Dear Dr. Newton,

Thank you for submitting your manuscript to PLOS ONE. After careful consideration, we feel that it has merit but does not fully meet PLOS ONE’s publication criteria as it currently stands. Therefore, we invite you to submit a revised version of the manuscript that addresses the points raised during the review process.

In your revised manuscript, please extensively work on the abstract. I recommend revising the abstract to improve its logical flow and thematic coherence. The current version is difficult to follow and unlikely to engage a broad scientific audience. As Reviewer #2 suggested, some of the data presentation needs to be reformatted and with suitable statistic analysis. 

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Helene Minyi Liu, Ph.D.

Academic Editor

PLOS ONE

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Please state what role the funders took in the study.  If the funders had no role, please state: "The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript."

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Reviewers' comments:

Reviewer's Responses to Questions

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #2: Yes

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2. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #1: Yes

Reviewer #2: Yes

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3. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy

Reviewer #1: Yes

Reviewer #2: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1: This manuscript examines the regulation of IRFs in human pulmonary epithelial cells, with a particular emphasis on the NFκB–dependent induction of IRF1 by IL1β and TNFα. The authors use a combination of cell lines (A549 and BEAS-2B) and primary human bronchial epithelial cells, applying multiple complementary approaches, including mRNA-seq, ChIP-seq, siRNA silencing, pharmacologic inhibition, and reporter assays, to dissect the underlying mechanisms.

The data convincingly demonstrate that IRF1 is robustly induced in an NFκB-dependent manner, with several upstream enhancer regions contributing to its transcriptional control. Notably, IRF1 induction appears largely resistant to glucocorticoid suppression, a finding with potential implications for glucocorticoid resistant airway inflammation and severe asthma.

Overall, this is a technically rigorous, well controlled, and carefully executed study that makes an original contribution to the field. The conclusions are well supported by the data, the manuscript is clearly written, and all required ethics approvals and data availability statements are in place.

I have only one minor suggestion:

The abstract is difficult to follow in its current form. It should be revised for clarity and improved flow so that the key findings are more easily understood.

Reviewer #2: During inflammation, cytokines such as IL-1β and TNF⍺ activate signaling cascades through their respective receptors, leading to the activation of transcription factors like NF-κB and interferon regulatory factors (IRFs). NF-κB is activated through the phosphorylation and degradation of IκBα, allowing NF-κB to translocate to the nucleus and drive inflammatory gene expression. Although IRFs are primarily known for their role in antiviral and interferon responses, emerging evidence suggests they may also be activated through NF-κB-associated pathways, though this remains underexplored. Airway epithelial cells (AECs), as the frontline defenders against inhaled pathogens and irritants, play a central role in lung inflammation and are key targets of inhaled glucocorticoids used to manage asthma. However, the regulation of IRFs—particularly IRF1—in AECs is not well understood. IRF1 is inducible by cytokines and may regulate inflammatory mediators such as CXCL10, which has been linked to severe asthma. This study investigates how IL-1β and TNF⍺ influence IRF1 expression and inflammatory signaling in AECs, and how these responses are modulated by glucocorticoids like dexamethasone and budesonide. There are some comments as the follows:

1. In some graphs, such as Fig 1A, Fig 10A, Supplementary Fig 1C, and Supplementary Fig 6A, the time intervals in the x-axis are different, so they are not suitable for line chart.

2. In Fig 3B and 3C, Fig 4, Fig 6A and Fig 9, the statistical results of some charts need to be fully labeled and presented. For example, in Fig 3, some are labeled with “ns”, but it’s confused whether non-labeled are all “ns”.

3. In Fig 3 and Fig 9, should TNF⍺ treatment groups be labeled as yellow or orange?

4. In line 99, “This revealed transcripts for all nine IRFs under basal conditions…." should be “These revealed transcripts……".

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Reviewer #1: No

Reviewer #2: No

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While revising your submission, please upload your figure files to the Preflight Analysis and Conversion Engine (PACE) digital diagnostic tool, https://pacev2.apexcovantage.com/ . PACE helps ensure that figures meet PLOS requirements. To use PACE, you must first register as a user. Registration is free. Then, login and navigate to the UPLOAD tab, where you will find detailed instructions on how to use the tool. If you encounter any issues or have any questions when using PACE, please email PLOS at figures@plos.org

Inflammatory cytokines promote interferon regulatory factor (IRF) transcriptional activity in human pulmonary epithelial cells through the induction of IRF1 by nuclear factor-κB

PONE-D-25-37763R1

Dear Dr. Newton,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

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Kind regards,

Helene Minyi Liu, Ph.D.

Academic Editor

PLOS ONE

Additional Editor Comments (optional):

Reviewers' comments: