A Therapeutic Perspective

AF was funded by a Ph.D. fellowship (PD/BD/135506/2018) from the Fundação para a Ciência e a Tecnologia (FCT) and DB by the FCT Investigator Program (IF/00501/2014/CP1252/CT0001).The Adenosine diphosphate-Ribosylation Factor (ARF) family belongs to the RAS superfamily of small GTPases and is involved in a wide variety of physiological processes, such as cell proliferation, motility and differentiation by regulating membrane traffic and associating with the cytoskeleton. Like other members of the RAS superfamily, ARF family proteins are activated by Guanine nucleotide Exchange Factors (GEFs) and inactivated by GTPase-Activating Proteins (GAPs). When active, they bind effectors, which mediate downstream functions. Several studies have reported that cancer cells are able to subvert membrane traffic regulators to enhance migration and invasion. Indeed, members of the ARF family, including ARF-Like (ARL) proteins have been implicated in tumorigenesis and progression of several types of cancer. Here, we review the role of ARF family members, their GEFs/GAPs and effectors in tumorigenesis and cancer progression, highlighting the ones that can have a pro-oncogenic behavior or function as tumor suppressors. Moreover, we propose possible mechanisms and approaches to target these proteins, toward the development of novel therapeutic strategies to impair tumor progression.publishersversionpublishe

Subversion of Ras Small GTPases in Cutaneous Melanoma Aggressiveness

The rising incidence and mortality rate associated with the metastatic ability of cutaneous melanoma represent a major public health concern. Cutaneous melanoma is one of the most invasive human cancers, but the molecular mechanisms are poorly understood. Moreover, currently available therapies are not efficient in avoiding melanoma lethality. In this context, new biomarkers of prognosis, metastasis, and response to therapy are necessary to better predict the disease outcome. Additionally, the knowledge about the molecular alterations and dysregulated pathways involved in melanoma metastasis may provide new therapeutic targets. Members of the Ras superfamily of small GTPases regulate various essential cellular activities, from signaling to membrane traffic and cytoskeleton dynamics. Therefore, it is not surprising that they are differentially expressed, and their functions subverted in several types of cancer, including melanoma. Indeed, Ras small GTPases were found to regulate melanoma progression and invasion. Hence, a better understanding of the mechanisms regulated by Ras small GTPases that are involved in melanoma tumorigenesis and progression may provide new therapeutic strategies to block these processes. Here, we review the current knowledge on the role of Ras small GTPases in melanoma aggressiveness and the molecular mechanisms involved. Furthermore, we summarize the known involvement of these proteins in melanoma metastasis and how these players influence the response to therapy.publishersversionpublishe

The pursuit of skin pigmentation control mechanisms

Funding: We would like to thank our group for critical reading of the manuscript. This work was supported by Fundação para a Ciência e a Tecnologia (FCT) through projects EXPL/BEXBCM/0379/2013 and PTDC/BIA-CEL/29765/2017. H.M. was supported by a PhD fellowship from FCT (PD/BD/114118/2015), and D.C.B. by the FCT Investigator Program (IF/00501/2014/CP1252/ CT0001). This article is supported by the LYSOCIL project. This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No. 811087.The mechanisms by which the pigment melanin is transferred from melanocytes and processed within keratinocytes to achieve skin pigmentation remain ill-characterized. Nevertheless, several models have emerged in the past decades to explain the transfer process. Here, we review the proposed models for melanin transfer in the skin epidermis, the available evidence supporting each one, and the recent observations in favor of the exo/phagocytosis and shed vesicles models. In order to reconcile the transfer models, we propose that different mechanisms could co-exist to sustain skin pigmentation under different conditions. We also discuss the limited knowledge about melanin processing within keratinocytes. Finally, we pinpoint new questions that ought to be addressed to solve the long-lasting quest for the understanding of how basal skin pigmentation is controlled. This knowledge will allow the emergence of new strategies to treat pigmentary disorders that cause a significant socio-economic burden to patients and healthcare systems worldwide and could also have relevant cosmetic applications.publishersversionpublishe

Commensal-to-pathogen transition: One-single transposon insertion results in two pathoadaptive traits in Escherichia coli -macrophage interaction

There are no funders and sponsors indicated explicitly in the document. This deposit is composed by the main article plus the supplementary materials of the publication.Escherichia coli is both a harmless commensal in the intestines of many mammals, as well as a dangerous pathogen. The evolutionary paths taken by strains of this species in the commensal-to-pathogen transition are complex and can involve changes both in the core genome, as well in the pan-genome. One way to understand the likely paths that a commensal strain of E. coli takes when evolving pathogenicity is through experimentally evolving the strain under the selective pressures that it will have to withstand as a pathogen. Here, we report that a commensal strain, under continuous pressure from macrophages, recurrently acquired a transposable element insertion, which resulted in two key phenotypic changes: increased intracellular survival, through the delay of phagosome maturation and increased ability to escape macrophages. We further show that the acquisition of the pathoadaptive traits was accompanied by small but significant changes in the transcriptome of macrophages upon infection. These results show that under constant pressures from a key component of the host immune system, namely macrophage phagocytosis, commensal E. coli rapidly acquires pathoadaptive mutations that cause transcriptome changes associated to the host-microbe duet.There are no funders and sponsors indicated explicitly in the document.info:eu-repo/semantics/publishedVersio

Unraveling the relevance of arl gtpases in cutaneous melanoma prognosis through integrated bioinformatics analysis

Funding Information: Funding: This research was funded by iNOVA4Health—UIDB/04462/2020, a program financially supported by the Fundação para a Ciência e Tecnologia/Ministério da Educação e Ciência. M.P. was funded by the Liga Portuguesa Contra o Cancro ‒ Núcleo Regional do Sul (LPCC‐NRS). D.B. was funded by the FCT Investigator Program (IF/00501/2014/CP1252/CT0001). Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland.Cutaneous melanoma (CM) is the deadliest skin cancer, whose molecular pathways underlying its malignancy remain unclear. Therefore, new information to guide evidence‐based clinical decisions is required. Adenosine diphosphate (ADP)‐ribosylation factor‐like (ARL) proteins are membrane trafficking regulators whose biological relevance in CM is undetermined. Here, we investigated ARL expression and its impact on CM prognosis and immune microenvironment through integrated bioinformatics analysis. Our study found that all 22 ARLs are differentially expressed in CM. Specifically, ARL1 and ARL11 are upregulated and ARL15 is downregulated regardless of mutational frequency or copy number variations. According to TCGA data, ARL1 and ARL15 represent independent prognostic factors in CM as well as ARL11 based on GEPIA and OncoLnc. To investigate the mechanisms by which ARL1 and ARL11 increase patient survival while ARL15 reduces it, we evaluated their correlation with the immune microenvironment. CD4+ T cells and neutrophil infiltrates are significantly increased by ARL1 expression. Furthermore, ARL11 expression was correlated with 17 out of 21 immune infiltrates, including CD8+ T cells and M2 macrophages, described as having anti‐tumoral activity. Likewise, ARL11 is interconnected with ZAP70, ADAM17, and P2RX7, which are implicated in immune cell activation. Collectively, this study provides the first evidence that ARL1, ARL11, and ARL15 may influence CM progression, prognosis, and immune microenvironment remodeling.publishersversionpublishe

Influence of the university environment in the entrepreneurial intention in public and private universities

This study analyses the entrepreneurial intention (EI) in different higher education institutional environments – public and private universities. To achieve the objectives, an EI model adapted from Krueger et al. (2000) was used, which is known as Entrepreneurial Intention Classical Model. Data was collected using a structured questionnaire applied in three public universities and three private universities in the State of Rio Grande do Sul. Through comparative analysis and employing the Difference-in-Differences econometric method, it was seen that the results are in accordance with part of previous studies which pointed out that private university students have higher entrepreneurial intentions. However, the results also revealed that private university students already had higher EI before starting their graduation courses. Therefore, there is no evidence of difference of the influence of public and private university environments in EI, or in its underlying factors (that form EI), except for the factor that expresses the desire to learn about entrepreneurship.Neste artigo, tem-se como objetivo a analise da intenção empreendedora em diferentes ambientes institucionais de educação superior – universidades públicas e privadas. Para tanto, utilizou-se um modelo de intenção empreendedora adaptado de Krueger et al. (2000), conhecido como Modelo Clássico de Intenções Empreendedoras. Os dados foram coletados por meio de um questionário estruturado aplicado em três universidades públicas e três privadas do estado do Rio Grande do Sul. Através da análise comparativa utilizando-se do método econométrico de diferenças em diferenças evidenciou-se que os resultados estão de acordo com parte de estudos anteriores mostrando que os estudantes de universidades privadas têm intenções empreendedoras mais elevadas. No entanto, os resultados revelaram que os alunos das universidades privadas já tinham intenção empreendedora mais elevada antes de entrarem no curso superior. Assim, não há evidências de que exista diferença na influência do ambiente universitário público e privado na intenção empreendera, e nem nos fatores subjacentes desta (que formam a mesma), exceto para o fator que expressa o desejo de aprender sobre empreendedorismo

Influence of the university environment in the entrepreneurial intention in public and private universities

This study analyses the entrepreneurial intention (EI) in different higher education institutional environments – public and private universities. To achieve the objectives, an EI model adapted from Krueger et al. (2000) was used, which is known as Entrepreneurial Intention Classical Model. Data was collected using a structured questionnaire applied in three public universities and three private universities in the State of Rio Grande do Sul. Through comparative analysis and employing the Difference-in-Differences econometric method, it was seen that the results are in accordance with part of previous studies which pointed out that private university students have higher entrepreneurial intentions. However, the results also revealed that private university students already had higher EI before starting their graduation courses. Therefore, there is no evidence of difference of the influence of public and private university environments in EI, or in its underlying factors (that form EI), except for the factor that expresses the desire to learn about entrepreneurship.Neste artigo, tem-se como objetivo a analise da intenção empreendedora em diferentes ambientes institucionais de educação superior – universidades públicas e privadas. Para tanto, utilizou-se um modelo de intenção empreendedora adaptado de Krueger et al. (2000), conhecido como Modelo Clássico de Intenções Empreendedoras. Os dados foram coletados por meio de um questionário estruturado aplicado em três universidades públicas e três privadas do estado do Rio Grande do Sul. Através da análise comparativa utilizando-se do método econométrico de diferenças em diferenças evidenciou-se que os resultados estão de acordo com parte de estudos anteriores mostrando que os estudantes de universidades privadas têm intenções empreendedoras mais elevadas. No entanto, os resultados revelaram que os alunos das universidades privadas já tinham intenção empreendedora mais elevada antes de entrarem no curso superior. Assim, não há evidências de que exista diferença na influência do ambiente universitário público e privado na intenção empreendera, e nem nos fatores subjacentes desta (que formam a mesma), exceto para o fator que expressa o desejo de aprender sobre empreendedorismo

Rab11 is required for lysosome exocytosis through the interaction with Rab3a, Sec15 and GRAB

Funding: This study was supported by Fundaçã o para a Ciência e Tecnologia (FCT): C.E. was supported by a post-doctoral fellowship (SFRH/BPD/78491/2011), L.B.-L. by a PhD fellowship (SFRH/BD/131938/2017) and D.C.B. by the FCT Investigator Program (IF/00501/2014/CP1252/CT0001). This work was developed with the support from the research infrastructure PPBI-POCI-01-0145-FEDER-022122, co-financed by FCT (Portugal) and Lisboa2020, under the PORTUGAL2020 agreement (European Regional Development Fund). This article was supported by the LYSOCIL project. This project has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No. 811087. Deposited in PMC for immediate release.Lysosomes are dynamic organelles, capable of undergoing exocytosis. This process is crucial for several cellular functions, namely plasma membrane repair. Nevertheless, the molecular machinery involved in this process is poorly understood. Here, we identify Rab11a and Rab11b as regulators of calcium-induced lysosome exocytosis. Interestingly, Rab11-positive vesicles transiently interact with lysosomes at the cell periphery, indicating that this interaction is required for the last steps of lysosome exocytosis. Additionally, we found that the silencing of the exocyst subunit Sec15, a Rab11 effector, impairs lysosome exocytosis, suggesting that Sec15 acts together with Rab11 in the regulation of lysosome exocytosis. Furthermore, we show that Rab11 binds the guanine nucleotide exchange factor for Rab3a (GRAB) and also Rab3a, which we described previously as a regulator of the positioning and exocytosis of lysosomes. Thus, our study identifies new players required for lysosome exocytosis and suggest the existence of a Rab11-Rab3a cascade involved in this process.publishersversionpublishe

Towards a precise test for malaria diagnosis in the Brazilian Amazon: comparison among field microscopy, a rapid diagnostic test, nested PCR, and a computational expert system based on artificial neural networks

<p>Abstract</p> <p>Background</p> <p>Accurate malaria diagnosis is mandatory for the treatment and management of severe cases. Moreover, individuals with asymptomatic malaria are not usually screened by health care facilities, which further complicates disease control efforts. The present study compared the performances of a malaria rapid diagnosis test (RDT), the thick blood smear method and nested PCR for the diagnosis of symptomatic malaria in the Brazilian Amazon. In addition, an innovative computational approach was tested for the diagnosis of asymptomatic malaria.</p> <p>Methods</p> <p>The study was divided in two parts. For the first part, passive case detection was performed in 311 individuals with malaria-related symptoms from a recently urbanized community in the Brazilian Amazon. A cross-sectional investigation compared the diagnostic performance of the RDT Optimal-IT, nested PCR and light microscopy. The second part of the study involved active case detection of asymptomatic malaria in 380 individuals from riverine communities in Rondônia, Brazil. The performances of microscopy, nested PCR and an expert computational system based on artificial neural networks (MalDANN) using epidemiological data were compared.</p> <p>Results</p> <p>Nested PCR was shown to be the gold standard for diagnosis of both symptomatic and asymptomatic malaria because it detected the major number of cases and presented the maximum specificity. Surprisingly, the RDT was superior to microscopy in the diagnosis of cases with low parasitaemia. Nevertheless, RDT could not discriminate the <it>Plasmodium </it>species in 12 cases of mixed infections (<it>Plasmodium vivax </it>+ <it>Plasmodium falciparum</it>). Moreover, the microscopy presented low performance in the detection of asymptomatic cases (61.25% of correct diagnoses). The MalDANN system using epidemiological data was worse that the light microscopy (56% of correct diagnoses). However, when information regarding plasma levels of interleukin-10 and interferon-gamma were inputted, the MalDANN performance sensibly increased (80% correct diagnoses).</p> <p>Conclusions</p> <p>An RDT for malaria diagnosis may find a promising use in the Brazilian Amazon integrating a rational diagnostic approach. Despite the low performance of the MalDANN test using solely epidemiological data, an approach based on neural networks may be feasible in cases where simpler methods for discriminating individuals below and above threshold cytokine levels are available.</p

Regulation of CD1 Antigen-presenting Complex Stability

For major histocompatibility complex class I and II molecules, the binding of specific peptide antigens is essential for assembly and trafficking and is at the center of their quality control mechanism. However, the role of lipid antigen binding in stabilization and quality control of CD1 heavy chain (HC).beta(2)-microglobulin (beta(2)m) complexes is unclear. Furthermore, the distinct trafficking and loading routes of CD1 proteins take them from mildly acidic pH in early endososmal compartments (pH 6.0) to markedly acidic pH in lysosomes (pH 5.0) and back to neutral pH of the cell surface (pH 7.4). Here, we present evidence that the stability of each CD1 HC.beta(2)m complex is determined by the distinct pH optima identical to that of the intracellular compartments in which each CD1 isoform resides. Although stable at acidic endosomal pH, complexes are only stable at cell surface pH 7.4 when bound to specific lipid antigens. The proposed model outlines a quality control program that allows lipid exchange at low endosomal pH without dissociation of the CD1 HC.beta(2)m complex and then stabilizes the antigen-loaded complex at neutral pH at the cell surface