Identification and analysis of an outer-seed-coat-specific promoter from Arabidopsis thaliana

Abstract

Differentiation of the Arabidopsis thaliana (Arabidopsis) seed coat epidermal cells involves pronounced changes highlighted by the synthesis and secretion of copious amounts of dispensable, pectinaceous mucilage followed by a thick cellulosic secondary cell wall. This cell type, therefore, represents an excellent molecular-genetic model to study the biosynthesis and modification of cell wall components, particularly pectin. To support such research, we sought to identify a promoter that drives expression specifically in the Arabidopsis seed coat epidermis. Arabidopsis seed coat microarray data was analysed for genes expressed in the wild type seed coat but not the seed coat of the apetala2 mutant where the epidermal cells fail to differentiate. Of 14 candidate genes, 9 showed a seed-specific expression pattern by reverse transcriptase-PCR. Transcriptional regulatory region-β-glucuronidase (GUS) reporter gene fusions introduced into Arabidopsis identified one promoter, that of the DIRIGENT PROTEIN1 (DP1) gene, as seed coat specific. The specificity of the expression was confirmed using a second reporter gene, Citrine YFP. Expression of both reporter genes was limited to the epidermal and palisade cell layers of the seed coat. Quantitative PCR data using wild type seed coat RNA suggested that the promoter is particularly active at 7 days post anthesis. The DP1 promoter was able to direct transcription of GUS in a similar pattern in the Brassica napus seed coat. Thus, in addition to its application in studying the plant cell wall, this promoter will provide an experimental tool for expressing high-valued recombinant proteins as well as modifying seed coat traits in economically important crops.

Unidirectional Movement of Cellulose Synthase Complexes in Arabidopsis Seed Coat Epidermal Cells Deposit Cellulose Involved in Mucilage Extrusion, Adherence, and Ray Formation

“…The unique deposition of cellulose in mucilage provides an excellent system to further study the interactions between pectins and cellulose and their effects on cell wall rheology. Additional genes that are hypothesized to facilitate cellulose biosynthesis or interactions between pectins and cellulose could be investigated in vivo in the seed coat epidermis using a cell-specific promoter (Esfandiari et al, 2013).…”

Detection and Adaptation in Parasitic Angiosperm Host Selection

“…Based on cDNA sequences, three genomic sequences were obtained from S. asiatica genomic DNA, extracted using the DNeasy Plant Mini Kit (Qiagen). The promoter region for each putative SaNOX homologue was identified by the thermal asymmetric interlaced (TAIL)-PCR method [16].…”

“…Distinct promoter sites for each of these genes were obtained using thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR) [16]. Exon-intron analyses of the full-length genomic sequences, as well as structural predictions based on the mRNA, are available in the Supplementary Information and summarized below (Supplementary Figures S2-S4).…”

Expression ofPRX36,PMEI6andSBT1.7is controlled by complex transcription factor regulatory networks for proper seed coat mucilage extrusion

“…24,35 Finally, the nearly identical regulatory pattern of PRX36 and DIR12 is in good agreement with the very high score of DIR12 within the PRX36 ATTED-II network and their very similar spatiotemporal expression patterns. 24,28 However, this does not preclude the existence of a common mucilage release phenotype as demonstrated here. It could be hypothesized that MSC are not only linked to abundant polysaccharidic secretion but also harbors specialized natural products for defense purpose such as neolignans.…”

“…25 Among the numerous genes identified as members of the same network, 2 additional putative candidates have been chosen: another class III peroxidase, PRX56; a dirigent protein, DIR12 ( Table 1). While PRX56 role has not been previously studied, DIR12, also named DP1, belongs to a class of genes necessary to control stereochemical phenoxy radical coupling during natural product synthesis 26,27 and has been shown to be specifically expressed in the outer-seed-coat 28 with a role in production of a specific neolignan. 29 Here, the expression of genes coding for the 3 cell wall proteins, PRX36/PMEI6/ SBT1.7, and the 2 candidates genes, PRX56/DIR12, will be studied in a panel of T-DNA mutants affected in master TFs described above (ap2, egl3/gl3, gl2, gl3, myb5, tt8, ttg1, ttg2 and luh1/mum1).…”