Abstract
The human skin and in particular its outermost layer, the epidermis, protects the body from potentially harmful substances, radiation as well as excessive water loss. However, the interference between the various stress responses of the epidermal keratinocytes, which often occur simultaneously, is largely unknown. The focus of this study was to investigate the interference between osmotic stress and DNA damage response. In addition to revealing the already well-described regulation of diverse gene sets, for example, cellular processes such as transcription, translation, and metabolic pathways (e.g., the KEGG citrate cycle and Reactome G2/M checkpoints), gene expression analysis of osmotically stressed keratinocytes revealed an influence on the transcription of genes also related to UV-induced DNA damage response. A gene network regulating the H2AX phosphorylation was identified to be regulated by osmotic stress. To analyze and test the interference between osmotic stress and DNA damage response, which can be triggered by UV stress on the one hand and oxidative stress on the other, in more detail, primary human keratinocytes were cultured under osmotic stress conditions and subsequently exposed to UV light and H2O2, respectively. γH2AX measurements revealed lower γH2AX levels in cells previously cultured under osmotic stress conditions.
Highlights
The human skin consists of a multilayered epithelium, which is the barrier between the environment and the organism [1]
We investigate the interference between osmotic stress and DNA damage response in primary human skin keratinocytes
The interference was validated by γH2AX measurements in simultaneous stress treatment, and a functional gene network controlling phosphorylation of H2AX was identified to be affected by osmotic stress, explaining the interference between these stress responses in human keratinocytes
Summary
The human skin consists of a multilayered epithelium, which is the barrier between the environment and the organism [1]. The normal sodium concentration in blood serum is between 135–145 mmol/L [9] This can increase due to a loss of water [10] and can lead to an osmotic stress response. ATM senses double-stranded breaks, whereas ATR predominantly senses single-stranded breaks These checkpoint kinases transfer signals to effector molecules and phosphorylate histone family member X (H2AX), which is the key event in DNA damage response [20,22]. Increased DNA double-strand breaks were reported after osmotic stress in murine kidney cells [27], which persisted even when cells adapted to the osmotic stress [28] This shows that cellular stress pathways form complex networks and that the interference between different stress responses is of major importance, as it could help to develop new therapeutic targets [29]. The interference was validated by γH2AX measurements in simultaneous stress treatment, and a functional gene network controlling phosphorylation of H2AX was identified to be affected by osmotic stress, explaining the interference between these stress responses in human keratinocytes
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