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© The Author(s), 2021. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Weizman, E., Rinsky, M., Simon-Blecher, N., Lampert-Karako, S., Yaron, O., Tarrant, A. M., & Levy, O. Chromatin dynamics and gene expression response to heat exposure in field-conditioned versus laboratory-cultured Nematostella vectensis. International Journal of Molecular Sciences, 22(14), (2021): 7454, https://doi.org/10.3390/ijms22147454., Organisms’ survival is associated with the ability to respond to natural or anthropogenic environmental stressors. Frequently, these responses involve changes in gene regulation and expression, consequently altering physiology, development, or behavior. Here, we present modifications in response to heat exposure that mimics extreme summertime field conditions of lab-cultured and field-conditioned Nematostella vectensis. Using ATAC-seq and RNA-seq data, we found that field-conditioned animals had a more concentrated reaction to short-term thermal stress, expressed as enrichment of the DNA repair mechanism pathway. By contrast, lab animals had a more diffuse reaction that involved a larger number of differentially expressed genes and enriched pathways, including amino acid metabolism. Our results demonstrate that pre-conditioning affects the ability to respond efficiently to heat exposure in terms of both chromatin accessibility and gene expression and reinforces the importance of experimentally addressing ecological questions in the field.