Biology subjects

Godinho, S.

Publications and source records attributed to Godinho, S..

4 recordsLinked to original sources

bioRxiv2025

Endothelial β-PIX (ARHGEF7) drives exocytosis through enabling the dynamic reorganisation of the cytoskeleton

Endothelial cells rapidly respond to blood vessel injury or infection by releasing haemostatic and inflammatory proteins from their secretory organelles called Weibel-Palade Bodies (WPBs). Upon stimulation, WPBs traffic to the cell surface to secrete cargo such as von Willebrand factor (VWF), essential for platelet recruitment, and P-selectin, which facilitates the capture and subsequent rolling of immune cells. While this process is critical, many aspects of WPB trafficking and exocytosis remain unclear--particularly how release sites are selected across the cell surface and how fusion is directed to either the apical (lumen-facing) or basolateral (basement membrane-facing) side. These decisions directly impact the biological consequences and effectiveness of the secreted proteins. Using live-cell imaging, fibronectin micropatterning, loss-of-function and biochemical assays, we identified trafficking and release at hot spots near focal adhesions (FAs). We also discovered that the FA-resident guanine nucleotide exchange factor (GEF), {beta}-PIX (ARHGEF7), regulates VWF secretion. {beta}-PIX is known to activate Rho GTPases (Cdc42/Rac), triggering downstream p21-activated kinase 2 (PAK2) signalling and cytoskeletal remodelling. Here, depletion of {beta}-PIX impaired VWF secretion and delayed its release by perturbing cytoskeletal reorganisation. Knockdown and rescue experiments using truncated mutants further revealed which domains of {beta}-PIX are necessary for exocytosis and cytoskeletal reorganisation. This is the first demonstration of {beta}-PIXs role in VWF secretion from endothelial cells and our data provides new insights into spatial targeting of WPB exocytosis. Such targeting may be essential for guiding leukocyte transmigration or platelet binding, thereby maintaining vascular integrity. SchematicEndothelial secretory organelles, known as Weibel-Palade bodies (WPBs), store von Willebrand factor (VWF), and their regulated exocytosis is critical for blood clotting and immune responses. This process can be stimulated experimentally by phorbol esters as well as physiological stimuli such as adrenaline and histamine. Cytoskeletal remodelling is required for both pre- and post-fusion events in this secretory pathway. Here, we present evidence supporting the involvement of focal adhesion (FA)-associated exocytosis in the release of WPBs. For the first time, we demonstrate a regulatory role for the FA-resident guanine nucleotide exchange factor (GEF) {beta}-PIX in VWF secretion. Through its GEF activity toward Cdc42 and Rac, {beta}-PIX activates p21-activated kinases, driving the necessary cytoskeletal dynamics. Disruption of {beta}-PIX function results in abnormal cytoskeletal architecture that fails to respond appropriately to phorbol ester stimulation. Consequently, F-actin, nonmuscle myosin IIA (NMIIA), and septins become mislocalised, forming prominent filaments above the nucleus. These data provide a novel insight into the spatial targeting and molecular events underpinning the exocytosis of endothelial secretory organelles. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/652370v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@1dbc615org.highwire.dtl.DTLVardef@1a9a959org.highwire.dtl.DTLVardef@1ad31aforg.highwire.dtl.DTLVardef@1990000_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2024

Multi-Temporal Remote Sensing of Inland Surface Waters: A Fusion of Sentinel-1 Data Applied to Small Seasonal Ponds in Semiarid Environments

Inland freshwater resources in semiarid environments play a key role in maintaining ecological systems and supporting human development. Space-based remote sensing spatiotemporal data have emerged as a new paradigm for understanding ecohydrological processes and trends, particularly in water-stressed areas. However, comprehensive cataloging is still lacking, especially in semi-arid regions and for small-sized water bodies (i.e., ponds), which are often overlooked despite their ecological relevance. In this study, high-resolution optical and radar Sentinel data (Sentinel-1 and Sentinel-2) were used to construct Sentinel-1&2-based local surface water (SLSW) models, to infer surface water occurrence and extent. To assess the reliability of this model, the results were compared with verification data, and separately with Landsat-based global water (LGSW) models. Three distinct semiarid regions were selected in SW Iberia, within a Mediterranean climate, each encompassing special protection areas for conservation and subjected to marked seasonality and bioclimatic changes. Surface water attributes were modeled using Random Forests for SLSW time series forecasting, which included the period from January 1, 2020, to December 31, 2021. During this period, the completeness of the archived information was compared between SLSW and LGSW, considering both intra-annual and inter-annual variations. The predictive performance of these models was then compared for specific periods (dry and wet), and each was independently validated with verification data. The results showed that SLWM achieved satisfactory predictive performances in detecting surface water occurrence ({approx}72%), with far greater completeness and reconstructed seasonality patterns compared to LGSW. The relatedness between SLSW and LGSW was stronger during wet periods (R2=0.38) than dry periods (R2=0.05), and SLSW related much better with the verification data (R2=0.66) than when compared to LGSW (R2=0.24). The proposed SLSW approach may therefore provide advantages in the delineation of dynamic surface water characteristics (occurrence and extent) in very small-sized water bodies (i.e., <0.5 ha), allowing for uninterrupted surface water time series forecasting at high spatiotemporal detail, and over extensive areas. Given the water constraints in semiarid regions and water resources vulnerability to climate change, our results show high potential for supporting a variety of activities underlying rural development and biodiversity conservation. Additionally, the socio-ecological applications of this research may help identify surface water anomalies (e.g., drought events) and enhance sustainable water supply governance, a particular priority in climate change hotspots. HighlightsO_LISurface water occurrence and extent was modeled across three semiarid regions C_LIO_LISentinel-1&2 data was compared with Landsat for characterizing very small water bodies C_LIO_LIModels based on Sentinel-1&2 resulted in a satisfactory classification precision C_LIO_LIVery high series completeness across seasons was found using Sentinel-1&2 data C_LIO_LISentinel-1&2 data was more reliable than Landsat when compared to verification data C_LI

ecology↗

bioRxiv2024

Dysregulated SASS6 expression promotes increased ciliogenesis and cell invasion phenotypes

Centriole and/or cilia defects are characteristic of cancer cells and have been linked to cancer cell invasion. However, the mechanistic basis of these effects is unknown. Spindle assembly abnormal protein 6 homolog (SAS-6) is essential for centriole biogenesis and cilia formation. In cycling cells, SAS-6 undergoes APCCdh1-mediated targeted degradation by the 26S proteasome at the end of mitosis. Little is known about the function of SAS-6 outside of centrosome biogenesis. To examine this, we expressed a non-degradable SAS-6 mutant (SAS-6ND). Expression of SAS-6ND led to an increase in ciliation and cilia-dependent cell invasion, and caused an upregulation of the YAP/TAZ pathway. YAP/TAZ or ciliogenesis inhibition prevented SAS-6-induced invasion. SAS-6ND caused increased actin alignment and stress fiber coherency, and nuclear flattening known to promote YAP nuclear import. Finally, data from The Cancer Genome Atlas showed that SAS-6 overexpression is associated with poor prognosis in various cancers. Our data provide evidence for a defined role of SAS-6 in cancer cell invasion and offers mechanistic insight into the role of YAP/TAZ in this cilia-sensitive process. Synopsis O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=157 SRC="FIGDIR/small/576599v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@1cec0daorg.highwire.dtl.DTLVardef@d73051org.highwire.dtl.DTLVardef@1a53e47org.highwire.dtl.DTLVardef@19bb05d_HPS_FORMAT_FIGEXP M_FIG C_FIG SAS-6 overexpressing cells show increased ciliation, actin cytoskeleton reorganization, cell flattening, YAP pathway activation and increased invasion

cell biology↗

bioRxiv2022

Low HER2 enables dedifferentiation and transformation of normal breast epithelial cells via chromatin opening

Overexpression of the human epidermal growth factor 2 (HER2) protein in breast cancer patients is a predictor of poor prognosis and resistance to therapies. Despite significant advances in the development of targeted therapies and improvements in the 5-year survival rate of metastatic HER2-positive breast cancer patients, a better understanding of the disease at an early stage is needed to prevent its progression. Here, we used an inducible breast cancer transformation system that allows investigation of early molecular changes at high temporal resolution. HER2 overexpression to similar levels as those observed in a subtype of HER2 positive breast cancer patients induced transformation of MCF10A cells and resulted in gross morphological changes, increased anchorage-independent growth of cells, and altered transcriptional programme of genes associated with oncogenic transformation. Global phosphoproteomic analysis during the first few hours of HER2 induction predominantly detected an increase in protein phosphorylation. Intriguingly, this correlated with a wave of chromatin opening, as measured by ATAC-seq on acini isolated from 3D cell culture. We observed that HER2 overexpression leads to reprogramming of many distal regulatory regions and promotes reprogramming-associated heterogeneity. We found that a subset of cells acquired a dedifferentiated breast stem-like phenotype, making them likely candidates for malignant transformation. Our data show that this population of cells, which counterintuitively enriches for relatively low HER2 protein abundance and increased chromatin accessibility, possesses transformational drive, resulting in increased anchorage-independent growth in vitro compared to cells not displaying a stem-like phenotype. Our data provide a discovery platform for signalling to chromatin pathways in HER2-driven cancers, offering an opportunity for biomarker discovery and identification of novel drug targets.

cancer biology↗