BZR1 Antibody
Description
Introduction to BZR1 Antibody
BZR1 antibodies are polyclonal or monoclonal reagents designed to recognize the BZR1 protein, which coordinates BR-mediated gene expression. BZR1 acts as a transcriptional repressor/activator, modulating BR biosynthesis and signaling while integrating environmental cues . Antibodies against BZR1 are critical for:
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Detecting BZR1 protein levels in plant tissues via Western blotting.
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Localizing BZR1 in cellular compartments (e.g., nucleus vs. cytoplasm).
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Studying post-translational modifications (e.g., phosphorylation) .
Immunogen Design
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Agrisera AS16 3219: Targets a synthetic peptide derived from Oryza sativa BZR1 (UniProt: Q7XI96) .
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PhytoAB PHY7824S: Developed using Arabidopsis thaliana BZR1 (AT1G75080) .
Cross-Reactivity
Validated reactivity spans multiple plant species:
| Species | Confirmed Reactivity |
|---|---|
| Arabidopsis | Yes |
| Oryza sativa | Yes |
| Zea mays | Predicted |
| Hordeum vulgare | Predicted |
Mechanistic Insights into BR Signaling
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Phosphorylation Regulation: BZR1 antibodies identified BIN2 kinase-mediated phosphorylation, which destabilizes BZR1 via proteasomal degradation .
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Transcriptional Repression: BZR1 recruits TOPLESS (TPL) corepressors via its EAR motif, enabling histone deacetylase (HDA)-dependent gene silencing .
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Immune-Growth Trade-off: BZR1 antibodies confirmed its interaction with WRKY40, suppressing pathogen-triggered immunity to prioritize growth during stress .
Genome-Wide Studies
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ChIP-chip Analysis: BZR1 antibodies facilitated chromatin immunoprecipitation, revealing 953 BR-regulated BZR1 target genes, including WRKY transcription factors .
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Protein Interaction Networks: Co-immunoprecipitation (Co-IP) identified BZR1 partners like MAPK4, MKK5, and PP2A, linking BR signaling to stress responses .
Table 1: BZR1 Functional Roles
Table 2: Antibody Performance in Key Studies
| Study Focus | Antibody Used | Outcome |
|---|---|---|
| BZR1-TPL Interaction | PhytoAB PHY7824S | Confirmed EAR motif-dependent binding |
| BZR1 Phosphorylation Dynamics | Agrisera AS16 3219 | Detected dephosphorylation post-BL treatment |
Limitations and Considerations
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- The brassinosteroid-regulated transcription factors BZR1/BES1 act as coordinators in multisignal-regulated plant growth. PMID: 29673687
- Light-regulated phosphorylation changes of BZR1 depend on the presence of endogenous brassinosteroid and functional BIN2 and protein phosphatase 2A (PP2A). PMID: 28396515
- BZR1, a homologue of BES1, promotes cell divisions in the quiescent center (QC) while suppressing columella stem cell differentiation. This action contrasts with that of BES1. PMID: 26136267
- BZR1 represses target genes by recruiting the Groucho/TUP1-like transcriptional corepressor TOPLESS. PMID: 24938363
- BZR1 serves as a significant regulator mediating the trade-off between growth and immunity upon integration of environmental cues. PMID: 24381244
- BIL1/BZR1 enhances plant resistance to insect feeding. PMID: 25036120
- BZR1-interacting proteins have been identified as potential components of the brassinosteroid signaling pathway in Arabidopsis. PMID: 24019147
- Phosphatidic acid reduces BZR1 dephosphorylation, inhibiting BZR1 activity and brassinosteroid signal response by regulating PP2A activity. PMID: 24121289
- BZR1 and AP2 likely influence Arabidopsis ovule number determination antagonistically. PMID: 22914576
- BZR1 dissociates from the DWF4 (cytochrome P450 90B1) promoter in the presence of auxin, indicating that DWF4 expression involves BZR1-mediated de-repression mechanisms. PMID: 21284753
- Studies reveal that BZR1 not only controls the expression of numerous signaling components of other hormonal and light pathways, but also co-regulates common target genes with light-signaling transcription factors. PMID: 21074725
- Findings demonstrate that BZR1 coordinates brassinosteroid (BR) homeostasis and signaling by playing dual roles in regulating BR biosynthesis and downstream growth responses. PMID: 15681342
- BZR1 functions as a nucleocytoplasmic shuttling protein, and GSK3-like kinases induce the nuclear export of BZR1 by modulating BZR1 interaction with the 14-3-3 proteins. PMID: 17873094
Q&A
FAQs for BZR1 Antibody in Academic Research
What experimental systems are optimal for studying BZR1 phosphorylation dynamics?
Use tissue-specific reporters combined with phospho-mimetic/-dead mutants:
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Hypocotyl elongation assays: Monitor BZR1-T177 phosphorylation status using Phos-tag™ gels in Arabidopsis etiolated seedlings .
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Bimolecular fluorescence complementation (BiFC): Visualize BZR1-14-3-3 interactions in epidermal cells under BR/auxin treatments .
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Live-cell imaging: Track BZR1-GFP nuclear-cytoplasmic shuttling in root tips using ProBZR1:BZR1-GFP lines .
Key phosphorylation sites:
| Site | Function | Regulatory Kinase |
|---|---|---|
| T177 | 14-3-3 binding, nuclear export | BIN2 |
| S173 | DNA binding attenuation | MPK3/6 |
| Y200 | Protein stability modulation | Unknown |
How to resolve contradictory data on BZR1’s role as transcriptional activator vs. repressor?
Context-dependent mechanisms explain apparent contradictions:
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Repressor function: BZR1 recruits TOPLESS/TPL via its EAR motif to downregulate BR biosynthesis genes (e.g., CPD, DWF4) .
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Activator function: BZR1 binds BIC1 coactivator to induce cell elongation genes (e.g., PRE1, EXP8) .
Methodological recommendations:
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Perform cell-type-specific ATAC-seq to map chromatin accessibility changes .
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Use dominant-negative TPL variants to uncouple repression effects .
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Conduct time-resolved RNA-seq (0–120 min post-BR treatment) to distinguish primary vs. secondary targets .
What controls are critical for co-immunoprecipitation (Co-IP) studies of BZR1 protein complexes?
Include five essential controls to minimize false positives:
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Empty vector IP: Express bait/prey proteins separately in N. benthamiana .
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Crosslinking validation: Compare results with/without 1% formaldehyde fixation .
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Phosphatase treatment: Assess phosphorylation-dependent interactions using λ-phosphatase .
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Compartment-specific IP: Fractionate nuclear/cytosolic extracts before IP .
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Endogenous competition: Spike recombinant BZR1 into WT extracts during IP .
Commonly identified interactors:
| Protein | Function | Validation Method |
|---|---|---|
| WRKY40 | Immune suppression | Y2H, Co-IP |
| PIF4 | Thermomorphogenesis | BiFC, Co-IP |
| PP2A (B’ subunit) | Dephosphorylation | TAP-MS, Genetic |
How to design experiments investigating BZR1’s role in growth-immunity trade-offs?
Adopt a multi-omics approach:
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Parallel phenotyping: Quantify ROS bursts (PTI) and hypocotyl lengths under varying BR/FLG22 ratios .
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Dual RNA-seq/ChIP-seq: Profile BZR1 targets in BZR1-ox vs. bak1-5 mutants .
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Metabolite profiling: Link transcriptional changes to JA/SA pathway intermediates .
Key findings from integrated studies:
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BZR1 suppresses RBOHD-mediated ROS by upregulating WRKY11/15/40 .
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BR-induced growth reduces callose deposition by 78% in Pseudomonas syringae infections .
What advanced techniques address BZR1 antibody limitations in plant-fungus pathosystems?
Implement cross-species compatibility testing:
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Epitope mapping: Compare BZR1 orthologs in Fusarium oxysporum-infected tomatoes using peptide arrays .
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Single-domain antibodies: Engineer camelid VHH antibodies for enhanced penetration in lignified tissues .
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Proximity ligation (PLA): Detect BZR1-WRKY40 complexes in arbuscular mycorrhiza-colonized roots .
Performance metrics:
| Method | Resolution | Tissue Compatibility | Background Reduction |
|---|---|---|---|
| Conventional IgG | 5–10 µm | Low (high autofluorescence) | 30% |
| VHH-nanobody | 1–2 µm | High | 85% |
How to troubleshoot nonspecific signals in BZR1 immunohistochemistry?
Apply three orthogonal strategies:
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Pre-absorption blocking: Incubate antibody with bzr1-1D mutant protein extracts .
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Multiplexed imaging: Combine with ProBZR1:GUS histochemical staining for spatial validation .
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Algorithmic unmixing: Use HALO® or Visiopharm® to separate signal from chloroplast autofluorescence .
Quantitative benchmarks:
| Tissue Type | Signal-to-Noise Ratio (Pre-optimization) | Post-optimization |
|---|---|---|
| Mature leaf | 2.1 | 8.7 |
| Root tip | 4.5 | 12.3 |
