The systemic immune response to viral infection is shaped by master transcription factors, such as NF-κB, STAT1, or PU.1. Although long noncoding RNAs (lncRNAs) have been suggested as important regulators of transcription factor activity, their contributions to the systemic immunopathologies observed during SARS-CoV-2 infection have remained unknown. Researchers at Philipps University Marburg have now employed a targeted single-cell RNA sequencing approach to reveal lncRNAs differentially expressed in blood leukocytes during severe COVID-19. These results uncover the lncRNA PIRAT (PU.1-induced regulator of alarmin transcription) as a major PU.1 feedback-regulator in monocytes, governing the production of the alarmins S100A8/A9, key drivers of COVID-19 pathogenesis. Knockout and transgene expression, combined with chromatin-occupancy profiling, characterized PIRAT as a nuclear decoy RNA, keeping PU.1 from binding to alarmin promoters and promoting its binding to pseudogenes in naïve monocytes. NF-κB–dependent PIRAT down-regulation during COVID-19 consequently releases a transcriptional brake, fueling alarmin production. Alarmin expression is additionally enhanced by the up-regulation of the lncRNA LUCAT1, which promotes NF-κB–dependent gene expression at the expense of targets of the JAK-STAT pathway. These results suggest a major role of nuclear noncoding RNA networks in systemic antiviral responses to SARS-CoV-2 in humans.
Identification of human myeloid lineage-specific lincRNAs
(A) Bulk and scRNA-seq strategy for the determination of myeloid lincRNAs relevant to COVID-19. (B) PCA (Upper) and hierarchical clustering (Lower, z-scores) of monocyte, granulocyte, B, NK, and CD4+ and CD8+ T cell lincRNA profiles. (C) qRT-PCR validation of PIRAT (LINC00211) and LUCAT1 as myeloid-specific lincRNAs (expression relative to human brain tissue). Horizontal bar indicates base-line (black) and twofold deviation from base-line (gray). (D) Relative abundance of PIRAT and LUCAT1 in RNA-seq profiles of hematopoietic stem cells (HSC), hematopoietic multipotent precursor cells (HMPC), common myeloid progenitors (CMP), granulocyte-macrophage progenitors (GMP), monocytes, and neutrophils. (E, Upper) PIRAT coexpression network, derived from RNA-seq data in B; (Lower) qRT-PCR analysis of PU.1 mRNA, PIRAT and LUCAT1 expression in PU.1 knockdown compared to control THP1 monocytes. (F and G) qRT-PCR analysis of lincRNA expression in response to indicated PAMPs and NF-κB inhibitor BAY-11-7082 (PAMP = 4 h LPS + polyI:C stimulation). (C–G) ≥3 independent experiments and one-way ANOVA test.