Molecular characterization and functional analysis of Glycine max sterol methyl transferase 2 genes involved in plant membrane sterol biosynthesis

Abstract

Sterol C24 methyltransferase (SMT2) genes governing the pattern of phytosterols synthesized in higher plants have been studied in Glycine seedlings and wild-type and engineered Arabidopsis thaliana plants. The SMT2 genes of soybean (SMT2-1 and SMT2-2) previously cloned and characterized (Neelakandan et al. 2009) were shown to complement the SMT deficient cvp1 mutant Arabidopsis plants, consistent with their role in regulation of 24-alkyl sterol-controlled plant physiology. Further analysis of these genes showed that environmental cues, including dehydration, cold, and abscisic acid induced differential changes in transcript levels of the SMT2 during soybean seedling growth. Sterol analyses of transgenic Arabidopsis seeds originating in variant constructs of AtHMGR1, GmSMT1, and GmSMT2 engineered in seeds showed relevant modifications in the ratio of 24-methyl to 24-ethyl sterol in the direction of sitosterol formation. To provide insight into the structural features of the sterol gene that affects transcript regulation, the upstream promoter sequences of soybean SMT2 genes were cloned and characterized. Sequence analysis revealed several important cis-elements and transcription factor binding sites. The analysis of promoter-GUS fusions in transgenic Arabidopsis plants revealed shared and distinct expression features in different developmental stages and tissues. The data are interpreted to imply that SMT2 is an important contributor to normal plant growth and development.

Arabidopsis ribosomal proteins control vacuole trafficking and developmental programs through the regulation of lipid metabolism

“…The SMT2 mutant displayed developmental defects, including aberrant cotyledon vein patterning, serrated petals, and reduced stature. The mutants also affected auxin response, demonstrated by reduced auxin sensitivity, enhanced axr1 auxin resistance, and ectopically expressed DR5:GUS signal (40,56). For the fatty acid pathway, we chose β-Ketoacyl-[acyl carrier protein] synthase I (KASI) (At5g46290), which catalyzes the elongation of de novo fatty acid synthesis.…”

Evaluation of Candidate Reference Genes for Normalization of Quantitative RT-PCR in Soybean Tissues under Various Abiotic Stress Conditions

“…Abiotic stressors are the adverse environmental conditions which are unfavorable to plant growth and include such circumstances as flooding, extreme temperatures, high soil salinity, and drought. These stressors can have detrimental effects on plants which generally result in major yield losses for the economically important crops, including soybean [1]–[5]. Understanding the mechanisms plants use to cope with such stresses is crucial for areas of research aimed at the engineering of soybean cultivars with increased stress tolerance [6]–[9].…”

Characterization, mutagenesis and mechanistic analysis of an ancient algal sterol C24-methyltransferase: Implications for understanding sterol evolution in the green lineage

“…Interestingly, red algae typically synthesize cholesterol as their dominant sterol (Beastall et al, 1974; Kodner et al, 2008), and cholesterol is often present in variable amounts in other organisms as diverse as the oomycetous fungi and humans (Nes, 2011; Weete et al, 2010), suggesting a down-regulation or loss of SMT activity from these lineages. Therefore, it is evident that retrograde evolution of SMT could be class-specific, and may have arisen in select lineages due to loss of promoters in the cis -sequence motifs located upstream of the SMT coding sequence (Neelakandan et al, 2010). Thus, directed mutations of regulatory and transcribed regions associated with the SMT gene may be responsible for shifts in sterol biosynthesis pathways which in the lineage generating vertebrate animals led to coupled loss in phytosterols with a gain in cholesterol production.…”