Lacerda, RafaelaMenezes, JulianeMarques-Ramos, AnaTeixeira, AlexandreRomão, Luísa2016-03-042020-12-312015-11-06http://hdl.handle.net/10400.18/3658Gene expression is a very intricate process comprising several tightly regulated steps. One of those is translation initiation that, under normal circumstances, is mostly cap-dependent. However, some proteins can initiate translation via a cap-independent mechanism. This allows the maintenance of protein synthesis under conditions that reduce global protein synthesis. Human up-frameshift 1 (UPF1) has a key role in several cellular processes such as nonsense-mediated mRNA decay, telomere replication and homeostasis, and cell cycle progression, suggesting a tight regulation in order to prevent abnormal proliferation. These data suggest UPF1 might initiate translation in a cap-independent way, allowing the cell to overcome stress conditions that impair cap-dependent translation. To test this hypothesis, we cloned the UPF1 5’UTR in a dicistronic vector and transfected cervical and colorectal cancer 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. Cells transfection with in vitro transcribed mRNAs resulted in a 2-fold increase in protein levels, confirming translation can occur in a cap-independent way. This is maintained under conditions of global protein synthesis inhibition. Deletional analysis of the UPF1 5’UTR revealed that the minimal core required for cap-independent activity is present either within the first 100 nucleotides or within the last 125. Further experiments are being undertaken to understand the biological role of a cap-independent mechanism for the translation of UPF1 and how it contributes to the roles UPF1 plays in the cell.engGenómica Funcional e EstruturalExpressão GénicaSíntese Proteicaconference object