Blocking IRES-mediated translation pathway as a new method to treat Alzheimer’s disease

AbstractScientists theorized that β-amyloid (Aβ) plaques and tau tangles are involved in the development of Alzheimer’s disease (AD), and amyloid precursor protein (APP) produces Aβ to trigger the disease process. However, the normal synaptic function of APP itself is not fully understood. Several findings cast APP as a potential key player in learning and memory under normal condition. Nevertheless, the regular operation of APP will be disrupted by abnormal accumulation of Aβ under cellular pathological conditions. Herein, there is a hypothesis that AD could be treated by attenuating APP synthesis during cellular pathophysiological stress. In virtue of a previous study, it was speculated that cells could not decrease APP synthesis via self-protection maybe because APP is synthesized via internal ribosome entry segment (IRES)-mediated translation. Consequently, the blockage of this translation might be a new inoffensive and high-level specificity treatment

Replication Origins and Timing of Temporal Replication in Budding Yeast: How to Solve the Conundrum?

Similarly to metazoans, the budding yeast Saccharomyces cereviasiae replicates its genome with a defined timing. In this organism, well-defined, site-specific origins, are efficient and fire in almost every round of DNA replication. However, this strategy is neither conserved in the fission yeast Saccharomyces pombe, nor in Xenopus or Drosophila embryos, nor in higher eukaryotes, in which DNA replication initiates asynchronously throughout S phase at random sites. Temporal and spatial controls can contribute to the timing of replication such as Cdk activity, origin localization, epigenetic status or gene expression. However, a debate is going on to answer the question how individual origins are selected to fire in budding yeast. Two opposing theories were proposed: the “replicon paradigm” or “temporal program” vs. the “stochastic firing”. Recent data support the temporal regulation of origin activation, clustering origins into temporal blocks of early and late replication. Contrarily, strong evidences suggest that stochastic processes acting on origins can generate the observed kinetics of replication without requiring a temporal order. In mammalian cells, a spatiotemporal model that accounts for a partially deterministic and partially stochastic order of DNA replication has been proposed. Is this strategy the solution to reconcile the conundrum of having both organized replication timing and stochastic origin firing also for budding yeast? In this review we discuss this possibility in the light of our recent study on the origin activation, suggesting that there might be a stochastic component in the temporal activation of the replication origins, especially under perturbed conditions

Lost in Transcription: Molecular Mechanisms that Control HIV Latency

Peer reviewe

Characterizing the Associations and Roles of DDK and Mcm2-7 DNA Replication Proteins in Saccharomyces Cerevisiae

The essential cell cycle kinase Dbf4/Cdc7 (DDK) triggers DNA replication through phosphorylation of the Mcm2-7 helicase at replication origins. Prior work has implicated various Mcm2-7 subunits as targets of DDK, however it is not well understood which specific subunits mediate the docking of the DDK complex. Through yeast two-hybrid and co-immunoprecipitation analyses, we found that Dbf4 and Cdc7 interact with distinct subunits of the Mcm2-7 helicase complex. Dbf4 showed the strongest interaction with Mcm2 while Cdc7 associated with Mcm4 and Mcm5. Dissection of the N-terminal region of Mcm2 revealed two regions that mediate the interaction with Dbf4, whereas in Mcm4, a region near the N-terminus has been previously identified by another group as the DDK docking domain. Mutant forms of Mcm2 (Mcm2ΔDDD) or Mcm4 (Mcm4ΔDDD) lacking the DDK docking domain were expressed in cells and resulted in modest growth and replication defects. Combining the two mutations resulted in synthetic lethality, suggesting a redundant mechanism of Mcm2 and Mcm4 in targeting the DDK complex to Mcm rings. Furthermore, growth inhibition could be induced in a Mcm4ΔDDD background by overexpressing Mcm2 to titrate Dbf4 from Mcm rings. These growth defects were exacerbated in the presence of genotoxic agents such as hydroxyurea and methyl methanesulfonate, suggesting that DDK-Mcm interactions may play a role in stabilizing replication forks under S-phase checkpoint conditions. Regions of Cdc7 were examined for their interaction with Mcm4 and Dbf4. Results have shown that the N-terminal amino acid region 55-124 and the C-terminal region 453-507 of Cdc7 are likely target regions for Dbf4-binding. Several conserved residues were identified within the N-terminal 55-124 Cdc7 region that interface with conserved residues within motif-C of Dbf4. Conserved residues were identified within the DDD domain of Mcm2 and mutating these residues resulted in a decreased interaction with Dbf4. Lastly, bioinformatics analysis has revealed potential conserved residues within the Mcm4DDD region, which may play a role in binding to Cdc7. This research is significant because these factors, which are conserved in all eukaryotes studied to date, should give further insight as to how DNA replication is triggered and how it is affected when cells are exposed to DNA damaging or replication compromising agents. This research also has implications in cancer genetics, as prior studies have shown elevated DDK and Mcm protein levels in tumour cell lines and melanomas, with Cdc7 showing great promise as a cancer therapeutic target. Such knowledge will further enhance our understanding of the DNA replication process and the roles of cell cycle proteins involved, under both normal and checkpoint conditions

Investigating signalling pathways that influence stem cell self-renewal and differentiation

Adult tissue homeostasis relies on stem cells dividing to provide cells that differentiate and replenish lost cells. To prevent depletion of the stem cell pool, some of the daughter cells resulting from stem cell divisions retain stem cell identity and continue to proliferate, or self-renew. How stem cell self-renewal and differentiation are balanced is poorly understood. The niche, in which stem cells reside, provides spatially restricted signals that promote self-renewal. Daughter cells displaced away from the niche are thought to differentiate upon losing access to such signals. The sequence of events that occurs leading up to and following stem cell division is not well-understood. The Drosophila testis is a well-characterised model to study these behaviours. Here, cyst stem cells (CySCs) give rise to cyst cells that support germline development. Previous work in fixed tissues has shown that dysregulating CySC signalling can bias CySC fate outcomes. It remains to be demonstrated that, under normal physiological conditions, endogenous differences in signalling pathway activity are responsible for stem cell fate. In this thesis, I present evidence that differentiation in the Drosophila testis is not a default state upon losing access to niche-derived signals but is actively induced by signalling, and that the germline also plays a role in CySC differentiation. To visualise signalling dynamics in vivo, we have adapted a kinase activity biosensor from mammalian cell culture. I demonstrate that this biosensor faithfully reports kinase activity in both larval and adult tissues, and that it can be implemented with live imaging to study real-time signalling dynamics in individual cells. Finally, I characterise CySC behaviours under normal physiological circumstances using time-lapse live imaging of adult Drosophila testis explants. Based on these data, I discuss the role that signalling pathways play in maintaining the balance between self-renewal and differentiation of stem cells during normal homeostasis

Targeting Polo-like Kinase 1 in Glioma Propagating Cells

Master'sMASTER OF SCIENC

Mitochondrial function in the evolutionary origin of the female germ line

PhDOxidative phosphorylation couples ATP synthesis to respiratory electron transport. This coupling occurs in mitochondria, which carry DNA. Respiratory electron transport in the presence of molecular oxygen generates mutagenic reactive oxygen species (ROS) at a frequency that is itself increased by mutation. Damage to mitochondrial DNA (mtDNA) therefore accumulates within the lifespan of individual organisms. Syngamy requires motility of one gamete, and this motility requires ATP. It has been proposed that that oxidative phosphorylation is absent in the special case of quiescent, template mitochondria, and that these remain sequestered in oocytes and female germ lines. Oocyte mtDNA is thus protected from damage. Here I present evidence that female gametes, which are immotile, repress mitochondrial DNA transcription, mitochondrial membrane potential (!!m), and ROS production. In contrast, somatic cells and male gametes are seen actively to transcribe mitochondrial genes for respiratory electron carriers, and to produce ROS. I find that this functional division of labour between sperm and egg is widely distributed within the animal kingdom, and characterised by contrasting mitochondrial size and morphology. If quiescent oocyte mitochondria alone retain the capacity for an indefinite number of accurate replications of mtDNA, then "female" can be defined as that sex which transmits genetic template mitochondria. Template mitochondria then give rise to mitochondria that perform oxidative phosphorylation in somatic cells and in male gametes of each new generation. Template mitochondria also persist within the female germ line, to populate the oocytes of daughters. Thus mitochondria are maternally inherited