PF456 BETA‐CATENIN/TCF SIGNALING PATHWAY PROMOTES GRANULOCYTIC DIFFERENTIATION OF HEMATOPOIETIC PROGENITORS BY FACILITATING IL‐1 RECEPTOR SIGNALING
Authors: Daněk P. , Kardosova M. , Zjablovskaja P. , Janeckova L. , Korinek V. , Alberich‐Jorda M. 
Published: December 2023

Background:

The canonical Wnt signaling pathway is an essential regulator of differentiation and cell fate decisions in many types of somatic stem cells. However, in hematopoietic stem and progenitor cells (HSPCs), the role of the canonical Wnt signaling remains unclear. A critical event in Wnt‐target gene activation is the interaction of ß‐catenin with TCF/LEF transcription factors which enables transcription of regulated regions. The majority of previous studies relied on ß‐catenin manipulation. Intriguingly, ß‐catenin has been described to have also important cytoplasmic functions and therefore, it is not clear whether the previously observed phenotypes are really caused by the alteration of expression of Wnt‐target genes.

Aims:

Our aim was to investigate how specific loss of ß‐catenin interaction with TCF/LEF factors will affect the function of HSPCs in vivo and to decipher molecular mechanism that is responsible for the functional changes.

Methods:

To investigate the function of canonical Wnt signaling pathway in HSPCs, we disrupted ß‐catenin interaction with TCF/LEF factors by expressing a dominant negative form of human TCF4 (dnTCF4) specifically in hematopoietic tissues. dnTCF4 forms a heterodimers with endogenous TCF factors and as it lacks ß‐catenin binding domain, it prevents the transactivation of Wnt‐target genes. The hematopoietic tissues of transgenic mice were analyzed using flow cytometry, while the function of HSPCs was assessed by bone marrow transplantation assays in vivo and colony formation assay in vitro. NF‐κB activation was assessed by intracellular staining of IκBa and analyzed by flow cytometry in various cell populations. RNA sequencing was performed in sorted short‐term HSC, defined as lin−, c‐kit+, Sca1+, CD48+, CD150−, using Illumina platform.

Results:

dnTCF4 transgenic mice presented decreased levels of granulocytes in peripheral blood accompanied by accumulation of HSPCs in bone marrow. Consistently, dnTCF4 bone marrow cells displayed granulocytic differentiation block, augmented colony‐forming abilities, and retained immature phenotype longer than wild type control in colony formation assays. The accumulation of HSPCs was also reflected by enhanced engraftment rate of dnTCF4‐expressing bone marrow after transplantation. Transcriptomic analysis revealed that dnTCF4‐expressing HSPCs have reduced expression of genes involved in granulocyte differentiation and function, including Csf3r, Il18r, and Il1r1r. Interestingly, IL‐1ß treatment fail to activate NF‐κB signaling pathway, a well described inducer of myeloid differentiation, in dnTCF4‐expressing HSPCs.

Summary/Conclusion:

Altogether, our data suggest that the canonical Wnt signaling promotes HSPCs differentiation towards granulocytes by upregulating IL‐1 receptor levels that allow proper NF‐κB activation. Since our murine model does present intact cytoplasmic ß‐catenin activity, our data supports that ß‐catenin controls HSPCs fate and differentiation via transactivation of Wnt‐target genes.