Secreted protein acidic and rich in cysteine (SPARC), or osteonectin, is a matricellular protein that modulates interactions between cells and their microenvironment. SPARC is expressed during extracellular matrix remodeling and is abundant in bone marrow and high-grade prostate cancer (PCa). In PCa, SPARC induces changes associated with epithelial–mesenchymal transition (EMT), enhancing migration and invasion and increasing the expression of EMT transcriptional factor Zinc finger E-box-binding homeobox 1 (ZEB1), but not Zinc finger protein SNAI1 (Snail) or Zinc finger protein SNAI2 (Slug). It is unknown whether the SPARC-induced downregulation of E-cadherin in PCa cells depends on ZEB1. Several integrins are mediators of SPARC effects in cancer cells. Because integrin signaling can induce EMT programs, we hypothesize that SPARC induces E-cadherin repression through the activation of integrins and ZEB1. Through stable knockdown and the overexpression of SPARC in PCa cells, we demonstrate that SPARC downregulates E-cadherin and increases vimentin, ZEB1, and integrin β3 expression. Knocking down SPARC in PCa cells decreases the tyrosine-925 phosphorylation of FAK and impairs focal adhesion formation. Blocking integrin αvβ3 and silencing ZEB1 revert both the SPARC-induced downregulation of E-cadherin and cell migration enhancement. We conclude that SPARC induces E-cadherin repression and enhances PCa cell migration through the integrin αvβ3/ZEB1 signaling pathway.

Mass spectrometry-based proteomics methods are widely used to identify and quantify protein complexes involved in diverse biological processes. Specifically, tandem mass spectrometry methods represent an accurate and sensitive strategy for identifying protein-protein interactions. However, most of these approaches provide only lists of peptide fragments associated with a target protein, without performing further analyses to discriminate physical or functional protein-protein interactions. Here, we present the PPI-MASS web server, which provides an interactive analytics platform to identify protein-protein interactions with pharmacological potential by filtering a large protein set according to different biological features. Starting from a list of proteins detected by MS-based methods, PPI-MASS integrates an automatized pipeline to obtain information of each protein from freely accessible databases. The collected data include protein sequence, functional and structural properties, associated pathologies and drugs, as well as location and expression in human tissues. Based on this information, users can manipulate different filters in the web platform to identify candidate proteins to establish physical contacts with a target protein. Thus, our server offers a simple but powerful tool to detect novel protein-protein interactions, avoiding tedious and time-consuming data postprocessing. To test the web server, we employed the interactome of the TRPM4 and TMPRSS11a proteins as a use case. From these data, protein-protein interactions were identified, which have been validated through biochemical and bioinformatic studies. Accordingly, our web platform provides a comprehensive and complementary tool for identifying protein-protein complexes assisting the future design of associated therapies.

Background: Myogenic tone is a fundamental property of resistance arteries that stabilizes tissue perfusion by coupling intraluminal pressure to smooth muscle cell (SMC) depolarization, Ca ²⁺ influx, and vasoconstriction. TRPM4 (transient receptor potential melastatin 4) cation channels are required for this response in cerebral arteries, but whether TRPM4-dependent mechanotransduction is conserved across the broader resistance vasculature has remained unknown. Methods: We combined droplet digital PCR, a newly generated Trpm4 -Cre transgenic reporter mouse line, native-cell patch-clamp electrophysiology, pressure myography, selective pharmacological inhibition, and novel SMC-specific Trpm4 -knockout ( Trpm4 -smKO) mice to define the expression, regulation, and functional importance of TRPM4 in cerebral, mesenteric, and skeletal muscle resistance arteries. Results: Trpm4 transcripts were detected in all three vascular beds, and genetic reporter-based mapping localized TRPM4 expression to SMCs in multiple organs. Using conventional whole-cell patch-clamp electrophysiology, we recorded cation currents activated by high intracellular [Ca ²⁺ ] and sensitive to the selective TRPM4 blocker 4-chloro-2-(1-naphthyloxyacetamido) benzoic acid (NBA) in native SMCs from all three beds. In cells patch-clamped using the amphotericin B-perforated configuration, stretching the plasma membrane by applying negative pressure (-20 mmHg) through the patch pipette activated transient inward cation currents that were suppressed by NBA. The selective angiotensin II type 1 receptor (AT ₁ R) blocker losartan also inhibited stretch-induced currents without affecting Ca ²⁺ -activated whole-cell TRPM4 currents, indicating that AT ₁ R signaling is required for mechanotransduction in SMCs from all three vascular beds. In pressurized arteries with established myogenic tone, NBA produced reversible, concentration-dependent suppression of pressure-induced constriction of cerebral, mesenteric, and skeletal muscle arteries while sparing constriction induced by direct depolarization of SMCs with high (60 mM) extracellular [K ⁺ ]. TRPM4-dependent whole-cell currents and stretch-induced cation currents were decreased in SMCs from Trpm4 -smKO mice, and myogenic tone was essentially absent in all three vascular beds from these animals. Conclusions: These findings show that TRPM4 is essential for pressure-induced SMC depolarization and myogenic constriction in the resistance vasculature.

Triple-negative breast cancer has a poor prognosis and is non-responsive to first-line therapies; hence, new therapeutic strategies are needed. Enhanced store-operated Ca²⁺ entry (SOCE) has been widely described as a contributing factor to tumorigenic behavior in several tumor types, particularly in breast cancer cells. SOCE-associated regulatory factor (SARAF) acts as an inhibitor of the SOCE response and, therefore, can be a potential antitumor factor. Herein, we generated a C-terminal SARAF fragment to evaluate the effect of overexpression of this peptide on the malignancy of triple-negative breast cancer cell lines. Using both in vitro and in vivo approaches, we showed that overexpression of the C-terminal SARAF fragment reduced proliferation, cell migration, and the invasion of murine and human breast cancer cells by decreasing the SOCE response. Our data suggest that regulating the activity of the SOCE response via SARAF activity might constitute the basis for further alternative therapeutic strategies for triple-negative breast cancer.

Neurovascular coupling (NVC), which is initiated by the brain’s dense capillary network, matches blood flow to neuronal activity. We found that ORAI1 channels and their regulator STIM1, the main drivers of store-operated Ca ²⁺ entry, were essential for communication from capillaries, which detect neuronal metabolic need, to upstream arterioles, which dilate to increase regional flow. Endothelial cell–specific knockout of either Stim1 or Orai1 disrupted capillary Ca ²⁺ signals, impaired sustained capillary-driven arteriole dilation, and reduced increases in blood flow in the somatosensory cortex evoked by whisker stimulation, indicating that ORAI1 and STIM1 sustain cerebral blood flow during prolonged neuronal stimulation. Moreover, mice with endothelial cell–specific deficiency of Stim1 or Orai1 showed cognitive impairment, whereas mice with endothelial cell–specific deficiency of Orai3 showed anxiety-like behaviors. These in vivo results link impaired capillary-to-arteriole signaling to isoform-specific behavioral aberrations. These findings demonstrate that intravascular communication mediated by ORAI channels and STIM1 is fundamental for NVC and brain health.