Abstract
Background Landoltia punctata can be used as renewable and sustainable biofuel feedstock because it can quickly accumulate high starch levels. ADP-glucose pyrophosphorylase (AGPase) catalyzes the first committed step during starch biosynthesis in higher plants. The heterotetrameric structure of plant AGPases comprises pairs of large subunits (LSs) and small subunits (SSs). Although several studies have reported on the high starch accumulation capacity of duckweed, no study has explored the underlying molecular accumulation mechanisms and their linkage with AGPase. Therefore, this study focused on characterizing the roles of different L. punctate AGPases. Methodology. Expression patterns of LpAGPs were determined through comparative transcriptome analyses, followed by coexpressing their coding sequences in Escherichia coli, Saccharomyces cerevisiae, Arabidopsis thaliana, and Nicotiana tabacum. Results Comparative transcriptome analyses showed that there are five AGPase subunits encoding cDNAs in L. punctata (LpAGPS1, LpAGPS2, LpAGPL1, LpAGPL2, and LpAGPL3). Nutrient starvation (distilled water treatment) significantly upregulated the expression of LpAGPS1, LpAGPL2, and LpAGPL3. Coexpression of LpAGPSs and LpAGPLs in Escherichia coli generated six heterotetramers, but only four (LpAGPS1/LpAGPL3, LpAGPS2/LpAGPL1, LpAGPS2/LpAGPL2, and LpAGPS2/LpAGPL3) exhibited AGPase activities and displayed a brownish coloration upon exposure to iodine staining. Yeast two-hybrid and bimolecular fluorescence complementation (BiFC) assays validated the interactions between LpAGPS1/LpAGPL2, LpAGPS1/LpAGPL3, LpAGPS2/LpAGPL1, LpAGPS2/LpAGPL2, and LpAGPS2/LpAGPL3. All the five LpAGPs were fusion-expressed with hGFP in Arabidopsis protoplasts, and their green fluorescence signals were uniformly localized in the chloroplast, indicating that they are plastid proteins. Conclusions This study uncovered the cDNA sequences, structures, subunit interactions, expression patterns, and subcellular localization of AGPase. Collectively, these findings provide new insights into the molecular mechanism of fast starch accumulation in L. punctata.
Highlights
Duckweed is a general term for plants belonging to the Lemnaceae family, comprising five genera (Spirodela, Landoltia, Lemna, Wolffiella, and Wolffia) and more than 40 species [1]
Comparative transcriptome analysis was performed in a previous study to investigate the high starch accumulation of L. punctata under nutrient starvation [11]
It was shown that ADP-glucose pyrophosphorylase (AGPase) large subunits (LSs) are encoded by three genes (LpAGPL1, LpAGPL2, and LpAGPL3), while small subunits (SSs) are encoded by two genes (LpAGPS1, LpAGPS2) [11]
Summary
Duckweed is a general term for plants belonging to the Lemnaceae family, comprising five genera (Spirodela, Landoltia, Lemna, Wolffiella, and Wolffia) and more than 40 species [1]. The plants in this family do not differentiate into stems and leaves but exhibit leaflike fronds bearing none or several roots on the underside. This study uncovered the cDNA sequences, structures, subunit interactions, expression patterns, and subcellular localization of AGPase. These findings provide new insights into the molecular mechanism of fast starch accumulation in L. punctata
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