Professor M. L. Chye

• Research Team
• Research Interests
  • Investigations on Arabidopsis acyl-CoA binding proteins
  • Role of HMGS in plant isoprenoid metabolism
  • Expression of siRNAs and heterologous proteins in transgenic plants
• Representative publications

Research Team

 

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Investigations on Arabidopsis acyl-CoA binding proteins

The focus of our research is on a new family of acyl-CoA-binding proteins (ACBPs) which bind acyl-CoA esters and transport them within the plant cell. We have shown that in the model plant Arabidopsis, six genes encode four structurally distinct classes of ACBPs (Leung et al., 2004). These classes are (i) cytoslic ACBPs which is the only class identified and characterized in other eukaryotes, (ii) membrane-associated ACBPs with ankyrin repeats, ACBP1 and ACBP2 (Chye et al., 1999; Li and Chye, 2003; 2004), (iii) ACBP3 (Leung et al., 2006) and (iv) kelch-motif containing ACBP4 and ACBP5 (Leung et al., 2004). We have identified the amino acid residues in the acyl-CoA-binding domain that are critical in binding acyl-CoA esters (Chye et al., 2000; Leung et al., 2004; 2006). A variation in their binding affinities to acyl-CoA esters suggests that each ACBP has a unique role in plant lipid metabolism. Further, ACBPs with ankyrin repeats (Li and Chye, 2004) and kelch motifs (Leung et al., 2004) can potentially mediate protein-protein interactions.

Role of HMGS in plant isoprenoid metabolism

We are investigating the role of 3-hydroxy-3-methylglutaryl-CoA synthase (HMGS) in plant isoprenoid metabolism. HMGS is an enzyme in the cytosolic mevalonate pathway which produces sterols, sesquiterpenes and polyterpenes. Its expression is stress-inducible and is highest during early development in flower, seed and seedling (Alex et al., 1999). Four isogenes encoding HMGS are differentially expressed in Brassica juncea (Nagegowda et al., 2005). Using green fluorescent protein fusions, B. juncea HMGS1 (BjHMGS) has been subcellularly localized to the cytosol (Nagegowda et al., 2005). We have biochemically purified and characterized His-tagged recombinant BjHMGS expressed in Escherichia coli, presenting a first detailed characterization of a plant HMGS and the amino acids involved in catalysis were identified by site-directed mutagenesis (Nagegowda et al., 2004).

Some of the approaches currently used to engineer plants to disease-resistance and pest tolerance exploit the natural defense mechanisms evolved in the plant in response to invasion. We are interested in isolating such genes from tropical plants for expression in transgenic crops. To this end we have cloned and characterized cDNAs encoding beta-1, 3-glucanase from Hevea brasiliensis (Chye and Cheung, 1995, Plant Mol Biol 29: 347-402) and an unusual Brassica juncea chitinase with two chitin-binding domains, designated BjCHI1 (Zhao and Chye, 1999; Fung et al., 2002). Potato transgenic for both proteins was protected from Rhizoctonia solani invasion (Chye et al., 2005). Investigations on anti-fungal BjCHI1 showed that its two chitin-binding domains confer agglutinating properties (Tang et al., 2004). BjCHI1-susceptible phytopathogens are currently being identified for further applications in the generation of anti-fungal transgenic crops. Research on proteinase inhibitor protein SaPIN2a from a weed, Solanum americanum (Xu et al., 2001), has not only led to the production of transgenic lettuce that are insect-resistant but that also show inhibition of endogenous trypsin- and chymotrysin-like activities (Xu et al., 2004). Investigations on SaPIN2a and SaPIN2b expression suggest that they have endogenous functions in inhibiting proteinase activities in phloem and floral development (Xu et al., 2001; Sin and Chye, 2004). We have used RNAi-based gene silencing to demonstrate that SaPIN2a and SaPIN2b are essential for seed development (Sin et al., 2006). A reduction in seed set due to seed abortion was observed in PIN2-RNAi transgenic S. americanum lines. Aborted seeds in transgenic fruits had an abnormal endothelium. The anomalous expansion of the endothelium prevented proper endosperm and embryo development, leading to seed abortion, hence suggesting that the endothelium may protect the embryo sac, allowing proper endosperm and embryo formation, through its ability to produce a proteinase inhibitor (Sin et al., 2006).

• FX Xu and ML Chye. 1999. Expression of cysteine proteinase during developmental events associated with programmed cell death in brinjal. Plant Journal 17: 321-327.
• ML Chye, BQ Huang and SY Zee. 1999. Isolation of a gene encoding Arabidopsis membrane-associated acyl-CoA binding protein and immunolocalization of its gene product. Plant Journal 18: 205-214.
• KJ Zhao and ML Chye. 1999. Methyl jasmonate induces expression of a Brassica juncea chitinase with two chitin-binding domains. Plant Mol Biol 40: 1009-1018.

• D Alex, TJ Bach and ML Chye. 2000. Expression of Brassica juncea 3-hydroxy-3-methylglutaryl-CoA synthase is developmentally regulated and stress-responsive. Plant Journal 22: 415-426.
• ML Chye, HY Li and MH Yung. 2000. Single amino acids substitutions at the acyl-CoA-binding domain interrupt ¹⁴[C]palmitoyl-CoA binding of ACBP2, an Arabidopsis acyl-CoA-binding protein with ankyrin repeats. Plant Mol Biol: 44: 711-721.

• ZF Xu, WQ Qi, XZ Ouyang, E Yeung and ML Chye. 2001. A proteinase inhibitor II of Solanum americanum is expressed in phloem. Plant Mol Biol 47: 727-738.

• KL Fung, KJ Zhao, ZM He and ML Chye. 2002. Tobacco-expressed Brassica juncea chitinase BjCHI1 shows antifungal activity in vitro. Plant Mol Biol 50: 283-294.

• ZF Xu, ML Chye, HY Li, FX Xu and KM Yao. 2003. G-box binding coincides with increased S. melongena cysteine proteinase expression in senescent fruits and circadian-regulated leaves. Plant Mol Biol 51: 9-19.
• HY Li and ML Chye. 2003. Membrane localization of Arabidopsis acyl-CoA-binding protein ACBP2. Plant Mol Biol 51: 483-492.

• ZF Xu, WL Teng and ML Chye. 2004. Inhibition of endogenous trypsin- and chymotrypsin-like activities in transgenic lettuce expressing heterogeneous proteinase inhibitor SaPIN2a. Planta 218: 623-629.
• HY Li and ML Chye. 2004. Arabidopsis acyl-CoA-binding protein ACBP2 interacts with an ethylene-responsive element binding protein AtEBP via its ankyrin repeats. Plant Mol Biol 54: 233-243.
• KC Leung, HY Li, G Mishra and ML Chye. 2004. ACBP4 and ACBP5, novel Arabidopsis acyl-CoA-binding proteins with kelch motifs that bind oleoyl-CoA. Plant Mol Biol 55: 297-309.
• SF Sin and ML Chye. 2004. Expression of proteinase inhibitor II proteins during floral development in Solanum americanum flowers. Planta 219: 1010-1022.
• D Nagegowda, TJ Bach and ML Chye. 2004. Brassica juncea HMG-CoA synthase 1: expression and characterization of recombinant wild-type and mutant enzymes. Biochem J 383: 517-527.
• CM Tang, ML Chye, S Ramalingam, SW Ouyang, KJ Zhao, W Ubhayasekera and S Mowbray. 2004. Functional analysis of the chitin-binding domains and the catalytic domain of Brassica juncea chitinase BjCHI1. Plant Mol Biol 56: 285-298.

• ML Chye, KJ Zhao, ZM He, S Ramalingam and KL Fung. 2005. Expression of an agglutinating chitinase BjCHI1 with two chitin-binding domains is R. solani-inducible and confers fungal protection in transgenic potato. Planta 220: 717-730.
• D Nagegowda, S Ramalingam, A Hemmerlin, TJ Bach and ML Chye. 2005. Brassica juncea HMG-CoA synthase: localization of mRNA and protein. Planta 221: 844-856.
• R Rawat, ZF Xu, KM Yao and ML Chye. 2005. Identification of cis-elements in ethylene and circadian regulation of gene encoding Solanum melongena cysteine proteinase. Plant Mol. Biol. 57: 629-643.

• KC Leung, HY Li, S Xiao, MH Tse, ML Chye. 2006. Arabidopsis ACBP3 is an extracellularly targeted acyl-CoA-binding protein. Planta 223: 871-881.
• SF Sin, EC Yeung, ML Chye. 2006. Down-regulation of Solanum americanum genes encoding proteinase inhibitor II causes defective seed development. Plant Journal 46: 58-70.
• HY Li, S Ramalingam and ML Chye. 2006. Production of the SARS-CoV spike protein in tobacco cytosol and chloroplasts. Expt. Biol. Medicine 231: 1346-1352.
• F Pojer, JL Ferrer, SB Richard, DA Nagegowda, ML Chye, TJ Bach and J P Noel. 2006. Structural basis for the design of potent and species specific inhibitors of 3-hydroxy-3-methylglutaryl-coenzyme A synthases. Proc. Natl. Acad. Sci. USA 103: 11491-11496.

Last modified: 30 August, 2007.

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