The Rho GTPase exchange factor Vav2 promotes extensive age-dependent rewiring of the hair follicle stem cell transcriptome

Both the number and regenerative activity of hair follicle stem cells (HFSCs) are regulated by Vav2, a GDP/GTP exchange factor involved in the catalytic stimulation of the GTPases Rac1 and RhoA. However, whether Vav2 signaling changes in HFSCs over the mouse lifespan is not yet known. Using a mouse knock-in mouse model, we now show that the expression of a catalytically active version of Vav2 (Vav2Onc) promotes an extensive rewiring of the overall transcriptome of HFSCs, the generation of new transcription factor hubs, and the synchronization of many transcriptional programs associated with specific HFSC states and well-defined signaling pathways. Interestingly, this transcriptome rewiring is not fixed in time, as it involves the induction of 15 gene expression waves with diverse distribution patterns during the life of the animals. These expression waves are consistent with the promotion by Vav2Onc of several functional HFSC states that differ from those normally observed in wild-type HFSCs. These results further underscore the role of Vav2 in the regulation of the functional state of HFSCs. They also indicate that, unlike other Vav2-dependent biological processes, the signaling output of this exchange factor is highly contingent on age-dependent intrinsic and/or extrinsic HFSC factors that shape the final biological readouts triggered in this cell type

HERC ubiquitin ligases in cancer

HERC proteins are ubiquitin E3 ligases of the HECT family. The HERC subfamily is composed of six members classified by size into large (HERC1 and HERC2) and small (HERC3-HERC6). HERC family ubiquitin ligases regulate important cellular processes, such as neurodevelopment, DNA damage response, cell proliferation, cell migration, and immune responses. Accumulating evidence also shows that this family plays critical roles in cancer. In this review, we provide an integrated view of the role of these ligases in cancer, highlighting their bivalent functions as either oncogenes or tumor suppressors, depending on the tumor type. We include a discussion of both the molecular mechanisms involved and the potential therapeutic strategies

Cancertool: A visualization and representation interface to exploit cancer datasets

With the advent of OMICs technologies, both individual research groups and consortia have spear-headed the characterization of human samples of multiple pathophysiologic origins, resulting in thousands of archived genomes and transcriptomes. Although a variety of web tools are now available to extract information from OMICs data, their utility has been limited by the capacity of nonbioinformatician researchers to exploit the information. To address this problem, we have developed CANCERTOOL, a web-based interface that aims to overcome the major limitations of public transcriptomics dataset analysis for highly prevalent types of cancer (breast, prostate, lung, and colorectal). CANCERTOOL provides rapid and comprehensive visualization of gene expression data for the gene(s) of interest in well-annotated cancer datasets. This visualization is accompanied by generation of reports customized to the interest of the researcher (e.g., editable figures, detailed statistical analyses, and access to raw data for reanalysis). It also carries out gene-to-gene correlations in multiple datasets at the same time or using preset patient groups. Finally, this new tool solves the time-consuming task of performing functional enrichment analysis with gene sets of interest using up to 11 different databases at the same time. Collectively, CANCERTOOL represents a simple and freely accessible interface to interrogate well-annotated datasets and obtain publishable representations that can contribute to refinement and guidance of cancer-related investigations at all levels of hypotheses and design.We are grateful to Iñaki Lazaro for the design of the tumor type logos, Evarist Planet and Antoni Berenguer for insightful discussions, and the Carracedo lab for valuable input. V. Torrano is funded by Fundación Vasca de Innovación e Investigación Sanitarias, BIOEF (BIO15/CA/052), the AECC J.P. Bizkaia and the Basque Department of Health (2016111109). The work of A. Carracedo is supported by the Basque Department of Industry, Tourism and Trade (Etortek) and the Department of Education (IKERTALDE IT1106-16, also participated by A. Gomez-Muñoz), the BBVA Foundation, the MINECO [SAF2016-79381-R (FEDER/EU)]; Severo Ochoa Excellence Accreditation SEV-2016-0644; Excellence Networks (SAF2016-81975-REDT), European Training Networks Project (H2020-MSCA-ITN-308 2016 721532), the AECC IDEAS16 (IDEAS175CARR), and the European Research Council (Starting Grant 336343, PoC 754627). CIBERONC was cofunded with FEDER funds. The work of A. Aransay is supported by the Basque Department of Industry, Tourism and Trade (Etortek and Elkartek Programs), the Innovation Technology Department of Bizkaia County, CIBERehd Network, and Spanish MINECO the Severo Ochoa Excellence Accreditation (SEV-2016-0644). I. Apaolaza is funded by a Basque Government predoctoral grant (PRE_2017_2_0028). X.R. Bustelo is supported by grants from the Castilla-León Government (BIO/SA01/15, CSI049U16), Spanish Ministry of Economy and Competitiveness (MINECO; SAF2015-64556-R), Worldwide Cancer Research (14-1248), Ramón Areces Foundation, and the Spanish Society against Cancer (GC16173472GARC). Funding from MINECO to X.R. Bustelo is partially contributed by the European Regional Development Fund. The work of F.J. Planes is supported by the MINECO (BIO2016-77998-R) and ELKARTEK Programme of the Basque Government (KK-2016/00026)

A hotspot mutation targeting the R-RAS2 GTPase acts as a potent oncogenic driver in a wide spectrum of tumors

A missense change in RRAS2 (Gln to Leu), analogous to the Gln-to-Leu mutation of RAS oncoproteins, has been identified as a long-tail hotspot mutation in cancer and Noonan syndrome. However, the relevance of this mutation for in vivo tumorigenesis remains understudied. Here we show, using an inducible knockin mouse model, that R-Ras2 triggers rapid development of a wide spectrum of tumors when somatically expressed in adult tissues. These tumors show limited overlap with those originated by classical Ras oncogenes. R-Ras2-driven tumors can be classified into different subtypes according to therapeutic susceptibility. Importantly, the most relevant R-Ras2-driven tumors are dependent on mTORC1 but independent of phosphatidylinositol 3-kinase-, MEK-, and Ral guanosine diphosphate (GDP) dissociation stimulator. This pharmacological vulnerability is due to the extensive rewiring by R-Ras2 of pathways that orthogonally stimulate mTORC1 signaling. These findings demonstrate that RRAS2 is a bona fide oncogenic driver and unveil therapeutic strategies for patients with cancer and Noonan syndrome bearing RRAS2 mutations.We thank M. Blázquez and the personnel of the CIC Flow Cytometry, Microscopy, Pathology, and Genomics Units for expert technical work. X.R.B.’s project leading to these results has received funding from the Spanish Association against Cancer (GC16173472GARC), the Castilla-León government (CSI252P18, CSI145P20, and CLC-2017-01), the RTI2018-096481-B-100 grant cofounded by MCIN/AEI/10.13039/501100011033 and the European Research Development Fund “A way of making Europe” of the European Union, and “la Caixa” Banking Foundation (HR20-00164). X.R.B.’s institution is supported by the Programa de Apoyo a Planes Estratégicos de Investigación de Estructuras de Investigación de Excelencia of the Castilla-León government (CLC-2017-01). J.R.-V. received funding from the Carlos III Health Institute (PI20/01724). J.R.-V.’s contract is supported by a senior postdoctoral contract of the Spanish Association against Cancer. L.C.’s contract was supported by contracts from the Spanish Association against Cancer and the CLC-2017-01 grant. L.F.L.-M.’s contract was mostly supported by funding from the Spanish Ministry of Education, Culture and Sports (FPU13/02923) and, subsequently, by the CLC-2017-01 grant. R.C. was supported by a predoctoral contract from the MSI (BES-2016-0077909) and the CLC-2017-01 grant

Computational and in vitro Pharmacodynamics Characterization of 1A-116 Rac1 Inhibitor: Relevance of Trp56 in Its Biological Activity

In the last years, the development of new drugs in oncology has evolved notably. In particular, drug development has shifted from empirical screening of active cytotoxic compounds to molecularly targeted drugs blocking specific biologic pathways that drive cancer progression and metastasis. Using a rational design approach, our group has developed 1A-116 as a promising Rac1 inhibitor, with antitumoral and antimetastatic effects in several types of cancer. Rac1 is over activated in a wide range of tumor types and and it is one of the most studied proteins of the Rho GTPase family. Its role in actin cytoskeleton reorganization has effects on endocytosis, vesicular trafficking, cell cycle progression and cellular migration. In this context, the regulatory activity of Rac1 affects several key processes in the course of the cancer including invasion and metastasis. The purpose of this preclinical study was to focus on the mode of action of 1A-116, conducting an interdisciplinary approach with in silico bioinformatics tools and in vitro assays. Here, we demonstrate that the tryptophan 56 residue is necessary for the inhibitory effects of 1A-116 since this compound interferes with protein-protein interactions (PPI) of Rac1GTPase involving several GEF activators. 1A116 is also able to inhibit the oncogenic Rac1P29S mutant protein, one of the oncogenic drivers found in sun-exposed melanoma. It also inhibits numerous Rac1-regulated cellular processes such as membrane ruffling and lamellipodia formation. These results deepen our knowledge of 1A-116 inhibition of Rac1 and its biological impact on cancer progression. They also represent a good example of how in silico analyses represent a valuable approach for drug development.Fil: González, Nazareno. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Cardama, Georgina Alexandra. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Chinestrad, Patricio Manuel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Farmacología Molecular; ArgentinaFil: Robles Valero, Javier. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; EspañaFil: Rodríguez Fdez, Sonia. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; EspañaFil: Lorenzo Martín, L. Francisco. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; EspañaFil: Bustelo, Xosé R.. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; EspañaFil: Lorenzano Menna, Pablo. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Farmacología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Gomez, Daniel Eduardo. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Oncología Molecular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Vagal afferents contribute to sympathoexcitation-driven metabolic dysfunctions

Multiple crosstalk between peripheral organs and the nervous system are required to maintain physiological and metabolic homeostasis. Using Vav3-deficient mice as a model for chronic sympathoexcitation-associated disorders, we report here that afferent fibers of the hepatic branch of the vagus nerve are needed for the development of the peripheral sympathoexcitation, tachycardia, tachypnea, insulin resistance, liver steatosis and adipose tissue thermogenesis present in those mice. This neuronal pathway contributes to proper activity of the rostral ventrolateral medulla, a sympathoregulatory brainstem center hyperactive in Vav3−/− mice. Vagal afferent inputs are also required for the development of additional pathophysiological conditions associated with deregulated rostral ventrolateral medulla activity. By contrast, they are dispensable for other peripheral sympathoexcitation-associated disorders sparing metabolic alterations in liver.X R B is supported by grants from the Castilla-León Government (CSI252P18, CLC-2017-01), the Spanish Ministry of Science, Innovation and Universities (MSIU) (SAF2015-64556-R), Worldwide Cancer Research (14-1248), the Ramón Areces Foundation, and the Spanish Association against Cancer (GC16173472GARC). X R B’s institution is supported by the Programa de Apoyo a Planes Estratégicos de Investigación de Estructuras de Investigación de Excelencia of the Ministry of Education of the Castilla-León Government (CLC-2017-01). S R-F and L F L-M contracts have been supported by funding from the MISIU (BES-2013-063573) and the Spanish Ministry of Education, Culture and Sports (L F L-M, FPU13/02923), respectively. Both Spanish and Castilla-León government-associated funding is partially supported by the European Regional Development Fund

Characterization of mutant versions of the R-RAS2/TC21 GTPase found in tumors

The R-RAS2 GTP hydrolase (GTPase) (also known as TC21) has been traditionally considered quite similar to classical RAS proteins at the regulatory and signaling levels. Recently, a long-tail hotspot mutation targeting the R-RAS2/TC21 Gln72 residue (Q72L) was identified as a potent oncogenic driver. Additional point mutations were also found in other tumors at low frequencies. Despite this, little information is available regarding the transforming role of these mutant versions and their relevance for the tumorigenic properties of already-transformed cancer cells. Here, we report that many of the RRAS2 mutations found in human cancers are highly transforming when expressed in immortalized cell lines. Moreover, the expression of endogenous R-RAS2Q72L is important for maintaining optimal levels of PI3K and ERK activities as well as for the adhesion, invasiveness, proliferation, and mitochondrial respiration of ovarian and breast cancer cell lines. Endogenous R-RAS2Q72L also regulates gene expression programs linked to both cell adhesion and inflammatory/immune-related responses. Endogenous R-RAS2Q72L is also quite relevant for the in vivo tumorigenic activity of these cells. This dependency is observed even though these cancer cell lines bear concurrent gain-of-function mutations in genes encoding RAS signaling elements. Finally, we show that endogenous R-RAS2, unlike the case of classical RAS proteins, specifically localizes in focal adhesions. Collectively, these results indicate that gain-of-function mutations of R-RAS2/TC21 play roles in tumor initiation and maintenance that are not fully redundant with those regulated by classical RAS oncoproteins

Ozone Eliminates SARS-CoV-2 from Difficult-to-Clean Office Supplies and Clinical Equipment.

Background: Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) continues to cause profound health, economic, and social problems worldwide. The management and disinfection of materials used daily in health centers and common working environments have prompted concerns about the control of coronavirus disease 2019 (COVID-19) infection risk. Ozone is a powerful oxidizing agent that has been widely used in disinfection processes for decades. The aim of this study was to assess the optimal conditions of ozone treatment for the elimination of heat-inactivated SARS-CoV-2 from office supplies (personal computer monitors, keyboards, and computer mice) and clinical equipment (continuous positive airway pressure tubes and personal protective equipment) that are difficult to clean. (2) Methods: The office supplies and clinical equipment were contaminated in an area of 1 cm2 with 1 × 104 viral units of a heat-inactivated SARS-CoV-2 strain, then treated with ozone using two different ozone devices: a specifically designed ozonation chamber (for low–medium ozone concentrations over large volumes) and a clinical ozone generator (for high ozone concentrations over small volumes). SARS-CoV-2 gene detection was carried out using quantitative real-time polymerase chain reaction (RT-qPCR). (3) Results: At high ozone concentrations over small surfaces, the ozone eliminated SARS-CoV-2 RNA in short time periods—i.e., 10 min (at 4000 ppm) or less. The optimum ozone concentration over large volumes was 90 ppm for 120 min in ambient conditions (24 °C and 60–75% relative humidity). (4) Conclusions: This study showed that the appropriate ozone concentration and exposure time eliminated heat-inactivated SARS-CoV-2 RNA from the surfaces of different widely used clinical and office supplies, decreasing their risk of transmission, and improving their reutilization. Ozone may provide an additional tool to control the spread of the COVID-19 pandemic.TRUEInstituto de Salud Carlos III, Madrid, Spain, and by the European Regional Development Funds (FEDER)Fundación Canaria del Instituto de Investigación Sanitaria de Canarias (FIISC), Las PalmasFundación Mapfre Guanarteme, Las PalmasGobierno de Canarias, Las Palmaspu

Functional specificity of the members of the Sos family of Ras-GEF Activators: Novel role of Sos2 in control of epidermal stem cell homeostasis

© 2021 by the authors.Prior reports showed the critical requirement of Sos1 for epithelial carcinogenesis, but the specific functionalities of the homologous Sos1 and Sos2 GEFs in skin homeostasis and tumorigenesis remain unclear. Here, we characterize specific mechanistic roles played by Sos1 or Sos2 in primary mouse keratinocytes (a prevalent skin cell lineage) under different experimental conditions. Functional analyses of actively growing primary keratinocytes of relevant genotypes—WT, Sos1-KO, Sos2-KO, and Sos1/2-DKO—revealed a prevalent role of Sos1 regarding transcriptional regulation and control of RAS activation and mechanistic overlapping of Sos1 and Sos2 regarding cell proliferation and survival, with dominant contribution of Sos1 to the RAS-ERK axis and Sos2 to the RAS-PI3K/AKT axis. Sos1/2-DKO keratinocytes could not grow under 3D culture conditions, but single Sos1-KO and Sos2-KO keratinocytes were able to form pseudoepidermis structures that showed disorganized layer structure, reduced proliferation, and increased apoptosis in comparison with WT 3D cultures. Remarkably, analysis of the skin of both newborn and adult Sos2-KO mice uncovered a significant reduction of the population of stem cells located in hair follicles. These data confirm that Sos1 and Sos2 play specific, cell-autonomous functions in primary keratinocytes and reveal a novel, essential role of Sos2 in control of epidermal stem cell homeostasis.The E.S. group was supported by grants from ISCIII-MCUI (FIS PI19/00934), JCyL (SA264P18-UIC 076), Areces Foundation (CIVP19A5942), Solorzano-Barruso Foundation (FS/32-2020), and by ISCIII-CIBERONC (group CB16/12/00352). Research was co-financed by FEDER funds. The J.M.P. lab is co-funded by European Regional Development Fund (FEDER) grants from Science and Innovation (SAF2015-66015-R and PID2019-110758RB-I00 to J.M.P.) and Instituto de Salud Carlos III (CIBERONC no. CB16/12/00228 to J.M.P.). The XRB lab is funded by “la Caixa” Banking Foundation (HR20-00164), the Castilla-León autonomous government (CSI252P18, CSI145P20, CLC-2017-01), the Spanish Ministry of Science and Innovation (MSI) (RTI2018-096481-B-100), and the Spanish Association against Cancer (GC16173472GARC). The CIC is supported by the Programa de Apoyo a Planes Estratégicos de Investigación de Estructuras de Investigación de Excelencia of the Castilla-León autonomous government (CLC-2017-01). L.F.L.-M. and N.F.-P. contracts have been supported by funding from the Spanish Ministry of Universities (FPU13/02923, FPU17/03912) and, in the case of L.F.L.M., by CLC-2017-01 grant

RAS at the Golgi antagonizes malignant transformation through PTPRκ-mediated inhibition of ERK activation

© The Author(s) 2018.RAS GTPases are frequently mutated in human cancer. H- and NRAS isoforms are distributed over both plasma-membrane and endomembranes, including the Golgi complex, but how this organizational context contributes to cellular transformation is unknown. Here we show that RAS at the Golgi is selectively activated by apoptogenic stimuli and antagonizes cell survival by suppressing ERK activity through the induction of PTPRκ, which targets CRAF for dephosphorylation. Consistently, in contrast to what occurs at the plasma-membrane, RAS at the Golgi cannot induce melanoma in zebrafish. Inactivation of PTPRκ, which occurs frequently in human melanoma, often coincident with TP53 inactivation, accelerates RAS-ERK pathway-driven melanomagenesis in zebrafish. Likewise, tp53 disruption in zebrafish facilitates oncogenesis driven by RAS from the Golgi complex. Thus, RAS oncogenic potential is strictly dependent on its sublocalization, with Golgi complex-located RAS antagonizing tumor development.We are grateful to Drs: Ignacio Rubio, Yardena Samuels, Mariano Barbacid and Javier León for providing reagents; and Alicia Noriega, Sandra Zunzunegui y Victor Campa for technical support. Crespo laboratory is supported by grant SAF-2015 63638R (MINECO/ FEDER, UE); by Red Temática de Investigación Cooperativa sobre el Cáncer (RTICC). RD/12/0036/0033 and by Asociación Española Contra el Cáncer (AECC), grant GCB141423113. Work in the Hurlstone laboratory was unded by a grant from the European Research Council (ERC-2011-StG-282059 PROMINENT). B.C. is supported by a Retos Jóvenes Investigadores grant SAF2015-73364-JIN (AEI/FEDER, UE) and a grant from Fundación Francisco Cobos. X.R.B. is supported by grants from the CastillaLeón Government (BIO/SA01/15, CSI049U16), MINECO (SAF2015-64556-R, RD12/ 0036/0002), Worldwide Cancer Research (14-1248), Ramón Areces Foundation, andAECC (GC16173472GARC). Spanish funding to P.C., B.C., and X.R.B. is partially supported by the European Regional Development Fund