ATX1 Antibody
Description
Definition and Target Specificity
ATX1 antibodies are designed to bind ATX1 proteins, which differ functionally across species:
2.1. Arabidopsis ATX1 in Chromatin Regulation
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Dual Regulatory Role: ATX1 activates ~780 genes and represses ~860 genes in Arabidopsis, impacting transcription factors, stress responses, and development .
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Mechanistic Insights:
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Binds phosphatidylinositol 5-phosphate (PI5P), linking lipid signaling to chromatin remodeling .
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Facilitates RNA Polymerase II elongation by establishing H3K4me3 marks downstream of transcription start sites .
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Regulates drought-responsive genes (e.g., WRKY70, NCED3), with 30% of targets encoding plasma membrane/cell wall functions .
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2.2. Chlamydomonas ATX1 in Metal Homeostasis
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Copper Chaperone Activity: Delivers Cu(I) to post-Golgi vesicles for iron uptake oxidase assembly, critical under iron-limited conditions .
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Antibody Validation: Knockdown (ami-atx1) and knockout (atx1-1, atx1-2) lines confirmed ATX1’s role in growth during iron deficiency .
2.3. Human ATOX1/Ataxin-1 in Disease
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Neurodegeneration: Mutant Ataxin-1 (polyglutamine-expanded) interacts with SMRT/HDAC3 repressors, causing transcriptional dysregulation in spinocerebellar ataxia .
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Subcellular Localization: Ataxin-1 associates with splicing factors (e.g., U2AF65) and forms nuclear aggregates linked to toxicity .
3.2. Experimental Workflows
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Western Blotting: Used to assess ATX1 expression in Chlamydomonas under iron-limited conditions .
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Chromatin Immunoprecipitation (ChIP): Identified ATX1-binding regions at H3K4me3-enriched loci in Arabidopsis .
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Immunofluorescence: Localized Ataxin-1 to nuclear aggregates in human cell models .
Technical Considerations
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Cross-Reactivity: Antibodies like PHY1582S are specific to Chlamydomonas ATX1, with no cross-reactivity reported in Arabidopsis or human systems .
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Batch Consistency: Recombinant monoclonal antibodies (e.g., 83785-1-PBS) ensure high reproducibility for quantitative assays .
Future Directions
Product Specs
Composition: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
References:
- The critical role of ATX1 in copper (Cu) homeostasis, conferring tolerance to both excess and deficiency, is well-established. PMID: 22555879
Q&A
Basic Research Questions
How to validate ATX1 antibody specificity in plant tissue samples?
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Perform Western blot with atx1 knockout mutants as negative controls (wild-type vs mutant comparisons)
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Use peptide blocking assays: Pre-incubate antibody with immunogen peptide to test signal disappearance
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Combine with mass spectrometry verification of immunoprecipitated proteins
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Recommended controls:
Control Type Purpose Expected Result Wild-type tissue Positive signal Clear band at ~150 kDa (full-length ATX1) atx1 mutant Specificity test No band at 150 kDa Secondary antibody-only Background check No bands
What experimental factors affect ATX1 subcellular localization studies?
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Fixation method: Use fresh formaldehyde (1-2%) for nuclear-cytoplasmic localization preservation
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Developmental stage: Localization shifts occur between:
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Phospholipid interactions: PI5P treatment causes nuclear-to-cytoplasmic relocalization within 95 mins
Advanced Research Challenges
3. Resolving contradictory data on ATX1's transcriptional roles
Context: ATX1 shows both activation (H3K4me3 deposition) and repression functions
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Experimental approach:
Interpreting unexpected 22 kDa band in Western blots
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Identification steps:
Designing cross-linking protocols for ATX1-Pol II interaction studies
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Optimized parameters:
Methodological Innovation
Integrating ATX1 ChIP-seq with single-cell RNA sequencing
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Workflow:
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Sort cell types using FACS (e.g., interfascicular fibers vs xylem)
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Parallel processing:
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10% cells for H3K4me3 ChIP-seq
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90% cells for scRNA-seq
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Key finding: 26/107 ATX1-direct targets show stage-specific hypermethylation
