## 1 Introduction  
The levels of carbon dioxide (CO\({}_{2}\)) in the blood are closely regulated in the body through acute chemosensory mechanisms that respond to changes in the levels of CO\({}_{2}\) and adjust the rate and depth of breathing. In certain lung diseases, e.g., chronic obstructive pulmonary disease (COPD), the partial pressure of CO\({}_{2}\) in the blood can accumulate to levels >45 mmHg, termed hypercapnia. Hypercapnia is known to suppress immune and inflammatory signalling and is a risk factor in several diseases, e.g., COPD [1]. Hypercapnia can elicit signalling acutely through modulation of chemosensory and neuronal signaling and more chronically via changes in gene expression [2]. The mechanisms through which CO\({}_{2}\) affects transcriptional responses are not fully understood [3] and may involve different transcription factors in different contexts. Work from our group and others indicates that multiple signalling pathways are modulated by increased CO\({}_{2}\). However, relatively few transcription factors have been tested in interventional (e.g., knockdown/knockout) studies, which is the approach we have taken here to provide mechanistic insight. The Orphan Nuclear Receptor Family 4A family members (NR4A) (_NR4A1-3_) are early response genes that are activated in response to a variety of stimuli and stressors [4; 5] and modulate the differentiation of monocytes: classical to non-classical monocytes (_NR4A1_) and to dendritic cells (_NR4A3_). NR4A expression is associated with several pathologies, including arthritis [6; 7], atherosclerosis [8], and cancer [9; 10]. Regulation of NR4A activity is primarily at the transcriptional level, with a correlation between mRNA levels and activity. In recent years, several NR4A modulators have been developed and have demonstrated efficacy in numerous models, supporting the idea that these receptors are feasible therapeutic targets [11; 12]. The NR4 As can also undergo post-translational modifications, specifically phosphorylation and SUMOylation, which can also affect their activity [11; 13; 14]. We have recently characterised the transcriptional roles of _NR4A2_ and _NR4A3_ in THP-1 monocytes, revealing significant overlap of transcriptional targets as well as more selective antigen presentation (_NR4A2_-dependent) and viral response (_NR4A3_-dependent) roles [15]. The NR4A family of transcription factors and their role in the response to elevated CO2 are the focus of this study for a number of reasons. Firstly, nuclear hormone receptors, of which _NR4A_s are members, represent a class of genes that was initially identified as being CO2 sensitive in _Caenorhabditis elegans_ (C. _elegans_) [16]. In this seminal model organism study, E3 ligases, which regulate protein degradation via the ubiquitin pathway, and genes associated with the innate immune response were key classes of genes implicated in the C. elegans response to CO2 elevation. Subsequent to this study in worms, several studies have corroborated the role of the ubiquitin pathway [17; 18] and immune regulation [19; 20] in the CO2 response in mammalian systems. Indeed, a recent study identified ubiquitin as a carbon-dioxide-binding protein [21]. Thus, we hypothesised that nuclear receptor family members (which were robustly CO2 sensitive in _C. elegans_) are worth pursuing further as potential conserved CO2-sensitive transcriptional regulators. Furthermore, we and others have demonstrated a role for altered /suppressed Nuclear Factor of Kappa Light Chain Gene Enhancer in B cells (NFkB)-dependent signalling in the cellular response to hypercapnia [21; 22; 23]. The NR4 As are known to be both regulated by NFkB-dependent signalling and capable of regulating NFkB-dependent transcription [24; 25]. Thus, the NR4A family members have the potential to act at the intersection of NFkB-dependent and CO2-dependent signalling.