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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 found that when a common plant (Arabidopsis, related to crops like broccoli and mustard) is exposed to PET plastic particles in water, it turns on genes using a previously unknown genetic "switch", not the usual stress-response switches plants use for things like drought. This is early, computer-based evidence (not yet lab-confirmed) that plants may respond to microplastics through a distinct biological pathway, which matters because it could help researchers understand how microplastic contamination affects the food crops we eat and the water systems we depend on.
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).