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IKKα plays a major role in canonical NF-κB signalling in colorectal cells

Inhibitor of kappa B (IκB) kinase β (IKKβ) has long been viewed as the dominant IKK in the canonical nuclear factor-κB (NF-κB) signalling pathway, with IKKα being more important in non-canonical NF-κB activation. Here we have investigated the role of IKKα and IKKβ in canonical NF-κB activation in co...

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Autores principales: Prescott, Jack A., Balmanno, Kathryn, Mitchell, Jennifer P., Okkenhaug, Hanneke, Cook, Simon J.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Portland Press Ltd. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8883499/
https://www.ncbi.nlm.nih.gov/pubmed/35029639
http://dx.doi.org/10.1042/BCJ20210783
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author Prescott, Jack A.
Balmanno, Kathryn
Mitchell, Jennifer P.
Okkenhaug, Hanneke
Cook, Simon J.
author_facet Prescott, Jack A.
Balmanno, Kathryn
Mitchell, Jennifer P.
Okkenhaug, Hanneke
Cook, Simon J.
author_sort Prescott, Jack A.
collection PubMed
description Inhibitor of kappa B (IκB) kinase β (IKKβ) has long been viewed as the dominant IKK in the canonical nuclear factor-κB (NF-κB) signalling pathway, with IKKα being more important in non-canonical NF-κB activation. Here we have investigated the role of IKKα and IKKβ in canonical NF-κB activation in colorectal cells using CRISPR–Cas9 knock-out cell lines, siRNA and selective IKKβ inhibitors. IKKα and IKKβ were redundant for IκBα phosphorylation and turnover since loss of IKKα or IKKβ alone had little (SW620 cells) or no (HCT116 cells) effect. However, in HCT116 cells IKKα was the dominant IKK required for basal phosphorylation of p65 at S536, stimulated phosphorylation of p65 at S468, nuclear translocation of p65 and the NF-κB-dependent transcriptional response to both TNFα and IL-1α. In these cells, IKKβ was far less efficient at compensating for the loss of IKKα than IKKα was able to compensate for the loss of IKKβ. This was confirmed when siRNA was used to knock-down the non-targeted kinase in single KO cells. Critically, the selective IKKβ inhibitor BIX02514 confirmed these observations in WT cells and similar results were seen in SW620 cells. Notably, whilst IKKα loss strongly inhibited TNFα-dependent p65 nuclear translocation, IKKα and IKKβ contributed equally to c-Rel nuclear translocation indicating that different NF-κB subunits exhibit different dependencies on these IKKs. These results demonstrate a major role for IKKα in canonical NF-κB signalling in colorectal cells and may be relevant to efforts to design IKK inhibitors, which have focused largely on IKKβ to date.
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spelling pubmed-88834992022-03-10 IKKα plays a major role in canonical NF-κB signalling in colorectal cells Prescott, Jack A. Balmanno, Kathryn Mitchell, Jennifer P. Okkenhaug, Hanneke Cook, Simon J. Biochem J Cancer Inhibitor of kappa B (IκB) kinase β (IKKβ) has long been viewed as the dominant IKK in the canonical nuclear factor-κB (NF-κB) signalling pathway, with IKKα being more important in non-canonical NF-κB activation. Here we have investigated the role of IKKα and IKKβ in canonical NF-κB activation in colorectal cells using CRISPR–Cas9 knock-out cell lines, siRNA and selective IKKβ inhibitors. IKKα and IKKβ were redundant for IκBα phosphorylation and turnover since loss of IKKα or IKKβ alone had little (SW620 cells) or no (HCT116 cells) effect. However, in HCT116 cells IKKα was the dominant IKK required for basal phosphorylation of p65 at S536, stimulated phosphorylation of p65 at S468, nuclear translocation of p65 and the NF-κB-dependent transcriptional response to both TNFα and IL-1α. In these cells, IKKβ was far less efficient at compensating for the loss of IKKα than IKKα was able to compensate for the loss of IKKβ. This was confirmed when siRNA was used to knock-down the non-targeted kinase in single KO cells. Critically, the selective IKKβ inhibitor BIX02514 confirmed these observations in WT cells and similar results were seen in SW620 cells. Notably, whilst IKKα loss strongly inhibited TNFα-dependent p65 nuclear translocation, IKKα and IKKβ contributed equally to c-Rel nuclear translocation indicating that different NF-κB subunits exhibit different dependencies on these IKKs. These results demonstrate a major role for IKKα in canonical NF-κB signalling in colorectal cells and may be relevant to efforts to design IKK inhibitors, which have focused largely on IKKβ to date. Portland Press Ltd. 2022-02-11 2022-02-04 /pmc/articles/PMC8883499/ /pubmed/35029639 http://dx.doi.org/10.1042/BCJ20210783 Text en © 2022 The Author(s) https://creativecommons.org/licenses/by/4.0/This is an open access article published by Portland Press Limited on behalf of the Biochemical Society and distributed under the Creative Commons Attribution License 4.0 (CC BY) (https://creativecommons.org/licenses/by/4.0/) .
spellingShingle Cancer
Prescott, Jack A.
Balmanno, Kathryn
Mitchell, Jennifer P.
Okkenhaug, Hanneke
Cook, Simon J.
IKKα plays a major role in canonical NF-κB signalling in colorectal cells
title IKKα plays a major role in canonical NF-κB signalling in colorectal cells
title_full IKKα plays a major role in canonical NF-κB signalling in colorectal cells
title_fullStr IKKα plays a major role in canonical NF-κB signalling in colorectal cells
title_full_unstemmed IKKα plays a major role in canonical NF-κB signalling in colorectal cells
title_short IKKα plays a major role in canonical NF-κB signalling in colorectal cells
title_sort ikkα plays a major role in canonical nf-κb signalling in colorectal cells
topic Cancer
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8883499/
https://www.ncbi.nlm.nih.gov/pubmed/35029639
http://dx.doi.org/10.1042/BCJ20210783
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