Plant Molecular Biology 34: 891–896, 1997.
1997 Kluwer Academic Publishers. Printed in Belgium.
Characterization of Zea mays endosperm C-24 sterol methyltransferase: one
of two types of sterol methyltransferase in higher plants
Robert J. Grebenok
, David W. Galbraith and Dean Della Penna
Department of Plant Sciences, University of Arizona, Tucson, AZ 85721, USA; Current address:
Biology, Canisius College, Buffalo, NY 14208, USA (
author for correspondence);
Department of Biochemistry,
University of Nevada-Reno, Reno, NV 89557, USA
Received 5 December 1996; accepted in revised form 28 April 1997
Key words: cloning, methyltransferase, phytosterol, yeast complementation, Zea mays
We report the characterization of a higher-plant C-24 sterol methyltransferase by yeast complementation. A Zea
mays endosperm expressed sequence tag (EST) was identiﬁed which, upon complete sequencing, showed 46%
identity to the yeast C-24 methyltransferase gene (ERG6) and 75% and 37% amino acid identity to recently isolated
higher-plant sterol methyltransferases from soybean and Arabidopsis, respectively. When placed under GAL4
regulation, the Z. mays cDNA functionally complemented the erg6 mutation, restoring ergosterol production and
conferring resistance to cycloheximide. Complementation was both plasmid-dependent and galactose-inducible.
The Z. mays cDNA clone contains an open reading frame encoding a 40 kDa protein containing motifs common
to a large number of S-adenosyl-L-methioninemethyltransferases (SMTs). Sequence comparisons and functional
studies of the maize, soybean and Arabidopsis cDNAs indicates two types of C-24 SMTs exist in higher plants.
In contrast to the situation in yeast, for which the path-
molecular level, in higher plants much less is known,
characterized. In higher plants the dominant sterols
contain C-24 alkyl additions, while the unmethylated
sterols such as cholesterol are present at low levels.
Production of the alkylated sterols in plants, after cyc-
lization of squalene, involves severalstages predicated
on the alkylation of the double bond at carbon 24 in
the sterol side chain , catalyzed by an S-adenosyl-
L-methionine methyltransferase (SMT). This step is
suggested to be a major site of regulation of carbon
ﬂux throughthe sterol biosynthetic pathway(Figure 1)
In higher plants and yeast, the SMT enzymes have
GenBank and DDBJ Nucleotide Sequence Databases, under the
accession number U79669.
reticulum of higher plant cells [2, 11]. The sunﬂower
SMTactivity, whichhas beencharacterizedin the most
detail , can methylate a large number of steroid
substrates, with varyingdegreesof catalytic efﬁciency.
Mechanistically, the nucleophiliccenters present at the
C-3 hydroxyl and the C-24 double bond are obligatory
for substrate binding and methylation.
Recent molecular genetic investigations of SMT
activity in higher plants [8, 14] have provided cDNAs
for sterol methyltransferases from soybean and Ara-
bidopsis, respectively. The two plant SMTs are highly
divergent with the soybean sequence having much
greater sequence similarity to the yeast ERG6 sterol
methyltransferase than does the Arabidopsis thaliana
cDNA. The reported sequence similarity between the
soybean cDNA, SMT1, and the yeast ERG6 cDNA
supports the ERG6 as a structural gene encoding a
SMT enzyme in yeast [6, 8, 14].
We present the isolation of a third plant C-24 ster-
ol methyltransferase from Zea mays and its function-
al characterization by yeast complementation. The
GR: 201001984, Pips nr. 140600 BIO2KAP
pla410us.tex; 16/07/1997; 16:02; v.7; p.1