HEN1 Antibody
Description
Definition and Biological Context
HEN1 Antibody is a specialized immunological reagent designed to detect and study HUA ENHANCER 1 (HEN1), a conserved RNA methyltransferase critical for small RNA stabilization through 2′-O-methylation. This enzyme is pivotal in RNA silencing pathways across plants, animals, and bacteria, with its inhibition implicated in viral pathogenesis and developmental regulation.
2.1. Epitope Specificity
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Monoclonal Antibody 2A3 targets the C-terminal region of human HEN1/HEN2 (NHLH2), recognizing amino acids 36–135 with a GST tag .
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Polyclonal antibodies against Arabidopsis thaliana HEN1 (AtHEN1) are generated using recombinant fragments for detecting endogenous HEN1 in plant studies .
2.2. Molecular Applications
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Immunocytochemistry: Localizes HEN1 in cytoplasmic and nuclear compartments .
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Co-Immunoprecipitation (Co-IP): Validates interactions between HEN1 and viral suppressors like HC-Pro .
3.1. Role in Viral RNA Silencing Suppression
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HC-Pro Inhibition: HEN1 antibodies enabled the discovery that potyviral HC-Pro variants (e.g., HC-Pro Tu) directly bind and inhibit HEN1’s methyltransferase activity, leading to unmodified miRNAs and enhanced AGO1 degradation .
3.2. Stability of Small RNAs
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In C. elegans, HENN-1 (HEN1 ortholog) methylates piRNAs and 26G RNAs, with antibody-based studies revealing:
5.1. Plant-Virus Interactions
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AtHEN1 Antibody confirmed HC-Pro’s role in sequestering HEN1 into cytoplasmic “H-bodies,” impairing miRNA methylation and promoting viral proliferation .
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EMSA assays using purified GST-HC-Pro and his-HEN1 demonstrated dose-dependent inhibition of HEN1’s RNA-binding capacity .
5.2. Animal Models
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C. elegans HENN-1 Antibodies revealed tissue-specific methylation patterns:
Challenges and Future Directions
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
Related Research
- Research indicates that HESO1 may have a redundant function with URT1 in miRNA uridylation when HEN1 is fully functional. PMID: 25928341
- Light-activated expression of HEN1 is dependent on the photoreceptors phytochrome A (phyA), phyB, cryptochrome 1 (cry1), and cry2, as well as key transcriptional regulators ELONGATED HYPOCOTYL5 (HY5) and HY5-HOMOLOG. PMID: 25052717
- Studies suggest a mechanism for plant miRNA maturation involving the binding of HEN1 methyltransferase to the DICER-LIKE 1 ribonuclease DCL1*RNA-binding protein HYL1*miRNA complex. PMID: 25680966
- Data suggests that the ago1 mutation in hen1 suppressed miRNA 3' modifications. PMID: 23839787
- AGO1, HYL1, and HEN1 have been found to play a role in the proximal-distal and vascular patterning of leaves. PMID: 22623415
- Motif mapping analysis revealed that amino acids between positions 139 and 320 of Zucchini yellow mosaic virus HC-Pro were associated with interaction with Arabidopsis HEN1. PMID: 21593273
- A mutation in RNA-dependent RNA polymerase 2 rescued the defects in miRNA methylation of hen1-2, revealing an unexpected, negative influence of siRNAs on HEN1-mediated miRNA methylation. PMID: 20448024
- Kinetic parameters with natural RNA substrates show that HEN1 exhibits high catalytic efficiency in the absence of supplementary proteins. The enzyme modifies individual strands in succession, leading to complete methylation of an RNA duplex. PMID: 20705645
- HEN1 is a miRNA methyltransferase that adds a methyl group to the 3'-most nucleotide of miRNAs, as well as siRNAs from sense transgenes, hairpin transgenes, and transposons or repeat sequences. It also targets a new class of siRNAs known as trans-acting siRNAs. PMID: 16111943
- A crystal structure of full-length HEN1 from Arabidopsis in complex with a 22-nucleotide small RNA duplex and cofactor product S-adenosyl-l-homocysteine has been determined. PMID: 19812675
Q&A
Here’s a structured FAQ for researchers working with HEN1 antibodies, designed to address academic research scenarios with methodological rigor and depth:
Advanced Research Questions
How to resolve contradictions in HEN1 localization data between immunocytochemistry and biochemical fractionation?
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Methodological reconciliation:
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Validate antibody cross-reactivity with organelle-specific markers (e.g., nucleolin for nucleus, COX2 for mitochondria) .
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Compare fixation methods (e.g., paraformaldehyde vs. methanol) to rule out epitope masking .
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Perform subcellular fractionation with protease/RNase inhibitors, followed by immunoblotting .
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What statistical approaches are robust for analyzing HEN1 knockout phenotypic variability?
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Apply mixed-effects models to account for batch effects in multi-experiment datasets .
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Use Bonferroni correction for small RNA sequencing data to address false discovery rates in differentially expressed miRNAs .
How to design a study linking HEN1 dysfunction to epigenetic instability in chronic stress models?
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Key parameters:
