Expression of mTOR in normal and pathological conditions

The H&TRC authors gratefully acknowledge the FCT/MCTES national support through the UIDB/05608/2020 and UIDP/05608/2020. This work was part of the IPL/2021/GATumor_ESTeSL project.The mechanistic/mammalian target of rapamycin (mTOR), a protein discovered in 1991, integrates a complex pathway with a key role in maintaining cellular homeostasis. By comprising two functionally distinct complexes, mTOR complex 1 (mTORC1) and mTORC2, it is a central cellular hub that integrates intra- and extracellular signals of energy, nutrient, and hormone availability, modulating the molecular responses to acquire a homeostatic state through the regulation of anabolic and catabolic processes. Accordingly, dysregulation of the mTOR pathway has been implicated in a variety of human diseases. While major advances have been made regarding the regulators and effectors of the mTOR signaling pathway, insights into the regulation of mTOR gene expression are beginning to emerge. Here, we present the currently available data regarding the mTOR expression regulation at the level of transcription, translation, and mRNA stability and systematize the current knowledge about the fluctuations of mTOR expression observed in several diseases, both cancerous and non-cancerous. In addition, we discuss whether mTOR expression changes can be used as a biomarker for diagnosis, disease progression, prognosis, and/or response to therapeutics. We believe that our study will contribute to the implementation of new disease biomarkers based on mTOR as it gives an exhaustive perspective on the regulation of mTOR gene expression in both normal and pathological conditions.info:eu-repo/semantics/publishedVersio

Workshop de pipetagem

Instrumentos que permitem a medição e transferência precisa e rigorosa de líquidos.N/

Síntese proteica e patogénese

Mecanismo canónico da iniciação da síntese proteica: regulação da iniciação da síntese proteica, tradução mediada por IRESs, tradução mediada por CITEs, outras formas da iniciação da síntese proteica: N6-methyladenosine, 5’TOP, tradução dependente de cap sem scanning, controlo da tradução por uORFs, função das proteínas de ligação ao RNA, elementos reguladores da tradução como alvos terapêuticos.N/

Endodontia: sessão única versus múltiplas sessões

Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Medicina DentáriaUm dos principais objetivos do tratamento Endodôntico é a tentativa de conseguir conduzir o sistema de canais radiculares a um ambiente de assepsia, sendo este conseguido através da eliminação dos microrganismos nocivos à cicatrização dos tecidos, permitindo assim a cura da infeção e posterior restabelecimento da função do dente. O tratamento endodôntico, seja preconizado numa ou em múltiplas sessões, visa a criação de condições ótimas para a obturação e selamento tridimensional dos canais radiculares. Assim, a seleção da modalidade de tratamento endodôntico terá de se basear, primeiramente, numa avaliação rigorosa do dente, propriamente dito, quanto à sua condição fisiopatológica pulpar e periapical, acessibilidade, caraterísticas morfológicas dos canais radiculares e restaurabilidade Uma das questões que mais se tem debatido é se o tratamento endodôntico numa sessão, possibilita que se atinjam tais condições, permitindo, portanto, alcançar os objetivos biológicos e mecânicos exigidos. Neste contexto, em que na atualidade paira a controvérsia e a incerteza sobre qual o procedimento mais indicado para a prática clínica, o objetivo deste trabalho foi, após efetuar uma revisão da literatura científica existente, tentar perceber se é mais indicado iniciar o tratamento e finaliza-lo na mesma sessão ou proceder a obturação na consulta seguinte. Hoje, devido aos avanços das técnicas de instrumentação e irrigação, que tem tornado o tratamento endodôntico mais eficaz num menor período de tempo, a sessão única é cada vez mais preconizada pelos profissionais. O sucesso será garantido se os procedimentos clínicos forem integralmente cumpridos e o tipo de modalidade escolhida resultar da reflexão das características de cada caso em particular. One of the main goals of Endodontic treatment is the attempt to achieve leading the root canal system to an aseptic environment, which is achieved through the elimination of the microorganisms that are harmful to the healing of tissues, allowing this way the healing of the infection and subsequent restoration of the tooth function. Endodontic treatment, whether recommended in one or in multiple sessions, aims to create optimal conditions for the tridimensional filling and sealing of the root canals. Thus, the selection of the type of endodontic treatment must be based, first, in a rigorous evaluation of the tooth itself, as far as its pathophysiological pulp and periapical condition, accessibility, morphological characteristics of the root canals and restorability are concerned. One of the most discussed issues is whether endodontic treatment in one session, enables one to get such conditions that allow the required mechanical and biological objectives to be achieved. In this context, in which nowadays hangs the controversy and the uncertainty about what the most suitable procedure for clinical practice is, the aim of this work, after carrying out a review of the existing scientific literature, was trying to understand whether it is more appropriate to start treatment and finish it in the same session or proceed with the filling in a second session. Today, due to the advances in the instrumentation and irrigation techniques, which have turned the endodontic treatment in a shorter period of time most effective, a Single Visit is increasingly recommended by health professionals. The success will be ensured if the clinical procedures are fully performed and if the type of modality chosen results of the reflection of the characteristics of each particular case

Higher education, stakeholders and collaborative work for entrepreneurial learning

Institutions of Higher Education can provide, in their internal environment, entrepreneurial learning opportunities for students stimulating new attitudes and behaviours towards entrepreneurship. The Portuguese universities have diversified their strategies to promote formal, informal and non formal entrepreneurial learning that can be encouraged in various strategies. The participation of key actors and stakeholders involved in awareness,mentoring and project implementation of the entrepreneurial learning process is crucial for collaborating work in a competitive world. Moreover, obtaining employment is increasingly dependent on not only the potentials of individuals, but also, and even more, the ability to build solid networks of partnerships in science and innovation and employment. Within the scope of the ongoing project "Entrepreneurial Learning, Cooperation and the Labour Market: Good Practice in Higher Education" this paper aims to highlight the importance of non formal and informal learning and to contribute to the reflection regarding added value of inter organizational cooperation and collaborative work

Human Argonaute 1 5’ untranslated region can mediate cap-independent translation initiation via an internal ribosome entry site

Argonaute proteins (AGOs) are essential effectors in RNA-mediated gene silencing pathways. There are eight AGO-like proteins in human cells, grouped in two families: the eIF2C/AGO subfamily and the PIWI subfamily. The eIF2C1 gene encodes AGO1, a member of the former subfamily, that is ubiquitously expressed at low to medium levels and it is highly conserved during evolution reflecting its important physiological roles. Moreover, recent studies concluded that AGO1 protein is overexpressed in colorectal cancer, relative to adjacent non-cancer tissue, without a concomitant increase in mRNA levels. These pieces of evidence lead us to suspect that high AGO1 protein levels may be due to internal ribosome entry site (IRES)-mediated translation. IRESs are structures that can mediate cap-independent translation initiation by directly recruiting ribosomes to the AUG vicinity, thus skipping the scanning of the whole 5’ untranslated region (UTR), in response to stress. To confirm this hypothesis, we transiently transfected colorectal cancer HCT116 and cervical cancer HeLa cells with an AGO1 5’UTR-containing dicistronic vector, and luciferase activity was measured by luminometry assays. Results have shown a 2-fold increase in relative luciferase activity in both cell lines, when compared to the cells transfected with the empty counterpart (P<0.05). Transfection of the corresponding promoterless plasmids ruled out the hypothesis of this fold to be due to the existence of a cryptic promoter. In addition, RT-PCR analyses of the dicistronic mRNAs confirmed that no cryptic splicing occurs. Besides, the knock-down of the eIF4E subunit induced a significant 2- to 4-fold increase in IRES activity. Taken together, these data suggest the presence of an IRES in the AGO1 5’UTR whose biological relevance is still under thorough investigation

Human UPF1: a cap-independent translation initiation mechanism and a cryptic promoter regulate its gene expression in cancer cells

Cervical (CC) and colorectal (CRC) cancers are among the leading causes of death worldwide and their development and progression is dependent on regulation of gene expression. Eukaryotic cells possess a variety of post-transcriptional mechanisms by which they regulate gene expression, namely at translation initiation level. Although in most cases it occurs via the cap-dependent mechanism, several oncogenes, growth factors or proteins involved in the regulation of programmed cell death, among others, have a different translation initiation mechanism. It involves the direct recruitment of the ribosome to the vicinity of the initiation codon without the involvement of the cap structure. This allows the maintenance of protein synthesis under several cellular stresses and promotes tumourigenesis. Apart from its role in nonsense-mediated decay, the human up-frameshift 1 (UPF1) DNA and RNA helicase protein plays a crucial role in telomere replication and homeostasis, and in cell cycle progression. In addition, its expression levels are maintained in every phase of the cell cycle, thus indicating that its translation may occur via a cap-independent mechanism. To test this hypothesis, we cloned the human UPF1 5’UTR in a dicistronic vector and transfected CC and CRC cell lines with either this construct or the control counterparts. We observed a 15- to 25-fold increase in relative luciferase activity of the UPF1 5’UTR-containing construct compared to the levels obtained from the empty counterpart in all tested cell lines, suggesting a cap-independent translation initiation. To control whether these levels of luciferase activity could be due to the presence of a cryptic promoter, we transfected cells with promoterless plasmids and observed the same result, demonstrating that UPF1 5’UTR contains a cryptic promoter. To check the cap-independent translation activity alone, we transfected cells with in vitro transcribed, capped and polyadenylated mRNAs and observed a 2-fold increase in protein levels, which shows that translation can occur in a cap-independent way. This is maintained under conditions of global protein synthesis inhibition. Deletional analysis of UPF1 5’UTR revealed that first 50 nucleotides are essential for both cryptic promoter and cap-independent activities. These results provide new insights on the regulation of UPF1 expression in human cancer cells

Endodontia: sessão única versus múltiplas sessões

Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Medicina DentáriaUm dos principais objetivos do tratamento Endodôntico é a tentativa de conseguir conduzir o sistema de canais radiculares a um ambiente de assepsia, sendo este conseguido através da eliminação dos microrganismos nocivos à cicatrização dos tecidos, permitindo assim a cura da infeção e posterior restabelecimento da função do dente. O tratamento endodôntico, seja preconizado numa ou em múltiplas sessões, visa a criação de condições ótimas para a obturação e selamento tridimensional dos canais radiculares. Assim, a seleção da modalidade de tratamento endodôntico terá de se basear, primeiramente, numa avaliação rigorosa do dente, propriamente dito, quanto à sua condição fisiopatológica pulpar e periapical, acessibilidade, caraterísticas morfológicas dos canais radiculares e restaurabilidade Uma das questões que mais se tem debatido é se o tratamento endodôntico numa sessão, possibilita que se atinjam tais condições, permitindo, portanto, alcançar os objetivos biológicos e mecânicos exigidos. Neste contexto, em que na atualidade paira a controvérsia e a incerteza sobre qual o procedimento mais indicado para a prática clínica, o objetivo deste trabalho foi, após efetuar uma revisão da literatura científica existente, tentar perceber se é mais indicado iniciar o tratamento e finaliza-lo na mesma sessão ou proceder a obturação na consulta seguinte. Hoje, devido aos avanços das técnicas de instrumentação e irrigação, que tem tornado o tratamento endodôntico mais eficaz num menor período de tempo, a sessão única é cada vez mais preconizada pelos profissionais. O sucesso será garantido se os procedimentos clínicos forem integralmente cumpridos e o tipo de modalidade escolhida resultar da reflexão das características de cada caso em particular. One of the main goals of Endodontic treatment is the attempt to achieve leading the root canal system to an aseptic environment, which is achieved through the elimination of the microorganisms that are harmful to the healing of tissues, allowing this way the healing of the infection and subsequent restoration of the tooth function. Endodontic treatment, whether recommended in one or in multiple sessions, aims to create optimal conditions for the tridimensional filling and sealing of the root canals. Thus, the selection of the type of endodontic treatment must be based, first, in a rigorous evaluation of the tooth itself, as far as its pathophysiological pulp and periapical condition, accessibility, morphological characteristics of the root canals and restorability are concerned. One of the most discussed issues is whether endodontic treatment in one session, enables one to get such conditions that allow the required mechanical and biological objectives to be achieved. In this context, in which nowadays hangs the controversy and the uncertainty about what the most suitable procedure for clinical practice is, the aim of this work, after carrying out a review of the existing scientific literature, was trying to understand whether it is more appropriate to start treatment and finish it in the same session or proceed with the filling in a second session. Today, due to the advances in the instrumentation and irrigation techniques, which have turned the endodontic treatment in a shorter period of time most effective, a Single Visit is increasingly recommended by health professionals. The success will be ensured if the clinical procedures are fully performed and if the type of modality chosen results of the reflection of the characteristics of each particular case

The expression of UPF1 is regulated by a cap-independent translation initiation mechanism and a cryptic promoter within its 5’ untranslated region

Regulation of gene expression occurs at several steps, including translation initiation. Under normal circumstances, translation initiation is mainly cap-dependent; however, several proteins can initiate translation in a cap-independent way, allowing the maintenance of these proteins under conditions that reduce global protein synthesis. hUPF1(human up-frameshift 1) plays important roles in several key cellular processes such as nonsense-mediated decay, telomere replication and homeostasis, and cell cycle progression, suggesting its expression must be tightly regulated to prevent abnormal cell proliferation. This protein is essential in the G2/M transition, a step known by a reduced overall protein synthesis. Taking these data into account, we hypothesized that UPF1 might initiate translation in a cap-independent way, allowing the cell to maintain its levels under conditions that impair cap-dependent translation initiation. To test this hypothesis, we cloned the hUPF1 5’UTR in a dicistronic vector and transfected CC and CRC cell lines with either this construct or the control counterparts. We observed a 15- to 25-fold increase in relative luciferase activity of the UPF1 5’UTR-containing construct compared to the levels obtained from the empty counterpart in all tested cell lines, suggesting a cap-independent translation initiation. To control whether luciferase activity levels are due to a cryptic promoter within UPF1 5’UTR, we transfected cells with promoterless plasmids and observed the same result, demonstrating that UPF1 5’UTR contains a cryptic promoter. Transfecting cells with in vitro transcribed mRNAs resulted in a 2-fold increase in protein levels, suggesting that translation can occur in a cap-independent way. This is maintained under conditions of global protein synthesis inhibition. Deletional analysis of UPF1 5’UTR revealed that the first 50 nucleotides are essential for cryptic promoter and cap-independent activities. These results provide new insights on the mechanisms that govern UPF1 expression

A motivação nos formadores

Atualmente, os Formadores e todos os Professsores em geral têm no seu contexto vários impedimentos, um deles de ordem emocional. Estes obstáculos podem resultar de um princípio natural ou apresentar-se como uma consequência. Afetando todos aqueles que com o Formador privam, comprometem as relações interpessoais e intrapessoais, refletindo-se desta forma na sua prática pedagógica e no grupo de formandos que o acompanha. Importa que a sociedade civil tome disto consciência e construa novos caminhos para o bem-estar emocional, desenvolvendo estratégias de autorregulação emocional na formação contínua do Formador e na procura de novas ferramentas de trabalho, com o objetivo de se contrariarem os constrangimentos que vão surgindo no exercício da profissão. Posto isto, evidencia-se a necessidade de áreas de competências da Inteligência Emocional nas entidades formativas, na medida em que a vida social e pessoal do formador se encontra interligada com as suas emoções e regulação. Precedeu este estudo de caso o objetivo de mostrar como o Formador, enquanto transmissor de um conhecimento e enquanto pessoa, se pode moldar consoante as emoções e sentimentos que vivencia nas instituições/entidades de formação. A metodologia baseou-se, essencialmente, em entrevistas a Formadores de várias áreas científicas e tecnológicas, previamente selecionados. Em seguida, procedeu-se a uma análise relevante dos dados, de forma a aferir os sentimentos/sensações e motivação interna/externa a que os formadores estão sujeitos e de que modo podem implicar no seu sucesso pessoal e profissional. Foi também realizada uma recolha paralela de informações/contributos que permitem demonstrar que as entidades/instituições formativas podem contribuir para o bem-estar emocional sentido pelos formadores e em todo o contexto de formação. Considerando os principais resultados obtidos, concluiu-se que o bem-estar emocional, provocado fundamentalmente pela forte persistência e motivação na profissão, decorre da operacionalização de algumas das habilidades provenientes da Inteligência Emocional pessoal desenvolvidas por cada um ao longo da sua experiência profissional.Actually, Formers and all Teachers in general have in their context several impediments, one of them is of emotional order. These obstacles may result from emotional dispositions. These obstacles may result from a natural principle or be presented as a consequence. They affect all the ones who deal directly with the Formers, they jeopardize the interpersonal and intrapersonal relations, reflecting themselves in the pedagogical practices as well as in the group of students who is with the Former. It is absolutely important that the civil society be conscious of it and make way to the emotional well being, developing some strategies of emotional autoregulation when forming, in a continuous way the Former, and when looking for new tools of work, with the main goal of contradict the constraints that emerge when practicing the profession. So, it is clear the need of Emotional Inteligence competences in the formative entities, as the social and personal Former’s life is connected to his/her emotions and regulations. In this case study, the main goal was to show the Former, as a knowledge transmitter and as a person, who can be changed according to the emotions and feelings that experiments in the forming institutions/entities. The methodology was based, mainly, in several areas Formers interviews, scientific and technological areas, previously selected. Then, the relevant analysis of all the data, as a way of getting all the feelings/sensations and internal/external motivations that formers are always exposed to and how they can manage his/her personal and professional success. It was also made a parallel compilation of informations/contributions, which allow demonstrate that forming entities/institutions can contribute to the emotional well-being felt by formers as in all forming context. Considering the obtained main results, it is concluded that the emotional well-being, caused mainly by the strong profession persistence and motivation, occurs from the operationalization that some skills proceed from the personal Emotional Inteligence developed by each one through his/her professional experience