mEAT

mEAT maps transcriptome-wide binding sites of RNA-binding proteins (RBPs) and microRNA-containing ribonucleoprotein complexes (miRNPs) by detecting thymidine-to-cytidine transitions in cDNAs from 4-thiouridine–labeled, immunopurified RNPs to enable analysis of posttranscriptional gene regulation.

Key Features:

• High-resolution transcriptome-wide mapping: Identifies binding sites of RBPs and miRNPs across the transcriptome at nucleotide resolution.
• Crosslink signature detection: Detects thymidine-to-cytidine (T-to-C) transitions in cDNAs as evidence of crosslinked sites.
• 4-thiouridine labeling and immunopurification: Employs 4-thiouridine incorporation and immunopurification of RNA-protein complexes (RNPs) to enrich crosslinked material.
• RBP and miRNP profiling: Determines binding sites for specific factors including PUM2, QKI, IGF2BP1-3, AGO/EIF2C1-4, and TNRC6A-C.
• Sequence motif identification: Reveals distinct sequence motifs associated with RBP and miRNP binding sites.
• Transcript region preference analysis: Assesses binding preferences for exonic versus intronic regions and coding sequences versus untranslated regions.
• Genetic variation interpretation: Provides mapping information useful for assessing how genetic variation influences complex genetic diseases.

Scientific Applications:

• Posttranscriptional gene regulation studies: Interprets the regulatory roles of RBPs and miRNPs in controlling RNA fate and function.
• Cell-type specific binding analysis: Elucidates binding patterns and regulatory consequences of RBPs and miRNPs across different cell types.
• Genetic disease research: Links precise binding-site maps to the impact of genetic variation on complex genetic diseases.

Methodology:

Cells are labeled with 4-thiouridine, RNA-protein complexes are immunopurified, and crosslinked sites are identified via thymidine-to-cytidine transitions in cDNAs derived from the purified RNPs.