Cloning, Expression, and Tobacco Overexpression Analyses of a PISTILLATA/GLOBOSA-like (OfGLO1) Gene from Osmanthus fragrans

GLOBOSA (GLO), a B-class MADS-box gene, is involved in floral organ determination but has rarely been studied in Osmanthus fragrans, which is a very popular ornamental tree species in China. Here, the full-length cDNA of a homologous GLO1 gene (named OfGLO1) was cloned from a flower bud of O. fragrans using the RACE technique. The OfGLO1 has a 645 bp open reading frame, encoding 214 amino acids. Similar to other PI/GLO proteins, OfGLO1 has two conserved domains, MADS MEF2-like and K-box, and a 16-amino-acid PI motif in the C terminal region. Our phylogeny analysis classified OfGLO1 as a PI-type member of the B-class MADS-box gene family. The qRT-PCR assay showed that the expression of OfGLO1 in O. fragrans was continuously upregulated from the tight bud stage to the full flowering stage but barely expressed in the pistils, sepals, and non-floral organs, such as root, leaf, and stem. The genetic effect of OfGLO1 was assayed by ectopic expression in tobacco plants. Compared with the wild-type, OfGLO1 transformants showed reduced plant size, earlier flowering, shorter stamens, and lower seed setting rates. Furthermore, some stamens were changed into petal-like structures. These findings indicate that OfGLO1 plays an important role in the regulation of flower development. This study improved our understanding of class B gene function in woody plants

Transcriptional and protein structural characterization of homogentisate phytyltransferase genes in barley, wheat, and oat

Abstract Background Homogentisate phytyltransferase (HPT) is the critical enzyme for the biosynthesis of tocopherols (vitamin E), which are the major lipid-soluble antioxidants and help plants adapt to various stress conditions. HPT is generally strictly conserved in various plant genomes; however, a divergent lineage HPT2 was identified recently in some Triticeae species. The molecular function and transcriptional profiles of HPT2 remain to be characterized. Results In this study, we performed comprehensive transcriptome data mining of HPT1 and HPT2 in different tissues and stages of barley (Hordeum vulgare), wheat (Triticum aestivum), and oat (Avena sativa), followed by qRT-PCR experiments on HPT1 and HPT2 in different tissues of barley and wheat. We found that the common HPT1 genes (HvHPT1, TaHPT1s, and AsHPT1s) displayed a conserved transcriptional pattern in the three target species and were universally transcribed in various tissues, with a notable preference in leaf. In contrast, HPT2 genes (HvHPT2, TaHPT2, and AsHPT2) were specifically transcribed in spike (developmentally up-regulated) and shoot apex tissues, displaying a divergent tissue-specific pattern. Cis-regulatory elements prediction in the promoter region identified common factors related to light-, plant hormone-, low temperature-, drought- and defense- responses in both HPT1s and HPT2s. We observed the transcriptional up-regulation of HvHPT1 and HvHPT2 under various stress conditions, supporting their conserved function in environmental adaption. We detected a clear, relaxed selection pressure in the HPT2 lineage, consistent with the predicted evolution pattern following gene duplication. Protein structural modelling and substrate docking analyses identified putative catalytic amino acid residues for HvHPT1 and HvHPT2, which are strictly conserved and consistent with their function in vitamin E biosynthesis. Conclusions We confirmed the presence of two lineages of HPT in Triticeae and Aveninae, including hexaploid oat, and characterized their transcriptional profiles based on transcriptome and qRT-PCR data. HPT1s were ubiquitously transcribed in various tissues, whilst HPT2s were highly expressed in specific stages and tissue. The active transcription of HPT2s, together with its conserved cis-elements and protein structural features, support HPT2s’ role in tocopherol production in Triticeae. This study is the first protein structural analysis on the membrane-bound plant HPTs and provides valuable insights into its catalytic mechanism