scholarly journals Identification and Characterization of MYB-bHLH-WD40 Regulatory Complex Members Controlling Anthocyanidin Biosynthesis in Blueberry Fruits Development

Genes ◽  
2019 ◽  
Vol 10 (7) ◽  
pp. 496 ◽  
Author(s):  
Zhao ◽  
Li ◽  
Zhu ◽  
Chang ◽  
Li ◽  
...  

Anthocyanins is the main representative of flavonoids in blueberry fruits. The anthocyanins biosynthetic pathway has been extensively studied in numerous model plants and fruit crops at biochemical, genetic, and molecular levels. However, the mechanisms by which the MYB transcription factor/basic helix-loop-helix (bHLH) domain protein/WD-repeat (MYB-bHLH-WD40) complexes regulate anthocyanin biosynthesis in blueberry is still limited. In the present study, we identified 11 MYB, 7 bHLH, and 6 WD40 genes in blueberry fruits, using amino acid sequences of homologous MYB-bHLH-WD40 complexes in Arabidopsis, apple, grape, and strawberry. To understand these mechanisms, the expression patterns of MYB-bHLH-WD40 genes were examined and validated using differentially expressed gene (DEG) analysis and quantitative real-time reverse transcription PCR (qRT-PCR), respectively. The expression patterns of MYB-bHLH-WD40 genes positively correlated with anthocyanin accumulation and color development in blueberry fruits. Consistent with the effects of other transcriptional regulators, the VcMYBL1::GFP, VcbHLH1::GFP, and VcWDL2::GFP fusion proteins were only observed in the nucleus. The protein-protein interactions (PPIs) and bimolecular fluorescence complementation (BiFC) assay suggested a possible link between VcbHLHL1 and VcMYBL1. Finally, a model was proposed and discussed for how the expression of the MYB-bHLH-WD40 complexes can promote anthocyanin biosynthesis in blueberry fruits. To our knowledge, this study was the first to evaluate MYB-bHLH-WD40 complexes in blueberry fruits, and it provides a foundation to dissect the function of the mechanism.

2021 ◽  
Vol 12 ◽  
Author(s):  
Zhaofei Duan ◽  
Shiyu Tian ◽  
Guobin Yang ◽  
Min Wei ◽  
Jing Li ◽  
...  

Many basic helix-loop-helix transcription factors (TFs) have been reported to promote anthocyanin biosynthesis in numerous plant species, but little is known about bHLH TFs that inhibit anthocyanin accumulation. In this study, SmbHLH1 from Solanum melongena was identified as a negative regulator of anthocyanin biosynthesis. However, SmbHLH1 showed high identity with SmTT8, which acts as a SmMYB113-dependent positive regulator of anthocyanin-biosynthesis in plants. Overexpression of SmbHLH1 in eggplant caused a dramatic decrease in anthocyanin accumulation. Only the amino acid sequences at the N and C termini of SmbHLH1 differed from the SmTT8 sequence. Expression analysis revealed that the expression pattern of SmbHLH1 was opposite to that of anthocyanin accumulation. Yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) assays showed that SmbHLH1 could not interact with SmMYB113. Dual-luciferase assay demonstrated that SmbHLH1 directly repressed the expression of SmDFR and SmANS. Our results demonstrate that the biological function of bHLHs in anthocyanin biosynthesis may have evolved and provide new insight into the molecular functions of orthologous genes from different plant species.


2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Na Sang ◽  
Hui Liu ◽  
Bin Ma ◽  
Xianzhong Huang ◽  
Lu Zhuo ◽  
...  

Abstract Background In plants, 14-3-3 proteins, also called GENERAL REGULATORY FACTORs (GRFs), encoded by a large multigene family, are involved in protein–protein interactions and play crucial roles in various physiological processes. No genome-wide analysis of the GRF gene family has been performed in cotton, and their functions in flowering are largely unknown. Results In this study, 17, 17, 31, and 17 GRF genes were identified in Gossypium herbaceum, G. arboreum, G. hirsutum, and G. raimondii, respectively, by genome-wide analyses and were designated as GheGRFs, GaGRFs, GhGRFs, and GrGRFs, respectively. A phylogenetic analysis revealed that these proteins were divided into ε and non-ε groups. Gene structural, motif composition, synteny, and duplicated gene analyses of the identified GRF genes provided insights into the evolution of this family in cotton. GhGRF genes exhibited diverse expression patterns in different tissues. Yeast two-hybrid and bimolecular fluorescence complementation assays showed that the GhGRFs interacted with the cotton FLOWERING LOCUS T homologue GhFT in the cytoplasm and nucleus, while they interacted with the basic leucine zipper transcription factor GhFD only in the nucleus. Virus-induced gene silencing in G. hirsutum and transgenic studies in Arabidopsis demonstrated that GhGRF3/6/9/15 repressed flowering and that GhGRF14 promoted flowering. Conclusions Here, 82 GRF genes were identified in cotton, and their gene and protein features, classification, evolution, and expression patterns were comprehensively and systematically investigated. The GhGRF3/6/9/15 interacted with GhFT and GhFD to form florigen activation complexs that inhibited flowering. However, GhGRF14 interacted with GhFT and GhFD to form florigen activation complex that promoted flowering. The results provide a foundation for further studies on the regulatory mechanisms of flowering.


2020 ◽  
Author(s):  
Xiya Zuo ◽  
Shixiang Wang ◽  
Wen Xiang ◽  
Huiru Yang ◽  
Muhammad Mobeen Tahir ◽  
...  

Abstract Background: Apple (Malus domestica Borkh.) is one of the most popular cultivated fruit crops in China. Apple floral transition is an important process but liable to be affected by various environmental factors. The 14-3-3 proteins are involved in regulating diverse biological processes in plants, and some 14-3-3 members play vital roles in flowering. However, little information was available about the 14-3-3 members in apple.Results: In the current study, we identified eighteen 14-3-3 gene family members from the apple genome database, designated MdGF14a to MdGF14r. The isoforms possess a conserved core region comprising nine antiparallel α-helices and divergent N and C termini. According to their structural and phylogenetic features, Md14-3-3 proteins could be classified into two major evolutionary branches, the epsilon (ɛ) group and the non-epsilon (non-ɛ) group. Moreover, expression profiles derived from transcriptome data and quantitative real-time reverse transcription PCR analysis showed diverse expression patterns of Md14-3-3 genes in various tissues and in response to different sugars and hormone treatments during the floral transition phase. Four Md14‑3-3 isoforms (MdGF14a, MdGF14d, MdGF14i, and MdGF14j) exhibiting prominent transcriptional responses to sugars and hormones were selected for further investigation. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation experiments showed that the four Md14-3-3 proteins interact with key floral integrators, MdTFL1 (TERMINAL FLOWER1) and MdFT (FLOWERING LOCUS T). Subcellular localization of four selected Md14-3-3 proteins demonstrated their localization in both the cytoplasm and nucleus.Conclusion: We identified the Md14-3-3s family in apple comprehensively. Certain Md14-3-3 genes are expressed predominantly during the apple floral transition stage, and may participate in the regulation of flowering through association with flower control genes. Our results provide a preliminary framework for further investigation into the roles of Md14-3-3s in floral transition.


2020 ◽  
Author(s):  
Xiya Zuo ◽  
Shixiang Wang ◽  
Wen Xiang ◽  
Huiru Yang ◽  
Muhammad Mobeen Tahir ◽  
...  

Abstract Background: Apple (Malus domestica Borkh.) is a popular cultivated fruit crop with high economic value in China. Apple floral transition is an important process but liable to be affected by various environmental factors. The 14-3-3 proteins are involved in regulating diverse biological processes in plants, and some 14-3-3 members play vital roles in flowering. However, little information was available about the 14-3-3 members in apple.Results: In the current study, we identified eighteen 14-3-3 gene family members from the apple genome database, designated MdGF14a to MdGF14r. The isoforms possess a conserved core region comprising nine antiparallel α-helices and divergent N and C termini. According to their structural and phylogenetic features, Md14-3-3 proteins could be classified into two major evolutionary branches, the epsilon (ɛ) group and the non-epsilon (non-ɛ) group. Moreover, expression profiles derived from transcriptome data and quantitative real-time reverse transcription PCR analysis showed diverse expression patterns of Md14-3-3 genes in various tissues and in response to different sugars and hormone treatments during the floral transition phase. Four Md14‑3-3 isoforms (MdGF14a, MdGF14d, MdGF14i, and MdGF14j) exhibiting prominent transcriptional responses to sugars and hormones were selected for further investigation. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation experiments showed that the four Md14-3-3 proteins interact with key floral integrators, MdTFL1 (TERMINAL FLOWER1) and MdFT (FLOWERING LOCUS T). Subcellular localization of four selected Md14-3-3 proteins demonstrated their localization in both the cytoplasm and nucleus.Conclusion: We identified the Md14-3-3s family in apple comprehensively. Certain Md14-3-3 genes are expressed predominantly during the apple floral transition stage, and may participate in the regulation of flowering through association with flower control genes. Our results provide a preliminary framework for further investigation into the roles of Md14-3-3s in floral transition.


HortScience ◽  
2015 ◽  
Vol 50 (5) ◽  
pp. 640-649 ◽  
Author(s):  
Ji Tian ◽  
Zhen-yun Han ◽  
Li-ru Zhang ◽  
Ting-Ting Song ◽  
Jie Zhang ◽  
...  

Anthocyanins are protective pigments that accumulate in plant organs such as fruits and leaves, and are nutritionally valuable components of the human diet. There is thus considerable interest in the factors that regulate synthesis. Malus crabapple leaves are rich sources of these compounds, and in this study we analyzed leaf coloration, anthocyanin levels, and the expression levels of anthocyanin biosynthetic and regulatory genes in three crabapple cultivars (Royalty, Prairifire, and Flame) following various temperature treatments. We found that low temperatures (LTs) promoted anthocyanin accumulation in ‘Royalty’ and ‘Prairifire’, leading to red leaves, but not in ‘Flame’, which accumulated abundant colorless flavonols and retained green colored leaves. Quantitative reverse transcript PCR (RT-PCR) analyses indicated that the expression of several anthocyanin biosynthetic genes was induced by LTs, as were members of the R2R3-MYB, basic helix–loop–helix (bHLH) and WD40 transcription factor families that are thought to act in a complex. We propose that anthocyanin biosynthesis is differentially regulated in the three cultivars by LTs via the expression of members of this anthocyanin regulatory complex.


2019 ◽  
Vol 70 (12) ◽  
pp. 3111-3123 ◽  
Author(s):  
Lili Xiang ◽  
Xiaofen Liu ◽  
Heng Li ◽  
Xueren Yin ◽  
Donald Grierson ◽  
...  

Abstract ‘Jimba’, a well-known white flowered chrysanthemum cultivar, occasionally and spontaneously produces red colored petals under natural cultivation, but there is little information about the molecular regulatory mechanism underlying this process. We analysed the expression patterns of 91 MYB transcription factors in ‘Jimba’ and ‘Turning red Jimba’ and identified an R3 MYB, CmMYB#7, whose expression was significantly decreased in ‘Turning red Jimba’ compared with ‘Jimba’, and confirmed it is a passive repressor of anthocyanin biosynthesis. CmMYB#7 competed with CmMYB6, which together with CmbHLH2 is an essential component of the anthocyanin activation complex, for interaction with CmbHLH2 through the bHLH binding site in the R3 MYB domain. This reduced binding of the CmMYB6–CmbHLH2 complex and inhibited its ability to activate CmDFR and CmUFGT promoters. Moreover, using transient expression assays we demonstrated that changes in the expression of CmMYB#7 accounted for alterations in anthocyanin content. Taken together, our findings illustrate that CmMYB#7 is a negative regulator of anthocyanin biosynthesis in chrysanthemum.


2020 ◽  
Author(s):  
Xiya Zuo ◽  
Shixiang Wang ◽  
Wen Xiang ◽  
Huiru Yang ◽  
Muhammad Mobeen Tahir ◽  
...  

Abstract Background: Apple ( Malus domestica Borkh.) is one of the most popular cultivated fruit crops in China. Apple floral transition is an important process but liable to be affected by various environmental factors. The 14-3-3 proteins are involved in regulating diverse biological processes in plants, and some 14-3-3 members have been identified to play vital roles in flowering. However, little information was available about the 14-3-3 members in apple. Results: In the current study, we identified eighteen 14-3-3 gene family members from apple genome database, designated MdGF14a to MdGF14r . The isoforms possess a conserved core region composed of nine antiparallel α-helices and divergent N and C termini. According to their structural and phylogenetic features, Md14-3-3s classified into two major evolutionary branches, the epsilon (ɛ) group and the non-epsilon (non-ɛ) group. Moreover, expression profiles derived from transcriptome data and quantitative reverse-transcription PCR (qRT-PCR) analysis exhibited diverse expression patterns of Md14-3-3 genes in various tissues and in response to different sugars and hormones treatments during floral transition phase. Four Md14-3-3 isoforms ( MdGF14a , MdGF14d , MdGF14i and MdGF14j ) exhibiting prominent transcriptional responses to sugars and hormones were selected for further investigation. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation (BiFC) experiments showed that the four Md14-3-3 proteins interact with key floral integrators, MdTFL1 (TERMINAL FLOWER1) and MdFT (FLOWERING LOCUS T). Subcellular localization of four selected Md14-3-3 proteins demonstrated their localization in both the cytoplasm and nucleus. Conclusion: We comprehensively identified Md14-3-3s family in apple. Some Md14-3-3 genes are predominantly expressed during apple floral transition stage, and may participate in regulation of flowering through association with flower control genes. Our results provide a preliminary framework for further investigation into the roles of Md14-3-3s for floral transition.


2020 ◽  
Author(s):  
Xiya Zuo ◽  
Shixiang Wang ◽  
Wen Xiang ◽  
Muhammad Mobeen Tahir ◽  
Huiru Yang ◽  
...  

Abstract Background: Apple ( Malus domestica Borkh.) is one of the most popular cultivated fruit crops in China. Apple floral transition is an important process but liable to be affected by various environmental factors. The 14-3-3 proteins are involved in regulating diverse biological processes in plants, and some 14-3-3 members have been identified to play vital roles in flowering. However, little information was available about the 14-3-3 members in apple. Results: In the current study, we identified eighteen 14-3-3 gene family members from apple genome database, designated MdGF14a to MdGF14r , 17 of them are transcribed. The isoforms possess a conserved core region composed of nine antiparallel α-helices and divergent N and C termini. According to their structural and phylogenetic features, Md14-3-3s classified into two major evolutionary branches, the epsilon (ɛ) group and the non-epsilon (non-ɛ) group. Moreover, expression profiles derived from transcriptome data and quantitative reverse-transcription PCR (qRT-PCR) analysis exhibited diverse expression patterns of Md14-3-3 genes in various tissues and in response to different sugars and hormones treatments during floral transition phase. Four Md14-3-3 isoforms ( MdGF14a , MdGF14d , MdGF14i and MdGF14j ) exhibiting prominent transcriptional responses to sugars and hormones were selected for further investigation. Subcellular localization of four selected Md14-3-3 proteins demonstrated their localization in both the cytoplasm and nucleus. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation (BiFC) experiments showed that the four Md14-3-3 proteins interact with key floral integrators, MdTFL1 (TERMINAL FLOWER1) and MdFT (FLOWERING LOCUS T). Conclusion: We comprehensively identified Md14-3-3s family in apple. Some Md14-3-3 genes are predominantly expressed during apple flowering transition stage, and may participate in regulation of flowering through association with flower control genes. Our results provide a preliminary framework for further investigation into the roles of Md14-3-3s for flower transition.


2020 ◽  
Author(s):  
Xiya Zuo ◽  
Shixiang Wang ◽  
Wen Xiang ◽  
Huiru Yang ◽  
Muhammad Mobeen Tahir ◽  
...  

Abstract Background: Apple (Malus domestica Borkh.) is one of the most popular cultivated fruit crops in China. Apple floral transition is an important process but liable to be affected by various environmental factors. The 14-3-3 proteins are involved in regulating diverse biological processes in plants, and some 14-3-3 members play vital roles in flowering. However, little information was available about the 14-3-3 members in apple.Results: In the current study, we identified eighteen 14-3-3 gene family members from the apple genome database, designated MdGF14a to MdGF14r. The isoforms possess a conserved core region comprising nine antiparallel α-helices and divergent N and C termini. According to their structural and phylogenetic features, Md14-3-3 proteins could be classified into two major evolutionary branches, the epsilon (ɛ) group and the non-epsilon (non-ɛ) group. Moreover, expression profiles derived from transcriptome data and quantitative real-time reverse transcription PCR analysis showed diverse expression patterns of Md14-3-3 genes in various tissues and in response to different sugars and hormone treatments during the floral transition phase. Four Md14‑3-3 isoforms (MdGF14a, MdGF14d, MdGF14i, and MdGF14j) exhibiting prominent transcriptional responses to sugars and hormones were selected for further investigation. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation experiments showed that the four Md14-3-3 proteins interact with key floral integrators, MdTFL1 (TERMINAL FLOWER1) and MdFT (FLOWERING LOCUS T). Subcellular localization of four selected Md14-3-3 proteins demonstrated their localization in both the cytoplasm and nucleus.Conclusion: We identified the Md14-3-3s family in apple comprehensively. Certain Md14-3-3 genes are expressed predominantly during the apple floral transition stage, and may participate in the regulation of flowering through association with flower control genes. Our results provide a preliminary framework for further investigation into the roles of Md14-3-3s in floral transition.


Genes ◽  
2019 ◽  
Vol 10 (10) ◽  
pp. 777 ◽  
Author(s):  
Yan Hong ◽  
Mengling Li ◽  
Silan Dai

The generation of chrysanthemum (Chrysanthemum × morifolium) flower color is mainly attributed to the accumulation of anthocyanins. In the anthocyanin biosynthetic pathway in chrysanthemum, although all of the structural genes have been cloned, the regulatory function of R2R3-MYB transcription factor (TF) genes, which play a crucial role in determining anthocyanin accumulation in many ornamental crops, still remains unclear. In our previous study, four light-induced R2R3-MYB TF genes in chrysanthemum were identified using transcriptomic sequencing. In the present study, we further investigated the regulatory functions of these genes via phylogenetic and alignment analyses of amino acid sequences, which were subsequently verified by phenotypic, pigmental, and structural gene expression analyses in transgenic tobacco lines. As revealed by phylogenetic and alignment analyses, CmMYB4 and CmMYB5 were phenylpropanoid and flavonoid repressor R2R3-MYB genes, respectively, while CmMYB6 was an activator of anthocyanin biosynthesis, and CmMYB7 was involved in regulating flavonol biosynthesis. Compared with wild-type plants, the relative anthocyanin contents in the 35S:CmMYB4 and 35S:CmMYB5 tobacco lines significantly decreased (p < 0.05), while for 35S:CmMYB6 and 35S:CmMYB7, the opposite result was obtained. Both in the 35S:CmMYB4 and 35S:CmMYB5 lines, the relative expression of several anthocyanin biosynthetic genes in tobacco was significantly downregulated (p < 0.05); on the contrary, several genes were upregulated in the 35S:CmMYB6 and 35S:CmMYB7 lines. These results indicate that CmMYB4 and CmMYB5 negatively regulate anthocyanin biosynthesis in chrysanthemum, while CmMYB6 and CmMYB7 play a positive role, which will aid in understanding the complex mechanism regulating floral pigmentation in chrysanthemum and the functional divergence of the R2R3-MYB gene family in higher plants.


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