Biology subjects

Skoczylas, R.

Publications and source records attributed to Skoczylas, R..

5 recordsLinked to original sources

bioRxiv2025

Distinct topologically associated domains underlie the regulatory logic in lymphatic endothelial cells contributing to proper cell differentiation

The activation and repression of genes is a fundamental part of proper embryonic development and functional tissue formation, ensuring that unique molecular codes are set up to orchestrate cell differentiation. Changes in chromatin organisation dictate accessibility to gene regulatory elements and control gene expression. Several molecular factors regulating lymphatic endothelial cell (LEC) specification and differentiation have been identified. However, it remains to be defined how chromatin is organised in lymphatic endothelium and how it orchestrates lymphatic vessel network formation. In this study, we combined Hi-C and ATAC-sequencing to characterise 3D chromatin architecture and accessibility in LECs and blood endothelial cells (BECs). We have identified cell type-specific topologically associated domains (TADs) in LECs and BECs. Specifically, our data revealed changes in the TAD boundaries and differentially segregating enhancers regions in lymphatic-associated loci, such as prox1a and tbx1. This multi-omic approach also defined the regulatory logic for nine genes whose expression is enriched in LECs. In vivo validation of their short- and long-range enhancers confirmed their LEC-confined activity. Leveraging these datasets, we reconstructed mafba tissue-specific regulatory networks and identified a genetic interaction with tfe3a in vivo necessary to limit ectopic vessel formation. Overall, our work provides a powerful resource of multi-omic datasets that can be used to systematically determine the regulatory networks governing LEC identity and genes linked to lymphatic disease.

developmental biology↗

bioRxiv2023

Multiple cis-regulatory elements control prox1a expression in distinct lymphatic vascular beds

Lymphatic vessels play a role in several physiological and pathological processes including tissue fluid homeostasis, dietary fat absorption, immunosurveillance, and immunomodulation. During embryonic development, lymphatic endothelial cell (LEC) precursors are distinguished from blood endothelial cells by the expression of the transcription factor Prospero-related homeobox 1 (PROX1). PROX1 is essential for lymphatic vascular network formation in mouse and zebrafish. The initiation of PROX1 expression precedes LEC sprouting and migration, serving as the definitive marker of specified LECs. Despite its crucial role in lymphatic development, the upstream regulation of PROX1 in LECs remains to be uncovered. SOX18 and COUP-TFII are thought to regulate Prox1 expression in mice by binding to its promoter region. However, how the specificity of Prox1 expression to LECs is achieved remains to be studied in detail. In this study, we analysed evolutionary conservation and chromatin accessibility to identify enhancer sequences located in the proximity of zebrafish prox1a active in developing LECs. We confirmed the functional role of the identified sequences through CRISPR/Cas9 mutagenesis of a lymphatic valve enhancer. The deletion of this genomic region results in impaired valve morphology and function. Overall, our results reveal the intricate control of prox1a expression through a collection of enhancers. Ray-finned fish-specific distal enhancers drive pan-lymphatic expression, while vertebrate-conserved proximal enhancers refine expression in functionally distinct subsets of lymphatic vessels. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=89 SRC="FIGDIR/small/550483v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@fed780org.highwire.dtl.DTLVardef@f62a8dorg.highwire.dtl.DTLVardef@163b04corg.highwire.dtl.DTLVardef@1cb7f87_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2021

The blood vasculature instructs lymphatics patterning in a SOX7-dependent manner

Despite a growing catalogue of secreted factors critical for lymphatic network assembly, little is known about the mechanisms that modulate the expression level of these molecular cues in blood vascular endothelial cells (BECs). Here, we show that a BEC-specific transcription factor, SOX7, plays a crucial role in lymphatic vessel patterning by modulating the transcription of lymphangiocrine signals. While SOX7 is not expressed in lymphatic endothelial cells (LECs), loss of SOX7 function in mouse embryos causes a dysmorphic dermal lymphatic phenotype. We identify novel distant regulatory regions in mice and humans that contribute to directly repressing the transcription of a major lymphangiogenic growth factor (Vegfc) in a SOX7-dependent manner. Further, we show that SOX7 directly binds HEY1, a canonical repressor of the Notch pathway, suggesting that transcriptional repression may also be modulated by the recruitment of this protein partner at Vegfc genomic regulatory regions. Our work unveils a role for SOX7 in modulating downstream signalling events crucial for lymphatic patterning, at least in part via the transcriptional repression of VEGFC levels in the blood vascular endothelium.

developmental biology↗

bioRxiv2021

Proper migration of lymphatic endothelial cells requires survival and guidance cues from arterial mural cells

The migration of lymphatic endothelial cells (LECs) is key for the development of the complex and vast lymphatic vascular network that pervades most of the tissues in an organism. In zebrafish, arterial intersegmental vessels together with chemokines have been shown to promote lymphatic cell migration from the horizontal myoseptum (HM). Here we found that LECs departure from HM coincides with the emergence of mural cells around the intersegmental arteries, raising the possibility that arterial mural cells promote LEC migration. Our live imaging and cell ablation experiments revealed that LECs migrate slower and fail to establish the lymphatic vascular network in the absence of arterial mural cells. We determined that mural cells are a source for the C-X-C motif chemokine 12 (Cxcl12a and Cxcl12b) and vascular endothelial growth factor C (Vegfc). We showed that ERK, a downstream component of Vegfc-Vegfr3 singling cascade, is activated in migrating LECs and that both chemokine and growth factor signalling is required for the robust migration. Furthermore, Vegfc-Vegfr3 has a pro-survival role in LECs during the migration. Together, the identification of mural cells a source for signals that guide LEC migration and survival will be important in the future design for rebuilding lymphatic vessels in the disease contexts.

developmental biology↗

bioRxiv2021

Pkd1 and Wnt5a genetically interact to control lymphatic vascular morphogenesis in mice

Lymphatic vascular development is regulated by well-characterised signalling and transcriptional pathways. These pathways regulate lymphatic endothelial cell (LEC) migration, motility, polarity and and morphogenesis. Canonical and non-canonical WNT signalling pathways are known to control LEC polarity and development of lymphatic vessels and valves. PKD1, encoding Polycystin-1, is the most commonly mutated gene in polycystic kidney disease but has also been shown to be essential in lymphatic vascular morphogenesis. The mechanism by which Pkd1 acts during lymphangiogenesis remains unclear. Here we find that loss of non-canonical WNT signalling components Wnt5a and Ryk phenocopy lymphatic defects seen in Pkd1 knockout mice. To investigate genetic interaction, we generated Pkd1/Wnt5a double knockout mice. Loss of Wnt5a suppressed phenotypes seen in the lymphatic vasculature of Pkd1-/- mice and Pkd1 deletion suppressed phenotypes observed in Wnt5a-/- mice. Thus, we report mutually suppressive roles for Pkd1 and Wnt5a, with developing lymphatic networks restored to a more wild-type state in double mutant mice. This genetic interaction between Pkd1 and the non-canonical WNT signalling pathway ultimately controls LEC polarity and the morphogenesis of developing vessel networks. Our work suggests that Pkd1 acts at least in part by regulating non-canonical WNT signalling during the formation of lymphatic vascular networks.

developmental biology↗