Transcriptional cascades in the regulation of 2-AP biosynthesis under Zn supply in fragrant rice.

Transcription factors (TFs) regulate gene expression to control certain genetic programs, such as growth and development, phytohormone regulation, and environmental stresses. 2-acetyl-1-pyrroline (2-AP) is the key element involved in aroma biosynthesis pathway, and the application of micronutrients can increase the 2-AP levels. However, little is known about the micronutrient-induced TFs involved in 2-AP biosynthesis. Here, we identify a number of TF families in two fragrant rice varieties, "Meixiangzhan-2" (M) and "Xiangyaxiangzhan" (X), in response to Zinc (Zn) application through transcriptomic analysis. A total of ~678 TFs were identified and grouped into 26 TF families, each of which was found to be involved in numerous signaling pathways. The WRKY TF family was found to be the most abundant, followed by bHLH and MYB. Furthermore, members of the WRKY, bHLH, MYB, ERF, HSF, MADS-box, NFY, and AP2 TF families were significantly upregulated and may be involved in the transcriptional regulation of aroma biosynthesis. In brief, this study enhances our understanding of the molecular mechanism of 2-AP biosynthesis and highlights the key TFs potentially involved in the production of aroma in fragrant rice.

Post-transcriptional regulation of 2-acetyl-1-pyrroline (2-AP) biosynthesis pathway, silicon, and heavy metal transporters in response to Zn in fragrant rice.

Fragrant rice (Oryza sativa L.) has a high economic and nutritional value, and the application of micronutrients regulates 2-acetyl-1-pyrroline (2-AP) production, which is responsible for aroma in fragrant rice. Alternative splicing (AS) is an important post-transcriptional regulatory mechanism to generate transcript variability and proteome diversity in plants. However, no systematic investigation of AS events in response to micronutrients (Zn) has been performed in fragrant rice. Furthermore, the post-transcriptional regulation of genes involved in 2-AP biosynthesis is also not known. In this study, a comprehensive analysis of AS events under two gradients of Zn treatment in two different fragrant rice cultivars (Meixiangzhan-2 and Xiangyaxiangzhan) was performed based on RNA-seq analysis. A total of 386 and 598 significant AS events were found in Meixiangzhan-2 treated with low and high doses of Zn, respectively. In Xiangyaxiangzhan, a total of 449 and 598 significant AS events were found in low and high doses of Zn, respectively. Go analysis indicated that these genes were highly enriched in physiological processes, metabolism, and cellular processes in both cultivars. However, genotype and dose-dependent AS events were also detected in both cultivars. By comparing differential AS (DAS) events with differentially expressed genes (DEGs), we found a weak overlap among DAS and DEGs in both fragrant rice cultivars indicating that only a few genes are post-transcriptionally regulated in response to Zn treatment. We further report that Zn differentially regulates the expression of 2-AP biosynthesis-related genes in both cultivars and Zn treatment altered the editing frequency of single nucleotide polymorphism (SNPs) in the genes involved in 2-AP biosynthesis. Finally, we showed that epigenetic modifications associated with active gene transcription are generally enriched over 2-AP biosynthesis-related genes. Similar to the 2-AP pathway, we found that heavy metal transporters (genes related to silicon, iron, Zn and other metal transport) are also regulated at transcriptional and post-transcriptional levels in response to Zn in fragrant rice. Taken together, our results provide evidence of the post-transcriptional gene regulation in fragrant rice in response to Zn treatment and highlight that the 2-AP biosynthesis pathway and heavy metal transporters may also be regulated through epigenetic modifications. These findings will serve as a cornerstone for further investigation to understand the molecular mechanisms of 2-AP biosynthesis and regulation of heavy metal transporters in fragrant rice.