Experimental and Computational Tools to Study P53 Dynamics at the Single-Cell Level

Abstract

One of the most commonly mutated genes found in cancer is the tumor suppressor p53. p53 is a transcription factor capable of inducing cell-cycle arrest, apoptosis, senescence, and other cellular processes thought to halt the progression of a nascent cancer. As part of a stress signaling pathway, p53 is acutely activated by ionizing radiation and the formation of DNA double-strand breaks. The appearence of this DNA damage causes the concentration of p53 within the nucleus to fluctuate and pulse regularly, which can be observed in single cells using fluorescence time-lapse microscopy. From the time this was first discovered, the connection between these p53 dynamics and p53 function has been speculated upon. A key insight into this connection came from a Lahav Lab publication that demonstrated the act of pulsing, itself, controls p53-dependent transcription and cell fate. The mechanisms and molecular details behind this relationship are now an area of intense study. Another area of high interest is the broader characterization of p53 dynamics in different time-scales, genetic backgrounds, and stresses. These lines of research each depend upon single-cell measurements that are often time consuming, noisy, and yield small sample sizes. The ongoing development of experiemental and computational tools for single-cell biology is needed to overcome these limitations. In the publication referenced earlier, a novel method was created to measure p53 dynamics and gene expression in the same cell. In a seperate study characterizing p53 dynamics over long time-scales, semi-automated tracking software aided in the discovery of new p53 dynamics: sustained elevation of p53 levels that follow a period of pulsing. Population measurements showing similarly elevated p53 levels on the same time-scale are shown to depend on the late induction of the p53-target PIDD.