform the three
N-methylation steps transforming P-EA to phosphocholine. This is in contrast to the MT from spinach (Spinacia
oleracea), suggesting a different functional evolution for the
monocot enzyme. The truncated C-terminal and the N-terminal mutated
enzyme were only able to methylate phosphomonomethylethanolamine and
phosphodimethylethanolamine, but not P-EA. This may suggest that the
C-terminal part is involved in regulating the rate and the equilibrium
of the three methylation steps. Northern and western analyses
demonstrated that both Wpeamt transcript and the
corresponding protein are up-regulated during cold acclimation. This
accumulation was associated with an increase in enzyme activity,
suggesting that the higher activity is due to de novo protein
synthesis. The role of this enzyme during cold acclimation and the
development of freezing tolerance are discussed.
1
This work was supported by the Natural Sciences
and Engineering Research Council of Canada and Fonds pour la Formation
de Chercheurs et l'Aide à la Recherche (research grants to F.S. and R.K.I.).
*
Corresponding author; e-mail sarhan.fathey{at}uqam.ca; fax
514-987-4647.
© 2002 American Society of Plant Physiologists
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