scholarly journals COMET: A toolkit for composing customizable genetic programs in mammalian cells

2019 ◽  
Author(s):  
Patrick S. Donahue ◽  
Joseph W. Draut ◽  
Joseph J. Muldoon ◽  
Hailey I. Edelstein ◽  
Neda Bagheri ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mathematical Model ◽  
Transcription Factors ◽  
Zinc Finger ◽  
Mammalian Cells ◽  
Single Layer ◽  
Performance Characteristics ◽  
Boolean Logic ◽  
Design And Construction

ABSTRACTEngineering mammalian cells to carry out sophisticated and customizable genetic programs requires a toolkit of multiple orthogonal and well-characterized transcription factors (TFs). To address this need, we developed the COmposable Mammalian Elements of Transcription (COMET)—an ensemble of TFs and promoters that enable the design and tuning of gene expression to an extent not previously possible. COMET currently comprises 44 activating and 12 inhibitory zinc-finger TFs and 83 cognate promoters, combined in a framework that readily accommodates new parts. This system can tune gene expression over three orders of magnitude, provides chemically inducible control of TF activity, and enables single-layer Boolean logic. We also develop a mathematical model that provides mechanistic insights into COMET performance characteristics. Altogether, COMET enables the design and construction of customizable genetic programs in mammalian cells.

scholarly journals Exploring the Changing Landscape of Cell-to-Cell Variation After CTCF Knockdown via Single Cell RNA-seq

2019 ◽  
Author(s):  
Wei Wang ◽  
Gang Ren ◽  
Ni Hong ◽  
Wenfei Jin
Keyword(s):  
Gene Expression ◽  
Transcription Factors ◽  
Single Cell ◽  
Zinc Finger ◽  
Ctcf Binding ◽  
Rna Seq ◽  
Expression Noise ◽  
Genome Wide ◽  
Cell Variation ◽  
Variable Genes

Abstract Background: CCCTC-Binding Factor (CTCF), also known as 11-zinc finger protein, participates in many cellular processes, including insulator activity, transcriptional regulation and organization of chromatin architecture. Based on single cell flow cytometry and single cell RNA-FISH analyses, our previous study showed that deletion of CTCF binding site led to a significantly increase of cellular variation of its target gene. However, the effect of CTCF on genome-wide landscape of cell-to-cell variation is unclear. Results: We knocked down CTCF in EL4 cells using shRNA, and conducted single cell RNA-seq on both wild type (WT) cells and CTCF-Knockdown (CTCF-KD) cells using Fluidigm C1 system. Principal component analysis of single cell RNA-seq data showed that WT and CTCF-KD cells concentrated in two different clusters on PC1, indicating gene expression profiles of WT and CTCF-KD cells were systematically different. Interestingly, GO terms including regulation of transcription, DNA binding, Zinc finger and transcription factor binding were significantly enriched in CTCF-KD-specific highly variable genes, indicating tissue-specific genes such as transcription factors were highly sensitive to CTCF level. The dysregulation of transcription factors potentially explain why knockdown of CTCF lead to systematic change of gene expression. In contrast, housekeeping genes such as rRNA processing, DNA repair and tRNA processing were significantly enriched in WT-specific highly variable genes, potentially due to a higher cellular variation of cell activity in WT cells compared to CTCF-KD cells. We further found cellular variation-increased genes were significantly enriched in down-regulated genes, indicating CTCF knockdown simultaneously reduced the expression levels and increased the expression noise of its regulated genes. Conclusions: To our knowledge, this is the first attempt to explore genome-wide landscape of cellular variation after CTCF knockdown. Our study not only advances our understanding of CTCF function in maintaining gene expression and reducing expression noise, but also provides a framework for examining gene function.

scholarly journals A tunable zinc finger-based framework for Boolean logic computation in mammalian cells

2012 ◽  
Vol 40 (11) ◽  
pp. 5180-5187 ◽  
Author(s):  
Jason J. Lohmueller ◽  
Thomas Z. Armel ◽  
Pamela A. Silver
Keyword(s):  
Zinc Finger ◽  
Mammalian Cells ◽  
Boolean Logic ◽  
Logic Computation

Identification and Use of Zinc Finger Transcription Factors That Increase Production of Recombinant Proteins in Yeast and Mammalian Cells

2008 ◽  
Vol 21 (3) ◽  
pp. 664-670 ◽  
Author(s):  
Kyung-Soon Park ◽  
Wongi Seol ◽  
Hyo-Young Yang ◽  
Seong-Il Lee ◽  
Sung Keun Kim ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Zinc Finger ◽  
Recombinant Proteins ◽  
Mammalian Cells

scholarly journals Zinc Finger Transcription Factors as Molecular Targets for Nitric Oxide-mediated Immunosuppression: Inhibition of IL-2 Gene Expression in Murine Lymphocytes

1999 ◽  
Vol 5 (11) ◽  
pp. 721-730 ◽  
Author(s):  
Denise Berendji ◽  
Victoria Kolb-Bachofen ◽  
Peter F. Zipfel ◽  
Christine Skerka ◽  
Carsten Carlberg ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nitric Oxide ◽  
Transcription Factors ◽  
Zinc Finger ◽  
Molecular Targets
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