Cadmium (Cd) is a heavy metal highly toxic to living organisms. Cd pollution of soils has become a serious problem worldwide, posing a severe threat to crop production and human health. When plants are poisoned by Cd, their growth and development are inhibited, chloroplasts are severely damaged, and respiration and photosynthesis are negatively affected. Therefore, elucidating the molecular mechanisms that underlie Cd tolerance in plants is important. Transcription factors can bind to specific plant cis-acting genes. Transcription factors are frequently reported to be involved in various signaling pathways involved in plant growth and development. Their role in the resistance to environmental stress factors, particularly Cd, should not be underestimated. The roles of several transcription factor families in the regulation of plant resistance to Cd stress have been widely demonstrated. In this review, we summarize the mechanisms of five major transcription factor families-WRKY, ERF, MYB, bHLH, and bZIP-in plant resistance to Cd stress to provide useful information for using molecular techniques to solve Cd pollution problems in the future.

The basic helix-loop-helix (bHLH) transcription factor family is the second largest in plants. bHLH transcription factor is not only universally involved in plant growth and metabolism, including photomorphogenesis, light signal transduction, and secondary metabolism, but also plays an important role in plant response to stress. However, the function of bHLH TFs in Pseudoroegneria species has not been studied yet. Pseudoroegneria (Nevski) Á. Löve is a perennial genus of the Triticeae. Pseudoroegneria species are mostly distributed in arid/semi-arid areas and they show good drought tolerance. In this study, we identified 152 PlbHLH TFs in Pseudoroegneria libanotica, which could be classified into 15 groups. Collinearity analysis indicates that 122 PlbHLH genes share homology with wbHLH genes in wheat, and it has lower homology with AtbHLH genes in Arabidopsis. Based on transcriptome profiling under an experiment with three PEG concentrations (0%, 10%, and 20%), 10 up-regulated genes and 11 down-regulated PlbHLH genes were screened. Among them, PlbHLH6, PlbHLH55 and PlbHLH64 as candidate genes may be the key genes related to drought tolerance response at germination, and they have been demonstrated to respond to drought, salt, oxidative, heat, and heavy metal stress in yeast. This study lays the foundation for an in-depth study of the biological roles of PlbHLHs in Pse. libanotica, and discovered new drought-tolerance candidate genes to enhance the genetic background of Triticeae crops.
UNASSIGNED: The online version contains supplementary material available at 10.1007/s12298-024-01433-w.

Anthocyanins play a paramount role in color variation and significantly contribute to the economic value of ornamental plants. The conserved activation complex MYB-bHLH-WD40 (MBW; MYB: v-myb avian myeloblastosis viral oncogene homolog; bHLH: basic helix-loop-helix protein; WD40:WD-repeat protein) involved in anthocyanin biosynthesis has been thoroughly researched, but there have been limited investigations into the function of repressor factors. In this study, we characterized TgMYB4, an R2R3-MYB transcriptional repressor which is highly expressed during petal coloration in red petal cultivars. TgMYB4-overexpressing tobaccos exhibited white or light pink petals with less anthocyanin accumulation compared to control plants. TgMYB4 was found to inhibit the transcription of ANTHOCYANIDIN SYNTHASE (TfANS1) and DIHYDRO-FLAVONOL-4-REDUCTASE (AtDFR), although it did not bind to their promoters. Moreover, the TgMYB4 protein was able to compete with the MYB activator to bind to the :bHLHprotein, thereby suppressing the function of the activator MBW complex. These findings demonstrate that TgMYB4 plays a suppressive role in the regulation of anthocyanin synthesis during flower pigmentation.

CONCLUSIONS: Sweet cherry PavbHLH106 was up-regulated under cold induction and overexpressed to enhance the cold resistance in tobacco by mediating the scavenging of ROS through increasing of antioxidant enzyme activity. Sweet cherry (Prunus avium L.) is an economically important fruit. Chilling requirements are critical during dormancy, but abnormally low temperatures unfavorably affect fruit growth and development. Differences were found in the transcript level of PavbHLH106 under salt, dehydration, and low-temperature treatments, especially in response to cold stress, suggesting that this gene is involved in the regulation of different abiotic stresses. PavbHLH106 is homologous to Arabidopsis thaliana AtbHLH106 with a conserved bHLH domain, and transient expression in tobacco suggests that the protein is localized in the nucleus and has transcriptional activity in yeast. The PavbHLH106 overexpression in tobacco resulted in weaker electrolyte leakages, lower malondialdehyde, and higher proline content than the wild type at low-temperature treatment. Reactive oxygen species accumulation was significantly reduced in the overexpressed lines, negatively correlated with the antioxidant enzyme activity. In addition, overexpression of PavbHLH106 delayed the germination of tobacco seeds and promoted plant growth. Resistance-related genes were expressed more in the overexpressed plants compared to the wild type. PavbHLH106 bound to the PavACO promoter in yeast and potentially interacted with a bHLH162-like transcription factor. These results indicate that PavbHLH106 has various functions and is particularly active in controlling low-temperature stress.

The tapetum, a crucial innermost layer encompassing male reproductive cells within the anther wall, plays a pivotal role in normal pollen development. The transcription factors (TFs) bHLH010/089/091 redundantly facilitate the rapid nuclear accumulation of DYSFUNCTIONAL TAPETUM 1, a gatekeeper TF in the tapetum. Nevertheless, the regulatory mechanisms governing the activity of bHLH010/089/091 remain unknown. In this study, we reveal that caffeoyl coenzyme A O-methyltransferase 1 (CCoAOMT1) is a negative regulator affecting the nuclear localization and function of bHLH010 and bHLH089, probably through their K259 site. Our findings underscore that CCoAOMT1 promotes the nuclear export and degradation of bHLH010 and bHLH089. Intriguingly, elevated CCoAOMT1 expression resulted in defective pollen development, mirroring the phenotype observed in bhlh010 bhlh089 mutants. Moreover, our investigation revealed that the K259A mutation in the bHLH089 protein disrupted its translocation from the nucleus to the cytosol and impeded its degradation induced by CCoAOMT1. Importantly, transgenic plants with the probHLH089::bHLH089K259A construct failed to rescue proper pollen development or gene expression in bhlh010 bhlh089 mutants. Collectively, these findings emphasize the need to maintain balanced TF homeostasis for male fertility. They firmly establish CCoAOMT1 as a pivotal regulator that is instrumental in achieving equilibrium between the induction of the tapetum transcriptional network and ensuring appropriate anther development.

The basic/helix-loop-helix (bHLH) family is a major family of transcription factors in plants. Although it has been reported that bHLH plays a defensive role against pathogen infection in plants, there is no comprehensive study on the bHLH-related defence response in rose (Rosa sp.). In this study, a genome-wide analysis of bHLH family genes (RcbHLHs) in rose was carried out, including their phylogenetic relationships, gene structure, chromosome localization and collinearity analysis. Via phylogenetic analysis, a total of 121 RcbHLH genes in the rose genome were divided into 21 sub-groups. These RcbHLHs are unevenly distributed in all 7 chromosomes of rose. The occurrence of gene duplication events indicates that whole-genome duplication and segmental duplication may play a key role in gene duplication. Ratios of non-synonymous to synonymous mutation frequency (Ka/Ks) analysis showed that the replicated RcbHLH genes mainly underwent purification selection, and their functional differentiation was limited. Gene expression analysis showed that 46 RcbHLHs were differentially expressed in rose petals upon B. cinerea infection. It is speculated that these RcbHLHs are candidate genes that regulate the response of rose plants to B. cinerea infection. Virus-induced gene silencing (VIGS) confirmed that RcbHLH112 in rose is a susceptibility factor for infection with B. cinerea. This study provides useful information for further study of the functions of the rose bHLH gene family.

The basic helix-loop-helix (bHLH) transcription factors possess DNA-binding and dimerization domains and are involved in various biological and physiological processes, such as growth and development, the regulation of secondary metabolites, and stress response. However, the bHLH gene family in C. tinctorius has not been investigated. In this study, we performed a genome-wide identification and analysis of bHLH transcription factors in C. tinctorius. A total of 120 CtbHLH genes were identified, distributed across all 12 chromosomes, and classified into 24 subfamilies based on their phylogenetic relationships. Moreover, the 120 CtbHLH genes were subjected to comprehensive analyses, including protein sequence alignment, evolutionary assessment, motif prediction, and the analysis of promoter cis-acting elements. The promoter region analysis revealed that CtbHLH genes encompass cis-acting elements and were associated with various aspects of plant growth and development, responses to phytohormones, as well as responses to both abiotic and biotic stresses. Expression profiles, sourced from transcriptome databases, indicated distinct expression patterns among these CtbHLH genes, which appeared to be either tissue-specific or specific to certain cultivars. To further explore their functionality, we determined the expression levels of fifteen CtbHLH genes known to harbor motifs related to abiotic and hormone responses. This investigation encompassed treatments with ABA, salt, drought, and MeJA. The results demonstrated substantial variations in the expression patterns of CtbHLH genes in response to these abiotic and hormonal treatments. In summary, our study establishes a solid foundation for future inquiries into the roles and regulatory mechanisms of the CtbHLH gene family.

The bHLH family, as a superfamily of transcription factors (TFs), has special functional characteristics in plants and plays a crucial role in a plant\'s growth and development and helping the plant cope with various stresses. In this study, 128 bHLH family genes were screened in the birch (B. platyphylla) genome using conservative domain scan and blast analysis. These genes are clustered into 21 subfamilies based on the phylogenetic tree construction and are unevenly distributed among the 14 birch chromosomes. In all, 22 segmental duplication pairs with 27 BpbHLH genes were identified. The duplications were distributed on eight chromosomes. Analysis of gene structures and protein motifs revealed intra-group conservation of BpbHLHs. Of the BpbHLH family genes, 16 contain only one intron each. The BPChr14G06667 gene contains the most introns, that is, 19. The cis-elements, which respond to plant hormones, light, defense, and stress, were found on the promoter of BHLH family genes. As per RNA-seq data analysis, under PEG osmotic stress, most BpbHLH genes were differentially expressed, and eight were highly differentially expressed. The qRT-PCR analysis results further indicated that BPChr06G09475 was the gene with the highest expression level in leaves, roots, and stems, and that the expression of these eight genes was higher in leaves than in roots and stems and upregulated in all three tissues under osmotic stress compared to the controls. The above analysis suggests that the BpbHLH family genes have a certain biological effect under drought stress that provides a basis for molecular breeding for stress resistance in birch.

Sunflowers are well-known ornamental plants, while sunflowers with red corolla are rare and the mechanisms underlying red coloration remain unclear. Here, a comprehensive analysis of metabolomics and transcriptomics on flavonoid pathway was performed to investigate the molecular mechanisms underlying the differential color formation between red sunflower Pc103 and two yellow sunflowers (Yr17 and Y35). Targeted metabolomic analysis revealed higher anthocyanin levels but lower flavonol content in Pc103 compared to the yellow cultivars. RNA-sequencing and phylogenetic analysis identified multiple genes involved in the flavonoid pathway, including series of structural genes and three MYB and bHLH genes. Specifically, HaMYBA and HabHLH1 were up-regulated in Pc103, whereas HaMYBF exhibited reduced expression. HaMYBA was found to interact with HabHLH1 in vivo and in vitro, while HaMYBF does not. Transient expression analysis further revealed that HabHLH1 and HaMYBA cooperatively regulate increased expression of dihydroflavonol 4-reductase (DFR), leading to anthocyanin accumulation. On the other hand, ectopic expression of HaMYBF independently modulates flavonol synthase (FLS) expression, but hindered anthocyanin production. Collectively, our findings suggest that the up-regulation of HaMYBA and HabHLH1, as well as the down-regulation of HaMYBF, contribute to the red coloration in Pc103. It offers a theoretical basis for improving sunflower color through genetic engineering.

Root hair is regarded as a pivotal complementary survival tactic for mycorrhizal plant like Abies beshanzuensis when symbiosis is disrupted. Relatively little is known about the mechanism underlying root hair morphogenesis in plant species that are strongly dependent on mycorrhizal symbiosis. Many of these species are endangered, and this knowledge is critical for ensuring their survival. Here, a MYB6/bHLH13-sucrose synthase 2 (AbSUS2) module was newly identified and characterized in A. beshanzuensis using bioinformatics, histochemistry, molecular biology, and transgenesis. Functional, expression pattern, and localization analysis showed that AbSUS2 participated in sucrose synthesis and was involved in root hair initiation in A. beshanzuensis. Additionally, the major enzymatic product of AbSUS2 was found to suppress root hair initiation in vitro. Our data further showed that a complex involving the transcription factors AbMYB6 and AbbHLH13 directly interacted with the promoter of AbSUS2 and strengthened its expression, thereby inhibiting root hair initiation in response to exogenous sucrose. Our findings offer novel insights into how root hair morphogenesis is regulated in mycorrhizal plants and also provide a new strategy for the preservation of endangered mycorrhizal plant species.