When Immune Cells Receive Opposing Signals – HUN-REN Researchers Uncover a New Adaptive Mechanism

Macrophages are highly adaptable immune cells. Their behaviour is not determined by a single signal: they continuously sense molecules in their surroundings and adjust their functions in response to the combined effects of these signals. A particularly intriguing question is what happens when they encounter signals that are considered to have opposing effects at the same time.

This was the question addressed by the Macrophage Polarization Research Group, led by Zsolt Czimmerer at the Institute of Genetics of the HUN-REN Biological Research Centre, Szeged. Working with national and international partners, the researchers investigated how macrophages respond to the simultaneous presence of two cytokines, IFNγ and IL-4.

‘Our results showed that, under these conditions, the cells do not simply adopt a state determined by one signal or the other, nor do the two effects merely cancel each other out. Instead, a distinct intermediate activation programme emerges, in which the expression of specific groups of genes is markedly enhanced by the combined presence of the two cytokines,’ explained Zsolt Czimmerer. 

One Signal May Prepare the Ground for the Other

Using transcriptomic, epigenomic and CRISPR-based analyses, the researchers investigated the molecular mechanisms underlying the phenomenon. In other words, they examined which genes become activated in the cells, how the accessibility of regulatory regions of DNA changes, and which genes and regulatory proteins are required for the observed response to develop. They found that the response requires the coordinated activity of the STAT1 and STAT6 transcription factors, increased histone acetylation at regulatory regions and the presence of the BRD4 cofactor.

The findings suggest that epigenetic changes initiated by one cytokine – including increased chromatin accessibility – can facilitate the binding of a transcription factor activated by the other, apparently opposing signal to the same regulatory DNA regions. As a result, selected genes can be activated in a coordinated and enhanced manner.

The researchers also identified another important regulatory component: IRF1, a transcription factor activated by IFNγ. IRF1 is indispensable for a substantial proportion of the genes involved in the intermediate macrophage activation programme, and its activity persists even in the presence of IL-4.

Dual Signalling Can Also Occur in Tumours

The researchers also investigated whether the processes identified in the laboratory could be observed in vivo. Analysis of single-cell RNA-sequencing data from a murine breast cancer model identified a macrophage population within the tumour microenvironment in which both STAT1 and STAT6 signalling were active and which displayed characteristic features of the newly identified gene programme.

The findings therefore provide a clearer picture of how macrophages may behave within the complex molecular microenvironments associated with different pathological conditions. The research also highlights that immune-cell responses are not necessarily determined by mutually exclusive signalling pathways. Instead, the final cellular response can be shaped by complex epigenetic and transcriptional regulatory processes that coordinate the accessibility and activity of genes.

In the longer term, a more detailed understanding of these mechanisms may help identify new therapeutic approaches and targets for conditions in which both IL-4 and IFNγ signalling shape the immune microenvironment, including certain severe forms of asthma and various types of cancer.

The research was carried out through a collaboration between research groups at the HUN-REN Biological Research Centre, Szeged and the University of Debrecen, together with researchers from four institutions in the United States: Johns Hopkins All Children’s Hospital, the University of South Florida, Stanford University and the Icahn School of Medicine at Mount Sinai. Zsófia Varga and Anna Poscher played key roles in conducting the research and preparing the publication.

The findings were published in Nucleic Acids Research, an internationally recognised journal in the fields of nucleic acid biology, gene regulation, chromatin research and epigenetics.