scholarly journals Hormonal Regulatory Patterns of LaKNOXs and LaBEL1 Transcription Factors Reveal Their Potential Role in Stem Bulblet Formation in LA Hybrid Lily

2021 ◽  
Vol 22 (24) ◽  
pp. 13502
Author(s):  
Yue Zhang ◽  
Zhen Zeng ◽  
Yubing Yong ◽  
Yingmin Lyu
Keyword(s):  
Potential Role ◽  
Transgenic Arabidopsis ◽  
Expression Patterns ◽  
Relative Increase ◽  
Bimolecular Fluorescence Complementation ◽  
Mechanism Of Formation ◽  
Yeast Two Hybrid ◽  
Physical Interactions ◽  
Two Hybrid ◽  
Strong Capacity

In lily reproduction, the mechanism of formation of bulbs has been a hot topic. However, studies on stem bulblet formation are limited. Stem bulblets, formed in the leaf axils of under- and above-ground stems, provide lilies with a strong capacity for self-propagation. First, we showed that above-ground stem bulblets can be induced by spraying 100 mg/L 6-BA on the LA hybrid lily ‘Aladdin’, with reduced endogenous IAA and GA4 and a higher relative content of cytokinins. Then, expression patterns of three potential genes (two KNOTTED1-like homeobox (KNOX) and one partial BEL1-like homeobox (BELL)), during stem bulblet formation from our previous study, were determined by RT-qPCR, presenting a down-up trend in KNOXs and a rising tendency in BELL. The partial BELL gene was cloned by RACE from L. ‘Aladdin’ and denoted LaBEL1. Physical interactions of LaKNOX1-LaBEL1 and LaKNOX1-LaKNOX2 were confirmed by yeast two-hybrid and bimolecular fluorescence complementation assays. Furthermore, hormonal regulatory patterns of single LaKNOX1, LaKNOX2, LaBEL1, and their heterodimers, were revealed in transgenic Arabidopsis, suggesting that the massive mRNA accumulations of LaKNOX1, LaKNOX2 and LaBEL1 genes during stem bulblet formation could cause the dramatic relative increase of cytokinins and the decline of GAs and IAA. Taken together, a putative model was proposed that LaKNOX1 interacts with LaKNOX2 and LaBEL1 to regulate multiple phytohormones simultaneously for an appropriate hormonal homeostasis, which suggests their potential role in stem bulblet formation in L. ‘Aladdin’.

scholarly journals Regulation of Glycosylphosphatidylinositol-Anchored Protein (GPI-AP) Expression by F-Box/LRR-Repeat (FBXL) Protein in Wheat (Triticum aestivum L.)

Plants ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1606
Author(s):  
Min Jeong Hong ◽  
Jin-Baek Kim ◽  
Yong Weon Seo ◽  
Dae Yeon Kim
Keyword(s):  
Triticum Aestivum ◽  
Plasma Membrane ◽  
Fluorescent Protein ◽  
Expression Patterns ◽  
Triticum Aestivum L ◽  
26S Proteasome ◽  
Bimolecular Fluorescence Complementation ◽  
Yeast Two Hybrid ◽  
Scf Complex ◽  
Two Hybrid

F-box proteins are substrate recognition components of the Skp1-Cullin-F-box (SCF) complex, which performs many important biological functions including the degradation of numerous proteins via the ubiquitin–26S proteasome system. In this study, we isolated the gene encoding the F-box/LRR-repeat (FBXL) protein from wheat (Triticum aestivum L.) seedlings and validated that the TaFBXL protein is a component of the SCF complex. Yeast two-hybrid assays revealed that TaFBXL interacts with the wheat glycosylphosphatidylinositol-anchored protein (TaGPI-AP). The green fluorescent protein (GFP) fusion protein of TaFBXL was detected in the nucleus and plasma membrane, whereas that of TaGPI-AP was observed in the cytosol and probably also plasma membrane. yeast two-hybrid and bimolecular fluorescence complementation (BiFC) assays revealed that TaFBXL specifically interacts with TaGPI-AP in the nucleus and plasma membrane, and TaGPI-AP is targeted by TaFBXL for degradation via the 26S proteasome system. In addition, TaFBXL and TaGPI-AP showed antagonistic expression patterns upon treatment with indole-3-acetic acid (IAA), and the level of TaGPI-AP was higher in tobacco leaves treated with both MG132 (proteasome inhibitor) and IAA than in leaves treated with either MG132 or IAA. Taken together, our data suggest that TaFBXL regulates the TaGPI-AP protein level in response to exogenous auxin application.

scholarly journals Key role of the motor protein Kinesin 13B in the activity of homeodomain-leucine zipper I transcription factors

2020 ◽  
Vol 71 (20) ◽  
pp. 6282-6296
Author(s):  
Virginia Natali Miguel ◽  
Karina Fabiana Ribichich ◽  
Jorge Ignacio Giacomelli ◽  
Raquel Lia Chan
Keyword(s):  
Transcription Factors ◽  
Leucine Zipper ◽  
Molecular Mechanisms ◽  
Vascular Tissue ◽  
Expression Patterns ◽  
Motor Protein ◽  
Bimolecular Fluorescence Complementation ◽  
Yeast Two Hybrid ◽  
Cauline Leaf ◽  
Two Hybrid

Abstract The sunflower (Helianthus annuus) homeodomain-leucine zipper I transcription factor HaHB11 conferred differential phenotypic features when it was expressed in Arabidopsis, alfalfa, and maize plants. Such differences were increased biomass, seed yield, and tolerance to flooding. To elucidate the molecular mechanisms leading to such traits and identify HaHB11-interacting proteins, a yeast two-hybrid screening of an Arabidopsis cDNA library was carried out using HaHB11 as bait. The sole protein identified with high confidence as interacting with HaHB11 was Kinesin 13B. The interaction was confirmed by bimolecular fluorescence complementation and by yeast two-hybrid assay. Kinesin 13B also interacted with AtHB7, the Arabidopsis closest ortholog of HaHB11. Histochemical analyses revealed an overlap between the expression patterns of the three genes in hypocotyls, apical meristems, young leaves, vascular tissue, axillary buds, cauline leaves, and cauline leaf nodes at different developmental stages. AtKinesin 13B mutants did not exhibit a differential phenotype when compared with controls; however, both HaHB11 and AtHB7 overexpressor plants lost, partially or totally, their differential phenotypic characteristics when crossed with such mutants. Altogether, the results indicated that Kinesin 13B is essential for the homeodomain-leucine zipper transcription factors I to exert their functions, probably via regulation of the intracellular distribution of these transcription factors by the motor protein.

scholarly journals The role of Clt1-regulated-Xylan metabolism in the melanin and toxin formation for the pathogenicity of Curvularia lunata in Maize

2021 ◽  
Author(s):  
Jinxin Gao ◽  
Jie Chen
Keyword(s):  
Toxin Production ◽  
Bimolecular Fluorescence Complementation ◽  
Curvularia Lunata ◽  
Yeast Two Hybrid ◽  
Acetyl Xylan Esterase ◽  
Btb Domain ◽  
Intermediate Metabolites ◽  
Regulative Mechanism ◽  
Two Hybrid

We previously reported that the BTB domain-containing protein Clt1 regulates melanin and toxin synthesis, conidiation, and pathogenicity in Curvularia lunata, but the interacting proteins and regulative mechanism of Clt1 are unclear. In this research, we identified two proteins, which respectively correspond to xylanase (Clxyn24) and acetyl xylan esterase (Claxe43) from C. lunata were regulated by Clt1. Yeast two-hybrid (Y2H), and bimolecular fluorescence complementation assays were conducted to verify the interaction of Clt1 with full-length Clxyn24 and Claxe43. Furthermore, the Y2H assay revealed that Clt1 physically interacted with Clxyn24 and Claxe43 through its BTB domain to degrade xylan which was used as a carbon source for C. lunata growth. The utilization of xylan provides acetyl-CoA for the synthesis of melanin and toxin, as well as energy and other intermediate metabolites for conidiation. Furthermore, transcriptome analysis revealed that PKS18 and its 13 flanking genes are found clustered in a region spanning 57.89 kb on scaffold 9 of the C. lunata CX-3 genome were down-regulated in toxin production deficient mutant T806, and this cluster is possibly responsible for toxin biosynthesis of C. lunata.

scholarly journals Identification of Beet necrotic yellow vein virus P25 Pathogenicity Factor–Interacting Sugar Beet Proteins That Represent Putative Virus Targets or Components of Plant Resistance

2009 ◽  
Vol 22 (8) ◽  
pp. 999-1010 ◽  
Author(s):  
Heike Thiel ◽  
Mark Varrelmann
Keyword(s):  
Sugar Beet ◽  
Resistance Mechanism ◽  
Yellow Vein ◽  
Bimolecular Fluorescence Complementation ◽  
Plant Proteins ◽  
Yeast Two Hybrid ◽  
Response To Stress ◽  
Bimolecular Fluorescence Complementation Assay ◽  
Two Hybrid ◽  
Important Disease

Beet necrotic yellow vein virus (BNYVV) induces the most important disease threatening sugar beet. The growth of partially resistant hybrids carrying monogenic dominant resistance genes stabilize yield but are unable to entirely prevent virus infection and replication. P25 is responsible for symptom development and previous studies have shown that recently occurring resistance-breaking isolates possess increased P25 variability. To better understand the viral pathogenicity factor's interplay with plant proteins and to possibly unravel the molecular basis of sugar beet antivirus resistance, P25 was applied in a yeast two-hybrid screen of a resistant sugar beet cDNA library. This screen identified candidate proteins recognized as orthologues from other plant species which are known to be expressed following pathogen infection and involved in plant defense response. Most of the candidates potentially related to host-pathogen interactions were involved in the ubiquitylation process and plants response to stress, and were part of cell and metabolism components. The interaction of several candidate genes with P25 was confirmed in Nicotiana benthamiana leaf cells by transient agrobacterium-mediated expression applying bimolecular fluorescence complementation assay. The putative functions of several of the candidates identified support previous findings and present first targets for understanding the BNYVV pathogenicity and antivirus resistance mechanism.

scholarly journals Silencing Suppressor Protein VPg of a Potyvirus Interacts With the Plant Silencing-Related Protein SGS3

2014 ◽  
Vol 27 (11) ◽  
pp. 1199-1210 ◽  
Author(s):  
Minna-Liisa Rajamäki ◽  
Janne Streng ◽  
Jari P. T. Valkonen
Keyword(s):  
Rna Silencing ◽  
Bimolecular Fluorescence Complementation ◽  
Mrna Levels ◽  
Yeast Two Hybrid ◽  
Main Determinant ◽  
Potato Virus A ◽  
Cytoplasmic Bodies ◽  
Multiple Protein ◽  
Suppressor Of Rna Silencing ◽  
Two Hybrid

Viral genome-linked protein (VPg) of potyviruses is involved in multiple steps of the potyvirus infection cycle, including viral multiplication and movement in plants. Recently, we showed that VPg of Potato virus A (PVA; genus Potyvirus) suppresses sense-mediated RNA silencing, which is linked to one or both nuclear or nucleolar localization. Here, we studied interactions between VPg and components of the plant RNA silencing pathway. Results showed that VPg interacts with the SGS3 protein of Solanum tuberosum and Arabidopsis thaliana, as shown by yeast two-hybrid analysis and bimolecular fluorescence complementation assays. VPg–SGS3 interactions co-localized with small cytoplasmic bodies that contained plant RNA-dependent RNA polymerase 6 (RDR6) (likely SGS3/RDR6 bodies). The N-terminal zinc finger (ZF) domain of SGS3 was the main determinant of the VPg interaction. Our data also suggest that the ZF domain controls SGS3 localization. SGS3 homodimerization was controlled by multiple protein regions. The VPg–SGS3 interaction appeared beneficial for PVA, as viral RNA levels correlated positively with sgs3 mRNA levels in the SGS3-silenced and SGS3-overexpressing leaves of Nicotiana benthamiana. The data support the idea that VPg acts as a suppressor of RNA silencing and suggest that an interaction with SGS3 may be important, especially in suppression of sense-mediated RNA silencing.

scholarly journals Genome-Wide Analysis of the IQM Gene Family in Rice (Oryza sativa L.)

Plants ◽  
2021 ◽  
Vol 10 (9) ◽  
pp. 1949
Author(s):  
Tian Fan ◽  
Tianxiao Lv ◽  
Chuping Xie ◽  
Yuping Zhou ◽  
Changen Tian
Keyword(s):  
Abiotic Stress ◽  
Stress Response ◽  
Gene Family ◽  
Expression Profiles ◽  
Expression Patterns ◽  
Abiotic Stress Response ◽  
Biotic And Abiotic Stress ◽  
Yeast Two Hybrid ◽  
Iq Motif ◽  
Two Hybrid

Members of the IQM (IQ-Motif Containing) gene family are involved in plant growth and developmental processes, biotic and abiotic stress response. To systematically analyze the IQM gene family and their expression profiles under diverse biotic and abiotic stresses, we identified 8 IQM genes in the rice genome. In the current study, the whole genome identification and characterization of OsIQMs, including the gene and protein structure, genome localization, phylogenetic relationship, gene expression and yeast two-hybrid were performed. Eight IQM genes were classified into three subfamilies (I–III) according to the phylogenetic analysis. Gene structure and protein motif analyses showed that these IQM genes are relatively conserved within each subfamily of rice. The 8 OsIQM genes are distributed on seven out of the twelve chromosomes, with three IQM gene pairs involved in segmental duplication events. The evolutionary patterns analysis revealed that the IQM genes underwent a large-scale event within the last 20 to 9 million years. In addition, quantitative real-time PCR analysis of eight OsIQMs genes displayed different expression patterns at different developmental stages and in different tissues as well as showed that most IQM genes were responsive to PEG, NaCl, jasmonic acid (JA), abscisic acid (ABA) treatment, suggesting their crucial roles in biotic, and abiotic stress response. Additionally, a yeast two-hybrid assay showed that OsIQMs can interact with OsCaMs, and the IQ motif of OsIQMs is required for OsIQMs to combine with OsCaMs. Our results will be valuable to further characterize the important biological functions of rice IQM genes.

scholarly journals Drosophila homologs of the proto-oncogene product PEBP2/CBF beta regulate the DNA-binding properties of Runt.

1996 ◽  
Vol 16 (3) ◽  
pp. 932-942 ◽  
Author(s):  
G Golling ◽  
L Li ◽  
M Pepling ◽  
M Stebbins ◽  
J P Gergen
Keyword(s):  
Dna Binding ◽  
Expression Patterns ◽  
Yeast Two Hybrid ◽  
Binding Studies ◽  
Runt Domain ◽  
Big Brother ◽  
Functional Homology ◽  
Two Hybrid

The Drosophila runt gene is the founding member of the Runt domain family of transcriptional regulators. Mammalian Runt domain genes encode the alpha subunit of the heterometric DNA-binding factor PEBP2/CBF. The unrelated PEBP2/CBF beta protein interacts with the Runt domain to increase its affinity for DNA. The conserved ability of the Drosophila Runt protein to respond to the stimulating effect of mammalian PEBP2/CBF beta indicated that flies were likely to have a homologous beta protein. Using the yeast two-hybrid system to isolate cDNAs for Runt-interacting proteins, we identified two Drosophila genes, referred to as Brother and Big-brother, that have substantial sequence homology with PEBP2/CBF beta. Yeast two-hybrid experiments as well as in vitro DNA-binding studies confirmed the functional homology of the Brother, Big-brother, and PEBP2/CBF beta proteins and demonstrated that the conserved regions of the Runt and Brother proteins are required for their heterodimeric interaction. The DNA-bending properties of Runt domain proteins in the presence and absence of their partners were also examined. Our results show that Runt domain proteins bend DNA and that this bending is influenced by Brother protein family members, supporting the idea that heterodimerization is associated with a conformational change in the Runt domain. Analysis of expression patterns in Drosophila embryos revealed that Brother and Big-brother are likely to interact with runt in vivo and further suggested that the activity of these proteins is not restricted to their interaction with Runt.

scholarly journals Screening and Identification of Host Proteins Interacting with Iris lactea var. chinensis Metallothionein IlMT2a by Yeast Two-Hybrid Assay

Genes ◽  
2021 ◽  
Vol 12 (4) ◽  
pp. 554
Author(s):  
Zhiquan Wang ◽  
Longjie Ni ◽  
Liangqin Liu ◽  
Haiyan Yuan ◽  
Suzhen Huang ◽  
...  
Keyword(s):  
Stress Responses ◽  
Transgenic Arabidopsis ◽  
Biological Significance ◽  
Significant Protection ◽  
Yeast Two Hybrid ◽  
Host Proteins ◽  
Screening And Identification ◽  
First Time ◽  
Gene Regulations ◽  
Two Hybrid

Iris lactea var. chinensis (Fisch.) (I. lactea var. chinensis) is a well-known cadmium (Cd)-tolerant plant and we have previously shown that the metallothionein gene, IlMT2a, of the plant may be playing a key role in conferring the Cd tolerance. In this study, we have identified several proteins interacting with the IlMT2a by screening yeast two-hybrid library constructed from cDNAs isolated from Cd-treated I. lacteal var. chinensis plants. Putative functions of these proteins include those involved in photosynthesis, ROS scavenge, nutrient transport, and transcriptional regulation, to name a few. In particular, another metallothionein, which we assigned the name of IlMT3, was identified as an interacting partner of the IlMT2a. Unlike IlMT2a, it did not provide any significant protection against Cd toxicity in transgenic Arabidopsis thaliana L. (A. thaliana). To our knowledge, this is the first time ever reporting the interaction of two metallothionein proteins in plants. Learning the biological significance of the interaction between IlMT2a and IlMT3 would be the focus of future study and would be able to provide valuable insights into the understanding plant metallothionein’s diverse and complex roles in coordinating many important cellular physiologies including stress responses, gene regulations, and energy metabolisms.

scholarly journals CmMLO17 and its partner CmKIC potentially support Alternaria alternata growth in Chrysanthemum morifolium

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Jingjing Xin ◽  
Ye Liu ◽  
Huiyun Li ◽  
Sumei Chen ◽  
Jiafu Jiang ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Alternaria Alternata ◽  
Defense Responses ◽  
Chrysanthemum Morifolium ◽  
Bimolecular Fluorescence Complementation ◽  
Resistance Locus ◽  
Plant Defense Responses ◽  
Yeast Two Hybrid ◽  
Two Hybrid ◽  
Insight Into

AbstractThe Mildew Resistance Locus O (MLO) gene family has been investigated in many species. However, there are few studies on chrysanthemum MLO genes. We report in this study that CmMLO17 in Chrysanthemum morifolium was upregulated after Alternaria alternata infection. Silencing of CmMLO17 by artificial microRNA resulted in reduced susceptibility of chrysanthemum to A. alternata infection. Genes in the abscisic acid (ABA) and Ca2+ signaling pathways were upregulated in the CmMLO17-silenced line R20 compared to the wild-type plants. We speculated that CmMLO17-silenced plants had a faster and stronger defense response that was mediated by the ABA and Ca2+ signaling pathways, resulting in reduced susceptibility of chrysanthemum to A. alternata infection. In addition, a candidate gene, CmKIC, that may interact with CmMLO17 was discovered by the yeast two-hybrid assay. The interaction between CmMLO17 and CmKIC was confirmed using the yeast two-hybrid assay and bimolecular fluorescence complementation (BiFC) analysis. CmMLO17 and CmKIC were both located on the plasma membrane, and CmKIC was also located on the nucleus. CmKIC overexpression increased the susceptibility of chrysanthemum to A. alternata, whereas CmKIC silencing resulted in reduced susceptibility. Therefore, CmMLO17 and CmKIC may work together in C. morifolium to support the growth of A. alternata. The results of this study will provide insight into the potential function of MLO and improve the understanding of plant defense responses to necrotrophic pathogens.

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