Sugitani, N. | bioRxiv Science

bioRxiv2026

Follistatin-like 1 Promotes Neutrophil-mediated Host Defense against Klebsiella pneumoniae

Klebsiella pneumoniae is a leading cause of pneumonia and highly burdensome etiology of antibiotic-resistant infections constituting an urgent public health threat. Follistatin-like 1 (FSTL-1) is implicated in many processes including lung development and homeostasis, tissue remodeling, and inflammation though its role in antibacterial immunity remains poorly defined. We have observed that FSTL-1 is elevated in the lungs of both humans and mice during bacterial pneumonia. Using an inducible Fstl1 knockout mouse model, we demonstrate that FSTL-1 is required for pulmonary bacterial clearance and neutrophil recruitment during K. pneumoniae infection. Recombinant FSTL-1 rescues Fstl1-deficient susceptibility and enhances clearance of K. pneumoniae in wild-type mice. Mechanistically, FSTL-1-mediated protection was lost upon neutrophil depletion and in Cd14-deficient or Nr4a1 deficient mice, components of a lung-specific neutrophil transcriptional program. These findings identify FSTL-1 as a lung-derived factor that licenses neutrophil-mediated antibacterial defense via a novel FSTL-1/CD14/NR4A1-dependent signaling pathway critical for protection against bacterial pneumonia.

immunology↗

bioRxiv2026

A Novel Role for FSTL-1 in Hematopoietic Stem Cell Homeostasis and Post-injury Regeneration

Background: The ability to repopulate all lineages of blood cells from a single hematopoietic stem cell (HSC) requires a constant balancing between maintenance of quiescence/self-renewal and the need for differentiation and adoption of specific lineage phenotypes. A classical interpretation suggests that during homeostasis, long- and then short-term HSCs differentiate into multipotent progenitors (MPPs), which then lack self-renewal capacity but can still differentiate into all downstream lineages. While much is known about the intrinsic and extrinsic regulators that promote HSC homeostasis, less is known about factors which influence HSC behavior following significant bone marrow (BM) injury, such as sublethal irradiation (SLI). Methods: Conditional knock-out of follistatin-like 1 (FSTL-1) was achieved by breeding mice with floxed Fstl1 sites to mice expressing CreERT2 under control of the ROSA locus, followed by systemic tamoxifen administration. ELISA and RT-PCR quantitated protein and RNA level loss of FSTL-1, respectively. HSC populations were assessed by flow cytometry under multiple conditions. Sublethal irradiation induced BM injury and blocking polyclonal FSTL-1 antibodies were used to recapitulate results seen with genetic ablation. Finally, single cell RNA sequencing defined both cell-specific expression of Fstl1 within the BM compartment, but also identified transcripts and pathways upregulated in the absence of FSTL-1. Results: These studies delineate a novel role for FSTL-1 in regulating HSC biology. In addition to selective expression of Fstl1 within HSCs, genetic deletion of Fstl1 increases Lineage negative;Sca-1+c-Kit+ (LSK) cell number and alters HSC homeostasis. These findings are further amplified in the BM in the context of recovery from sublethal irradiation. Importantly, the post-irradiation increase in BM cellularity seen with Fstl1 deletion is replicated by treatment with anti-FSTL-1 polyclonal antibodies. Finally, single cell RNA sequencing reveals distinct Fstl1 expression in specific LSK subpopulations, including HSCs and proliferating cells, and identifies a novel relationship between decreased FSTL-1 levels and enrichment of canonical metabolic pathways including oxidative phosphorylation, fatty acid metabolism, and glycolysis. Conclusions: Taken together, these results highlight a novel role for FSTL-1 in promoting HSC homeostasis. They also unlock an innovative way to promote HSPC recovery following BM injury, knowledge predicted to be of significant therapeutic value.

developmental biology↗

bioRxiv2022

ATR kinase inhibition induces thymineless death in proliferating CD8+ T cells

ATR kinase is a central regulator of the DNA damage response (DDR) and cell cycle checkpoints. ATR kinase inhibitors (ATRis) combine with radiation to generate CD8+ T cell-dependent responses in mouse models of cancer. We show that ATRis induce CDK1-dependent origin firing across active replicons in CD8+ T cells activated ex vivo while simultaneously decreasing the activity of rate-limiting enzymes for nucleotide biosynthesis. These pleiotropic effects of ATRi induce deoxyuridine (dU) contamination in genomic DNA, R loops, RNA-DNA polymerase collisions, and type-1 interferons (IFN-1). Remarkably, thymidine rescues ATRi-induced dU contamination, cell death, and IFN-1 expression in proliferating CD8+ T cells. Thymidine also rescues ATRi-induced cancer cell death. We propose that ATRi-induced dU contamination contributes to dose-limiting leukocytopenia and inflammation in the clinic and CD8+ T cell dependent anti-tumor responses in mouse models. We conclude that ATR is essential to limit dU contamination in genomic DNA and IFN-1 expression.

cancer biology↗