Abstract
Technological advances are continually improving our ability to obtain more accurate views about the inner workings of biological systems. One such rapidly evolving area is single cell biology, and in particular gene expression and its regulation by transcription factors in response to intrinsic and extrinsic factors. Regarding the study of transcription factors, we discuss some of the promises and pitfalls associated with investigating how individual cells regulate gene expression through modulation of transcription factor activities. Specifically, we discuss four leading experimental approaches, the data that can be obtained from each, and important considerations that investigators should be aware of when drawing conclusions from such data.
Highlights
To perform essential biological functions and respond to changes in their microenvironments, cells systematically change the expression levels of process-relevant genes, while maintaining homeostatic expression of so-called housekeeping genes
transcription factors (TFs) are thought to determine which genes are expressed, and influence the degree and timing of gene expression. Their temporal activity patterns can be unexpectedly complex [4,5,6,7,8,9,10,11], which raises questions about their functional significance, and complicates efforts to understand the role of TF dynamics in terms of gene regulation
A recent study combined imaging TF dynamics with microfluidic immunoassay of single cell protein secretion to study the impact of TF dynamics on single cell gene expression, and reported that nuclear factor κB (NF-κB) dynamics did not correlate with the expression of TNF-α protein [28]
Summary
To perform essential biological functions and respond to changes in their microenvironments, cells systematically change the expression levels of process-relevant genes, while maintaining homeostatic expression of so-called housekeeping genes. TFs are thought to determine which genes are expressed, and influence the degree and timing of gene expression. Their temporal activity patterns (hereafter referred to as TF dynamics) can be unexpectedly complex [4,5,6,7,8,9,10,11], which raises questions about their functional significance, and complicates efforts to understand the role of TF dynamics in terms of gene regulation. In investigations of TF dynamics in single cells, TFs are often expressed as fluorescent fusion proteins and imaged via live-cell fluorescence microscopy.
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