Bleris, Leonidas
Permanent URI for this collectionhttps://hdl.handle.net/10735.1/2891
Leonidas Bleris is interested in the intersection of synthetic and systems biology, mathematics and control theory. He is the head of the Synthetic & Systems Biology Research Lab. Learn more about Dr. Bleris's research on his Research Explorer page
Browse
Browsing Bleris, Leonidas by Subject "MicroRNAs"
Now showing 1 - 4 of 4
- Results Per Page
- Sort Options
Item Mapping the Operational Landscape of MicroRNAs in Synthetic Gene Circuits.(Nature Partner Journals, 2018-10-22) Quarton, Tyler; Ehrhardt, Kristina; Lee, James; Kannan, Srijaa; Li, Yi; Ma, Lan; Bleris, Leonidas; Quarton, Tyler; Ehrhardt, Kristina; Lee, James; Kannan, Srijaa; Li, Yi; Ma, Lan; Bleris, LeonidasMicroRNAs are a class of short, noncoding RNAs that are ubiquitous modulators of gene expression, with roles in development, homeostasis, and disease. Engineered microRNAs are now frequently used as regulatory modules in synthetic biology. Moreover, synthetic gene circuits equipped with engineered microRNA targets with perfect complementarity to endogenous microRNAs establish an interface with the endogenous milieu at the single-cell level. The function of engineered microRNAs and sensor systems is typically optimized through extensive trial-and-error. Here, using a combination of synthetic biology experimentation in human embryonic kidney cells and quantitative analysis, we investigate the relationship between input genetic template abundance, microRNA concentration, and output under microRNA control. We provide a framework that employs the complete operational landscape of a synthetic gene circuit and enables the stepwise development of mathematical models. We derive a phenomenological model that recapitulates experimentally observed nonlinearities and contains features that provide insight into the microRNA function at various abundances. Our work facilitates the characterization and engineering of multi-component genetic circuits and specifically points to new insights on the operation of microRNAs as mediators of endogenous information and regulators of gene expression in synthetic biology.Item Mir-192-Mediated Positive Feedback Loop Controls the Robustness of Stress-Induced P53 Oscillations in Breast Cancer Cells(Public Library of Science) Moore, Richard; Ooi, Hsu Kiang; Kang, Taek; Bleris, Leonidas; Ma, Lan; 55515673900 (Ma, L); Moore, Richard; Ooi, Hsu Kiang; Kang, Taek; Bleris, Leonidas; Ma, LanThe p53 tumor suppressor protein plays a critical role in cellular stress and cancer prevention. A number of post-transcriptional regulators, termed microRNAs, are closely connected with the p53-mediated cellular networks. While the molecular interactions among p53 and microRNAs have emerged, a systems-level understanding of the regulatory mechanism and the role of microRNAs-forming feedback loops with the p53 core remains elusive. Here we have identified from literature that there exist three classes of microRNA-mediated feedback loops revolving around p53, all with the nature of positive feedback coincidentally. To explore the relationship between the cellular performance of p53 with the microRNA feedback pathways, we developed a mathematical model of the core p53-MDM2 module coupled with three microRNA-mediated positive feedback loops involving miR-192, miR-34a, and miR-29a. Simulations and bifurcation analysis in relationship to extrinsic noise reproduce the oscillatory behavior of p53 under DNA damage in single cells, and notably show that specific microRNA abrogation can disrupt the wild-type cellular phenotype when the ubiquitous cell-to-cell variability is taken into account. To assess these in silico results we conducted microRNA-perturbation experiments in MCF7 breast cancer cells. Time-lapse microscopy of cell-population behavior in response to DNA double-strand breaks, together with image classification of single-cell phenotypes across a population, confirmed that the cellular p53 oscillations are compromised after miR-192 perturbations, matching well with the model predictions. Our study via modeling in combination with quantitative experiments provides new evidence on the role of microRNA-mediated positive feedback loops in conferring robustness to the system performance of stress-induced response of p53.;Item Transcription Activator-Like Effector Hybrids for Conditional Control and Rewiring of Chromosomal Transgene ExpressionLi, Yi; Moore, Richard; Guinn, Michael; Bleris, Leonidas; 0000 0001 2535 9739 (Bleris, L); 2012076942 (Bleris, L); Erik Jonsson School of Engineering and Computer Science. Center for Systems Biology.The ability to conditionally rewire pathways in human cells holds great therapeutic potential. Transcription activator-like effectors (TALEs) are a class of naturally occurring specific DNA binding proteins that can be used to introduce targeted genome modifications or control gene expression. Here we present TALE hybrids engineered to respond to endogenous signals and capable of controlling transgenes by applying a predetermined and tunable action at the single-cell level. Specifically, we first demonstrate that combinations of TALEs can be used to modulate the expression of stably integrated genes in kidney cells. We then introduce a general purpose two-hybrid approach that can be customized to regulate the function of any TALE either using effector molecules or a heterodimerization reaction. Finally, we demonstrate the successful interface of TALEs to specific endogenous signals, namely hypoxia signaling and microRNAs, essentially closing the loop between cellular information and chromosomal transgene expression. © 2012 Macmillan Publishers Limited. All rights reserved.Item Transcripts for Combined Synthetic MicroRNA and Gene Delivery(2013-06-26) Kashyap, Neha; Pham, Bich; Xie, Zhen; Bleris, Leonidas; 0000 0001 2535 9739 (Bleris, L)MicroRNAs (miRNAs) are a class of short noncoding RNAs which are endogenously expressed in many organisms and regulate gene expression by binding to messenger RNA (mRNA). MicroRNAs are either produced from their independent transcription units in intergenic regions or lie in intragenic regions. Intragenic miRNAs and their host mRNAs are produced from the same transcript by the microprocessor and the spliceosome protein complex respectively. The details and exact timing of the processing events have implications for downstream RNA interference (RNAi) efficiency and mRNA stability. Here we engineer and study in mammalian cells a range of synthetic intragenic miRNAs co-expressed with their host genes. Furthermore, we study transcripts which carry the target of the miRNA, thereby emulating a common regulation mechanism. We perform fluorescence microscopy and flow cytometry to characterize the engineered transcripts and investigate the properties of the underlying biological processes. Our results shed additional light on miRNA and pre-mRNA processing but importantly provide insight into engineering transcripts customized for combined delivery and use in synthetic gene circuits.;