Elucidation of Novel cis-Regulatory Elements and Promoter Structures Involved in Iron Excess Response Mechanisms in Rice Using a Bioinformatics Approach

Abstract

Iron (Fe) excess is a major constraint on crop production in flooded acidic soils, particularly in rice cultivation. Under Fe excess, plants activate a complex mechanism and network regulating Fe exclusion by roots and isolation in various tissues. In rice, the transcription factors and cis-regulatory elements (CREs) that regulate Fe excess response mechanisms remain largely elusive. We previously reported comprehensive microarray analyses of several rice tissues in response to various levels of Fe excess stress. In this study, we further explored novel CREs and promoter structures in rice using bioinformatics approaches with this microarray data. We first performed network analyses to predict Fe excess-related CREs through the categorization of the gene expression patterns of Fe excess-responsive transcriptional regulons, and found four major expression clusters: Fe storage type, Fe chelator type, Fe uptake type, and WRKY and other co-expression type. Next, we explored CREs within these four clusters of gene expression types using a machine-learning method called microarray-associated motif analyzer (MAMA), which we previously established. Through a comprehensive bioinformatics approach, we identified a total of 560 CRE candidates extracted by MAMA analyses and 42 important conserved sequences of CREs directly related to the Fe excess response in various rice tissues. We explored several novel cis-elements as candidate Fe excess CREs including GCWGCWGC, CGACACGC, and Myb binding-like motifs. Based on the presence or absence of candidate CREs using MAMA and known PLACE CREs, we found that the Boruta-XGBoost model explained expression patterns with high accuracy of about 83%. Enriched sequences of both novel MAMA CREs and known PLACE CREs led to high accuracy expression patterns. We also found new roles of known CREs in the Fe excess response, including the DCEp2 motif, IDEF1-, Zinc Finger-, WRKY-, Myb-, AP2/ERF-, MADS- box-, bZIP and bHLH- binding sequence-containing motifs among Fe excess-responsive genes. In addition, we built a molecular model and promoter structures regulating Fe excess-responsive genes based on new finding CREs. Together, our findings about Fe excess-related CREs and conserved sequences will provide a comprehensive resource for discovery of genes and transcription factors involved in Fe excess-responsive pathways, clarification of the Fe excess response mechanism in rice, and future application of the promoter sequences to produce genotypes tolerant of Fe excess.

Editorial: Nutrient Interactions in Plants

“…The perspective article is devoted to describing new approaches based on computational analysis to predict interactions between proteins related to different elements (Di Silvestre et al). Finally, within the 13 original research articles; eight of them are about the interactions between two or three elements, including non-essential ones, such as Fe-Cu, Si-Fe, Fe-P-S, S-N, Fe-Zn, K-Na, or P-Zn (Bernal and Kramer, 2021;Jian et al, 2021;Kakei et al, 2021;Li et al, 2021;Chaiwong et al;García et For instance, P deficiency, through organic acid release and rhizosphere acidification, can promote the mobilization of other nutrients, like Fe or Zn (Pueyo et al, 2021). In the same way, Si application can promote N and P acquisition by upregulating nitrate and phosphate transporters (Pavlovic et al, 2021).…”

“…The perspective article is devoted to describing new approaches based on computational analysis to predict interactions between proteins related to different elements ( Di Silvestre et al ). Finally, within the 13 original research articles; eight of them are about the interactions between two or three elements, including non-essential ones, such as Fe-Cu, Si-Fe, Fe-P-S, S-N, Fe-Zn, K-Na, or P-Zn (Bernal and Kramer, 2021 ; Jian et al, 2021 ; Kakei et al, 2021 ; Li et al, 2021 ; Chaiwong et al ; García et al ; Suman et al ; Yu et al ); four of them are about the interactions among many nutrients, such as ionome-macronutrients, ionome-micronutrients, ionome-N ( Courbet et al ; D'Oria et al ; Zhang C. et al ; Zhang J. et al ); and one is devoted to the substitution of K by Na ( Mateus et al ).…”

“…The articles included in this Research Topic reflect indirect interactions between nutrients, such as those simultaneously analyzing many nutrients ( Courbet et al ; D'Oria et al ; Zhang C. et al ; Zhang J. et al ), and also direct interactions, like those studying the interplay between 2 and 3 elements (Bernal and Kramer, 2021 ; Jian et al, 2021 ; Kakei et al, 2021 ; Li et al, 2021 ; Chaiwong et al ; García et al ; Mateus et al ; Suman et al ; Yu et al ). The depicted interactions occur at different steps of nutrient acquisition and translocation inside the plant.…”

Design and Analysis of Native Photorespiration Gene Motifs of Promoter Untranslated Region Combinations Under Short Term Abiotic Stress Conditions

“…Since synthetic promoters are assembled linking cis -regulatory elements of various promoter regions upstream of the core promoter (TATA box), one can combine tissue specificity with heightened expression within the same promoter construct. Unfortunately, unlike yeast or animal genomics, very few studies have been devoted to identifying core cis -regulatory elements underlying developmental and biochemical pathways in plants to help us generate an array of synthetic promoters for gene stacking in crop plants ( Kaplan et al, 2006 ; Kaur et al, 2017 ; Laxa and Fromm, 2018 ; Eseverri et al, 2020 ; Jores et al, 2021 ; Kakei et al, 2021 ). In addition to promoters, upstream and downstream untranslated regions (UTRs) also have a profound effect on gene expression ( Diamos and Mason, 2018 ; Yamamoto et al, 2018 ; de Felippes et al, 2020 ).…”

“…Despite the knowledge of the core photorespiratory enzymes very little is known about how they are regulated ( Rojas et al, 2012 ; Saji et al, 2017 ). Recent reports have shown the potential of utilizing cis -regulatory elements (CREs) of endogenous genes to fine-tune transgene expression for preserving spatiotemporal regulatory characteristics ( Kaur et al, 2017 ; Cai et al, 2020 ; Jores et al, 2021 ; Kakei et al, 2021 ; Melo et al, 2021 ). Synthetic promoters designed with single or multiple CREs identified from native photorespiratory genes can provide valuable insights into the potential positive and negative regulators of photorespiration.…”