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A candidate ACGTAA-core cis-regulatory element family from an Arabidopsis PET microplastic transcriptome: complete k-mer enrichment and TOMTOM novelty-check workflow
Summary
Scientists studying how plants respond to microplastic pollution found that when Arabidopsis (a common lab plant) roots were exposed to PET plastic particles, a specific, previously-unknown genetic "on switch" turned on certain genes, and this switch doesn't match any known stress-response pattern in plants' DNA. This is early-stage, computer-based detective work (not yet confirmed in the lab), but it suggests plants may have a unique, undiscovered way of reacting to microplastics, which matters because these same plastics are showing up in the food we eat and could be affecting plant health in ways we don't yet understand.
Background Plant cis-regulatory responses to microplastic exposure remain largely uncharacterised. This archive documents the complete, reproducible bioinformatics workflow that led to the identification of a candidate cis-regulatory element family — ACGTAA-core k-mers, culminating in the 8-mer CAACGTAA (BH-FDR q = 7.3e-5, fold enrichment = 3.12 in 1000-bp promoters) — enriched in promoters of Arabidopsis thaliana genes up-regulated by PET microplastic exposure. Data source Public transcriptomic dataset E-MTAB-13532 (Dainelli et al., 2025, Plant Physiology and Biochemistry): Arabidopsis hydroponic root transcriptome, 17-day exposure to transparent PET (Tr-PET) and black PET (Bl-PET) microplastics versus controls, 3 biological replicates per condition. Filtered normalised count matrices were retrieved from the EBI BioStudies database. Workflow (1) limma-trend differential expression analysis: Tr-PET vs control yielded 1,226 DEGs (641 up / 585 down; FDR < 0.05, |log2FC| >= 1), reproducing the pattern reported in the original publication; Bl-PET yielded 31 DEGs. (2) TAIR10 promoter extraction (Ensembl Plants release 59) for 12,128 genes, at multiple window lengths (300-3000 bp upstream of the start codon). (3) Exhaustive k-mer enumeration (k = 5-8, canonical two-strand merging) with 2x2 Fisher exact tests and Benjamini-Hochberg FDR correction, using genes non-responsive to either PET treatment as the negative set. The ACGTAA family is significant at multiple promoter lengths. (4) Known-element scanning: classical stress-responsive elements (as-1/TGACGTAA, G-box/CACGTG, ABRE variants, C-box, A-box) showed no significant enrichment in the PET up-regulated gene set (fold enrichment 0.95-1.23), indicating the PET response does not act through these canonical elements. (5) PWM construction from 340 genomic occurrences (10-bp window, two-strand orientation-normalised, consensus aaACGTAAat) and TOMTOM similarity search against JASPAR CORE 2026 Plants: no significant match (best hit DOF1.7, E = 0.18, q = 0.20; the closest as-1/TGA-family factor TGA1A ranked 18th with E = 4.61). A second candidate (ATTACGTG-family, 23 sites) also showed no significant match (best NAC055, E = 0.107). Pipeline positive control The same workflow applied to an independent ABA/drought-stress transcriptome (Populus) recovered ABRE enrichment as expected (ACGTG fold enrichment 1.29), and TOMTOM annotated one of its de novo motifs as a GAGA element (RAMOSA1 / BPC family, E = 0.013). This demonstrates that the workflow both detects and correctly annotates known elements, strengthening the novelty claim for the PET-responsive candidate. Key claim Within the JASPAR CORE 2026 Plants motif library, the ACGTAA-family candidate has no significant similarity match, while its enrichment in the PET up-regulated set is statistically robust. The claim is bioinformatic (candidate novelty element); functional validation (EMSA / yeast one-hybrid) is planned and not included in this archive. Contents 01_analysis_scripts - All analysis scripts (limma DE in R; promoter extraction, k-mer enumeration, PWM construction, TOMTOM output parsing in Python) 02_de_results - Full differential expression tables (Tr-PET and Bl-PET) 03_kmer_tables - Complete k-mer enrichment tables (4 promoter lengths x up/down sets), summary JSON, promoter extraction statistics 04_tomtom_evidence - TOMTOM input PWM (MEME format) and raw TOMTOM output HTML (JASPAR CORE 2026 Plants) 05_reports - Analysis report and TOMTOM interpretation (Chinese) 06_aba_positive_control - Positive-control materials from the ABA/drought pipeline (TOMTOM input and raw output HTML) Data citation Dainelli et al. (2025) Plant Physiology and Biochemistry. ArrayExpress/BioStudies accession E-MTAB-13532 (https://www.ebi.ac.uk/biostudies/studies/E-MTAB-13532). Reference genome: TAIR10 / Ensembl Plants release 59 (https://plants.ensembl.org/Arabidopsis_thaliana.Info/Index).