Characterization and mitigation of gene expression burden in mammalian cells

Despite recent advances in circuit engineering, the design of genetic networks in mammalian cells is still painstakingly slow and fraught with inexplicable failures. Here, we demonstrate that transiently expressed genes in mammalian cells compete for limited transcriptional and translational resources. This competition results in the coupling of otherwise independent exogenous and endogenous genes, creating a divergence between intended and actual function. Guided by a resource-aware mathematical model, we identify and engineer natural and synthetic miRNA-based incoherent feedforward loop (iFFL) circuits that mitigate gene expression burden. The implementation of these circuits features the use of endogenous miRNAs as elementary components of the engineered iFFL device, a versatile hybrid design that allows burden mitigation to be achieved across different cell-lines with minimal resource requirements. This study establishes the foundations for context-aware prediction and improvement of in vivo synthetic circuit performance, paving the way towards more rational synthetic construct design in mammalian cells

Homocysteine and Coronary Heart Disease: Meta-analysis of MTHFR Case-Control Studies, Avoiding Publication Bias

Robert Clarke and colleagues conduct a meta-analysis of unpublished datasets to examine the causal relationship between elevation of homocysteine levels in the blood and the risk of coronary heart disease. Their data suggest that an increase in homocysteine levels is not likely to result in an increase in risk of coronary heart disease

MiRNA-29a regulates the expression of numerous proteins and reduces the invasiveness and proliferation of human carcinoma cell lines

In this study we have identified a functional role for miR-29a in cancer cell invasion and proliferation. MiRNA expression profiling of human NSCLC cell lines indicated that miR- 29a levels were reduced in more invasive cell lines. Exogenous overexpression of miR-29a in both lung and pancreatic cancer cell lines resulted in a significant reduction in the invasion phenotype, as well as in proliferation. 2D DIGE proteomic profiling of cells transfected with pre-miR-29a or anti-miR-29a resulted in the identification of over 100 differentially regulated proteins. The fold change of protein expression was generally modest – in the range 1.2–1.7-fold. Only 14 were predicted computationally to have miR-29a seed sequences in their 30 UTR region. Subsequent studies using siRNA to knock down several candidate proteins from the 2D DIGE experiment identified RAN (a member of the RAS oncogene family) which significantly reduced the invasive capability of a model lung cancer cell line. We conclude that miR-29a has a significant anti-invasive and anti-proliferative effect on lung cancer cells in vitro and functions as an anti-oncomir. This function is likely mediated through the post-transcriptional fine tuning of the cellular levels of several proteins, both directly and indirectly, and in particular we provide some evidence that RAN represents one of these