Comprehensive Analysis of bHLH Transcription Factors in Ipomoea aquatica and Its Response to Anthocyanin Biosynthesis

Abstract

The basic helix-loop-helix (bHLH) proteins compose one of the largest transcription factor (TF) families in plants, which play a vital role in regulating plant biological processes including growth and development, stress response, and secondary metabolite biosynthesis. Ipomoea aquatica is one of the most important nutrient-rich vegetables. Compared to the common green-stemmed I. aquatica, purple-stemmed I. aquatica has extremely high contents of anthocyanins. However, the information on bHLH genes in I. aquatica and their role in regulating anthocyanin accumulation is still unclear. In this study, we confirmed a total of 157 bHLH genes in the I. aquatica genome, which were classified into 23 subgroups according to their phylogenetic relationship with the bHLH of Arabidopsis thaliana (AtbHLH). Of these, 129 IabHLH genes were unevenly distributed across 15 chromosomes, while 28 IabHLH genes were spread on the scaffolds. Subcellular localization prediction revealed that most IabHLH proteins were localized in the nucleus, while some were in the chloroplast, extracellular space, and endomembrane system. Sequence analysis revealed conserved motif distribution and similar patterns of gene structure within IabHLH genes of the same subfamily. Analysis of gene duplication events indicated that DSD and WGD played a vital role in the IabHLH gene family expansion. Transcriptome analysis showed that the expression levels of 13 IabHLH genes were significantly different between the two varieties. Of these, the IabHLH027 had the highest expression fold change, and its expression level was dramatically higher in purple-stemmed I. aquatica than that in green-stemmed I. aquatica. All upregulated DEGs in purple-stemmed I. aquatica exhibited the same expression trends in both qRT-PCR and RNA-seq. Three downregulated genes including IabHLH142, IabHLH057, and IabHLH043 determined by RNA-seq had opposite expression trends of those detected by qRT-PCR. Analysis of the cis-acting elements in the promoter region of 13 differentially expressed genes indicated that light-responsive elements were the most, followed by phytohormone-responsive elements and stress-responsive elements, while plant growth and development-responsive elements were the least. Taken together, this work provides valuable clues for further exploring IabHLH function and facilitating the breeding of anthocyanin-rich functional varieties of I. aquatica.

Research on Plant Genomics and Breeding 2.0

“…Liu et al identified 157 bHLH genes in Ipomoea aquatica and characterized the features and evolutionary relationships of the bHLH gene family through collinearity analysis, phylogenetic analysis, and sequence feature analysis. The subsequent transcriptome data quantification of bHLH in purple and green watercress and the analysis of 13 differentially expressed genes (DEGs) through cis-acting element analysis indicated that bHLH plays an important role in light response and plant hormone regulation, providing important insights into the role of bHLH in the anthocyanin biosynthesis pathway [ 22 ]. Determining the functions of homologous genes based on those of known functional genes can be a way to significantly enhance the study of gene functions.…”

Research on Plant Genomics and Breeding

“…The role of IabHLHs family transcription factors in the biosynthesis of anthocyanins in I. aquatica was analyzed using bioinformatics combined with molecular experiments. This work provides valuable clues to further explore IabHLH's function and facilitating the breeding of anthocyanin-rich varieties of I. aquatica [17]. Improving the cold tolerance of herbaceous peonies (Paeonia lactiflora) is essential to plant them at low latitudes.…”

Molecular Characterization and Expression of CmobHLH Genes in Pumpkin

“…Identification of bHLH family genes was initially recorded in maize (Zea mays L.) [11]. Thereafter, an increasing number of bHLH genes were widely characterized in numerous plant species, such as Arabidopsis [12], rice [13], tomato [14], poplar [1], apple [15], eggplant [16], grass pea [17], persimmon [18], sweet wormwood [19] and Ipomoea aquatica [20].…”