School of Agricultural Biotechnology, Punjab Agricultural University, Ludhiana, Punjab, India.
School of Biology and Ecology, University of Maine, Orono, ME, USA.
Mol Biol Rep. 2020 Jan;47(1):401-422. doi: 10.1007/s11033-019-05144-4. Epub 2019 Oct 22.
Rice bran oil is good quality edible oil, rich in antioxidants and comprised typically of oleic-linoleic type fatty acids. However, presence of a highly lipolytic enzyme Phospholipase D alpha1 (OsPLDα1) increases free fatty acid content in the oil which further leads to stale flavor and rancidity of the oil, making it unfit for human consumption. In this study, we compared the upstream regions of OsPLDα1 orthologs across 34 accessions representing 5 wild Oryza species and 8 cultivars, to uncover sequence variations and identify cis-elements involved in differential transcription of orthologs. Alignment of the upstream sequences to the Nipponbare OsPLDα1 reference sequence revealed the presence of 39 SNPs. Phylogenetic analysis showed that all the selected cultivars and wild species accessions are closely related to the reference except for three accessions of O. rufipogon (IRGC89224, IRGC104425, and IRGC105902). Furthermore, using exon-specific qRT-PCR, OsPLDα1 expression patterns in immature grains indicated significant differences in transcript abundance between the wild species accessions. In comparison to the control, lowest gene expression was observed in IRGC89224 accession (0.20-fold) followed by IRGC105902 (0.26-fold) and IRGC104425 (0.41-fold) accessions. In-silico analysis of the OsPLDα1 promoter revealed that the copy number variations of CGCGBOXAT, GT1CONSENSUS, IBOXCORE, NODCON2GM, OSE2ROOTNODULE, SURECOREATSULTR11, and SORLIP1AT cis-elements play an important role in the transcriptional activities of orthologous genes. Owing to the presence of ARFAT and SEBF elements only in the IRGC89224 accession, which had the lowest gene expression as well, these putative upstream regulatory sequences have been identified as novel cis-elements which may act as repressors in regulating the OsPLDα1 gene expression. The accessions identified with low OsPLDα1 expressions could be further deployed as potential donors of ideal OsPLDα1 allele for transfer of the desired trait into elite rice cultivars.
米糠油是一种优质食用油，富含抗氧化剂，通常由油酸-亚油酸型脂肪酸组成。然而，存在一种高度脂解的酶磷脂酶 D alpha1（OsPLDα1），它会增加油中的游离脂肪酸含量，从而导致油的味道变差和变质，使其不适合人类食用。在这项研究中，我们比较了代表 5 种野生稻种和 8 个品种的 34 个品系的 OsPLDα1 直系同源物的上游区域，以揭示序列变异，并鉴定参与直系同源物差异转录的顺式元件。将上游序列与 Nipponbare OsPLDα1 参考序列进行比对，发现存在 39 个 SNP。系统发育分析表明，除了 3 个 O. rufipogon （IRGC89224、IRGC104425 和 IRGC105902）外，所有选择的品种和野生种都与参考序列密切相关。此外，使用exon-specific qRT-PCR，对未成熟籽粒中 OsPLDα1 的表达模式进行分析表明，野生种间的转录物丰度存在显著差异。与对照相比，IRGC89224 品系（0.20 倍）的基因表达最低，其次是 IRGC105902 （0.26 倍）和 IRGC104425 （0.41 倍）品系。对 OsPLDα1 启动子的计算机分析表明，CGCGBOXAT、GT1CONSENSUS、IBOXCORE、NODCON2GM、OSE2ROOTNODULE、SURECOREATSULTR11 和 SORLIP1AT 顺式元件的拷贝数变异在同源基因的转录活性中起着重要作用。由于只有在基因表达最低的 IRGC89224 品系中存在 ARFAT 和 SEBF 元件，因此这些假定的上游调节序列已被确定为新的顺式元件，它们可能作为抑制物在调节 OsPLDα1 基因表达中起作用。鉴定出的 OsPLDα1 表达较低的品系可进一步作为理想 OsPLDα1 等位基因的潜在供体，将所需性状转移到优良水稻品种中。
