Valledor, L. | Search bioRxiv

bioRxiv2025

DNA Methylation Dynamics Reveal Unique Plant Responses and Transcriptional Reprogramming to Combined Heat and Phosphate Deficiency Stress

Plants adapt to environmental challenges through epigenetic mechanisms that modulate gene expression without altering DNA sequence. Among these, DNA methylation is central to balancing genome stability and transcriptional flexibility. We analyzed methylation dynamics in Arabidopsis thaliana under heat, phosphate deficiency, and their combination--conditions that frequently co-occur in nature--using whole-genome bisulfite sequencing, small RNA-seq and RNA-seq of shoots and roots in a setup closely mimicking field conditions. Stress-specific patterns emerged: heat and combined stress led to CHH hypomethylation in both shoot and root, while phosphate deficiency triggered hypermethylation in shoots but hypomethylation in roots. Importantly, the epigenetic response to combined stress was not a mere additive effect of individual stresses but displayed a distinct methylation signature. While both RdDM and CMT2 pathways contributed to heat-induced changes, CMT2 predominated under phosphate deficiency and combined stress, underscoring mechanistic specificity. Methylation changes concentrated in transposable elements (TEs) and intergenic regions, yet TE methylation shifts showed limited correlation with TE or adjacent gene expression, suggesting methylation does not act as a direct transcriptional switch. Instead, stress-induced methylation may influence chromatin accessibility at regulatory regions, particularly transcription factor binding sites; GATA motifs appeared as especially relevant in our analyses. A striking signature emerged in the nuclear mitochondrial DNA (NUMT) region, where hypomethylation under heat and combined stress correlated with upregulation of oxidative phosphorylation genes, critical for thermotolerance. Our findings highlight DNA methylation as an intricate regulatory layer integrating environmental signals into plant adaptive responses, offering a foundation for strategies to harness epigenetic plasticity for crop resilience under climate change.

plant biology↗

bioRxiv2022

Multiomics analyses reveal the central role of nucleolus and nucleoid machinery during heat stress acclimation in Pinus radiata

Climate warming is causing quick changes in mean annual temperature and more severe drought period. These are major contributors of forest dieback, which is becoming more frequent and widespread, particularly in warm and drought-prone regions. Despite being a hot topic in non-woody plant sciences, the information about how heatwaves impact in tree molecular biology is still scarce. In this work we investigated how the transcriptome of Pinus radiata changes during initial stress response and stress acclimation. To this end, and considering this species is non sequenced, we generated a deep dataset employing Illumina technology. This approach allowed us to reconstruct 77335 contigs which were annotated following gene ontology, and to define 12164 and 13590 transcripts as down- and upregulated, respectively, across the three sampled experimental points. Enrichment analysis allowed to distinguish 9 down-regulated pathways, the most of them related to the reduction of apoplast, and water transport. While 22 were upregulated, which followed two different trends those pathways that peaks at short-term (acute response) from those which accumulated long-term (acclimation response) being most of them related to heat shock response, redox machinery and RNA processing. Additionally, the combination of transcriptome data with other available omics layers, allowed an exceptional understanding of the mechanisms behind heat stress response, involving complex interrelated processes from molecular to physiological level. Nucleolus and nucleoid activities seem to be a central core in acclimating process, producing specific RNA isoforms and other essential elements for anterograde-retrograde stress signaling as NAC proteins, Helicase RVB, RZ1 RNA chaperone, or ribosomal RPS4. These mechanisms are connected by elements already known in heat stress-response (redox, heat shock proteins or ABA-related). But also, novel candidates, as photosynthetic pigments, shikimate, or proline centric proteases activities, have been identified effectively networking biochemical responses to its potential regulatory element. This work provides a first deep overview about what molecular mechanisms underlying heat stress response and acclimation in pines, supporting the development of new breeding strategies to face the challenges that the climate change will impose to forests.

plant biology↗