Structure and nucleotide sequence of tomato HMG2 encoding 3-hydroxy-3-methyl-glutaryl coenzyme A reductase.

Abstract

The enzyme 3-hydroxy-3-methylglutaryl coenzymeA reductase (HMGR; EC 1.1.1.34) catalyzes the conversion of hydroxymethylglutarylCoA to mevalonic acid, the proposed rate-limiting step in isoprenoid biosynthesis. HMGR regulation has been widely studied in mammalian systems due to its critical role in cholesterol biosynthesis. Plants utilize a broad array of isoprenoid compounds critical not only for sterol biosynthesis but also for growth and development (gibberellins, abscisic acid, cytokinins, steroids), disease resistance (phytoalexins), photosynthesis (plastiquinone, chlorophyll, carotenoids) and production of specialized isoprenoids associated with insect attraction, fragrance, flavor, feeding deterrents and allelopathy. Genes or cDNA sequences encoding HMGR have been reported for several plant species including Arabidopsis [3, 9], tomato [13], potato [14], radish [7], and Hevea rubber tree [5]. HMGR is encoded by a small multigene family in plants [3, 5, 14, 16] unlike animal systems where a single HMGR has been identified [4, 8, 10]. Genomic Southern analyses suggest that tomato contains at least four HMGR genes (H. Park and C.L. Cramer, unpublished results). We report here the sequence of one of these genes, HMG2. HMG2 mRNA levels are elevated in response to wounding or fungal elicitors suggesting that HMG2 is a defense-related gene ([16], H. Park et al., manuscript in preparation). A tomato genomic library (Lycopersicon esculentum cv. VFNT Cherry DNA in lambda Charon 35) was screened using a 1.75 kb Eco RI fragment of pJR326, encoding the region of yeast (Saccharomyces cerevisiae) HMG1 most highly conserved between species [1]. Low-stringency hybridization conditions (30~o formamide, 6 x SSC, 42 °C) were used and several crosshybridizing plaques were isolated. A 7 kb Hind III fragment of one clone was inserted into the Hind III site of vector pSP6/T7 ([ 12], BRL) and designated pTH295. Southern blots of total tomato DNA probed with HMGR-encoding regions of pTH295 (data not shown) indicated that the HMGR gene cloned in pTH295 is distinct from tomato HMG1 identified as a partial cDNA sequence by Narita and Gruissem [13]. Tomato HMG2 nucleotide and derived amino acid se-