Matteini, F. | bioRxiv Science

bioRxiv2025

Targeting RhoA activity rejuvenates aged hematopoietic stem cells

Biomechanical alterations contribute to the decreased regenerative capacity of hematopoietic stem cells (HSCs) upon aging. RhoA is a key regulator of mechano-signaling but its role for mechanotransduction in stem cell aging has not been investigated yet. Here, we show that murine HSCs respond to increased nuclear envelope (NE) tension by inducing NE translocation of P-cPLA2, which cell intrinsically activates RhoA. Interestingly, aged HSCs experience physiologically higher intrinsic NE tension, associated with increased NE P-cPLA2 and RhoA activity. Reducing RhoA activity lowers NE tension in aged HSCs. Feature image analysis of HSC nuclei reveals that chromatin remodeling is associated to RhoA inhibition, which includes the restoration of youthful levels of the heterochromatin marker H3K9me2 and a decrease in chromatin accessibility and transcription at retrotransposons. Eventually, we demonstrate that RhoA inhibition upregulates Klf4 expression and transcriptional activity, improving aged HSCs regenerative capacity and lympho/myeloid skewing in vivo. Overall, our data support that an intrinsic mechano-signaling axis dependent on RhoA can be pharmacologically targeted to rejuvenate stem cell function upon aging.

cell biology↗

bioRxiv2023

Radical fringe facilitates NOTCH1 and JAG1 cis interactions to sustain Hematopoietic stem cell fate

Hematopoietic stem cells (HSCs) develop within a short time window from the hemogenic endothelium in the aorta- gonads-and mesonephros (AGM) region during embryonic development. The first HSCs reside within Intra-aortic hematopoietic clusters (IAHC) along with hematopoietic progenitors (HPC). The signalling mechanisms that divert HSCs from HPCs are unknown. Notch signaling is essential for arterial specification, IAHC formation and HSC activity, but current studies on how Notch drives these different fates are inconsistent. To determine the role of Notch in the specification of hemogenic endothelium, HSC and/or HPCs, we extensively analyzed Notch dynamics in the period of HSC generation. We defined the expression pattern of Notch signalling molecules at the gene and protein level and established a molecular mechanism that reconcile previous studies demonstrating the loss of HSC activity in NOTCH1, JAG1 and RBPJ null mutants, the enhanced HSC generation by blocking specific Notch activities or the abrogation of emerging HSCs by high Notch activation. We now demonstrate that Notch activity is highest in a subset of Gfi1+ hemogenic endothelial cells and is gradually lost with HSC maturation. We uncover that the HSC phenotype is maintained through loss of Notch activity due to increasing levels of NOTCH1 and JAG1 interactions on the surface of the same cell (cis) that renders the NOTCH1 receptor from being activated. Forcing activation of the NOTCH1 receptor in IAHC cells activates a hematopoietic differentiation program and supports a cis-inhibitory function for JAG1 and NOTCH1. Furthermore, we demonstrate that this cis-inhibitory interaction is enabled by RADICAL FRINGE (RFNG), a glycosyltransferase that enhances the affinity of NOTCH1 to JAG1 in cis. Finally, our results indicate that NOTCH1-JAG1 cis-inhibition is necessary for preserving the HSC phenotype in the hematopoietic clusters of the aorta.

developmental biology↗