PMID- 29708974 OWN - NLM STAT- MEDLINE DCOM- 20180618 LR - 20240214 IS - 1553-7358 (Electronic) IS - 1553-734X (Print) IS - 1553-734X (Linking) VI - 14 IP - 4 DP - 2018 Apr TI - Quantitative analysis reveals crosstalk mechanisms of heat shock-induced attenuation of NF-kappaB signaling at the single cell level. PG - e1006130 LID - 10.1371/journal.pcbi.1006130 [doi] LID - e1006130 AB - Elevated temperature induces the heat shock (HS) response, which modulates cell proliferation, apoptosis, the immune and inflammatory responses. However, specific mechanisms linking the HS response pathways to major cellular signaling systems are not fully understood. Here we used integrated computational and experimental approaches to quantitatively analyze the crosstalk mechanisms between the HS-response and a master regulator of inflammation, cell proliferation, and apoptosis the Nuclear Factor kappaB (NF-kappaB) system. We found that populations of human osteosarcoma cells, exposed to a clinically relevant 43 degrees C HS had an attenuated NF-kappaB p65 response to Tumor Necrosis Factor alpha (TNFalpha) treatment. The degree of inhibition of the NF-kappaB response depended on the HS exposure time. Mathematical modeling of single cells indicated that individual crosstalk mechanisms differentially encode HS-mediated NF-kappaB responses while being consistent with the observed population-level responses. In particular "all-or-nothing" encoding mechanisms were involved in the HS-dependent regulation of the IKK activity and IkappaBalpha phosphorylation, while others involving transport were "analogue". In order to discriminate between these mechanisms, we used live-cell imaging of nuclear translocations of the NF-kappaB p65 subunit. The single cell responses exhibited "all-or-nothing" encoding. While most cells did not respond to TNFalpha stimulation after a 60 min HS, 27% showed responses similar to those not receiving HS. We further demonstrated experimentally and theoretically that the predicted inhibition of IKK activity was consistent with the observed HS-dependent depletion of the IKKalpha and IKKbeta subunits in whole cell lysates. However, a combination of "all-or-nothing" crosstalk mechanisms was required to completely recapitulate the single cell data. We postulate therefore that the heterogeneity of the single cell responses might be explained by the cell-intrinsic variability of HS-modulated IKK signaling. In summary, we show that high temperature modulates NF-kappaB responses in single cells in a complex and unintuitive manner, which needs to be considered in hyperthermia-based treatment strategies. FAU - Kardynska, Malgorzata AU - Kardynska M AD - Systems Engineering Group, Silesian University of Technology, Gliwice, Poland. FAU - Paszek, Anna AU - Paszek A AD - Systems Engineering Group, Silesian University of Technology, Gliwice, Poland. AD - System Microscopy Centre, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, United Kingdom. AD - Maria Sklodowska-Curie Institute-Oncology Center, Gliwice Branch, Gliwice, Poland. FAU - Smieja, Jaroslaw AU - Smieja J AUID- ORCID: 0000-0002-6120-4424 AD - Systems Engineering Group, Silesian University of Technology, Gliwice, Poland. FAU - Spiller, David AU - Spiller D AD - System Microscopy Centre, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, United Kingdom. FAU - Widlak, Wieslawa AU - Widlak W AD - Maria Sklodowska-Curie Institute-Oncology Center, Gliwice Branch, Gliwice, Poland. FAU - White, Michael R H AU - White MRH AD - System Microscopy Centre, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, United Kingdom. FAU - Paszek, Pawel AU - Paszek P AUID- ORCID: 0000-0002-0363-0716 AD - System Microscopy Centre, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester Academic Health Science Centre, Manchester, United Kingdom. FAU - Kimmel, Marek AU - Kimmel M AD - Systems Engineering Group, Silesian University of Technology, Gliwice, Poland. AD - Departments of Statistics and Bioengineering, Rice University, Houston, TX, United States of America. LA - eng GR - BB/I017976/1/BB_/Biotechnology and Biological Sciences Research Council/United Kingdom GR - BB/K003097/1/BB_/Biotechnology and Biological Sciences Research Council/United Kingdom PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20180430 PL - United States TA - PLoS Comput Biol JT - PLoS computational biology JID - 101238922 RN - 0 (NF-kappa B) RN - 0 (RELA protein, human) RN - 0 (Recombinant Fusion Proteins) RN - 0 (Transcription Factor RelA) RN - 0 (Tumor Necrosis Factor-alpha) RN - 0 (enhanced green fluorescent protein) RN - 147336-22-9 (Green Fluorescent Proteins) RN - EC 2.7.11.10 (I-kappa B Kinase) SB - IM MH - Cell Line MH - Computational Biology MH - Computer Simulation MH - Green Fluorescent Proteins/genetics/metabolism MH - Heat-Shock Response/*physiology MH - Humans MH - I-kappa B Kinase/metabolism MH - *Models, Biological MH - NF-kappa B/*metabolism MH - Recombinant Fusion Proteins/genetics/metabolism MH - Signal Transduction MH - Single-Cell Analysis MH - Transcription Factor RelA/genetics/metabolism MH - Tumor Necrosis Factor-alpha/metabolism PMC - PMC5945226 COIS- The authors have declared that no competing interests exist. EDAT- 2018/05/01 06:00 MHDA- 2018/06/19 06:00 PMCR- 2018/04/30 CRDT- 2018/05/01 06:00 PHST- 2018/03/09 00:00 [received] PHST- 2018/04/10 00:00 [accepted] PHST- 2018/05/10 00:00 [revised] PHST- 2018/05/01 06:00 [pubmed] PHST- 2018/06/19 06:00 [medline] PHST- 2018/05/01 06:00 [entrez] PHST- 2018/04/30 00:00 [pmc-release] AID - PCOMPBIOL-D-18-00385 [pii] AID - 10.1371/journal.pcbi.1006130 [doi] PST - epublish SO - PLoS Comput Biol. 2018 Apr 30;14(4):e1006130. doi: 10.1371/journal.pcbi.1006130. eCollection 2018 Apr.