Research
Tracking genomic dynamics
The CRISPR-based technology platforms we have recently developed for labeling and tracking specific DNA loci in live cells (see references below), both as instantaneous dynamics and more cursive ranging during progression of cells through G1, S and G2, now sets the stage for extended studies including the degree to which gene activation or silencing affects the dynamic range of these chromosomal sites (or conversely, may determine gene activation or silencing in the first place).
Multicolor CRISPR labeling of chromosomal loci in human cells.
Ma H, Naseri A, Reyes-Gutierrez P, Wolfe SA, Zhang S, Pederson T.
Proc. Natl. Acad. Sci. USA 2015. 112:3002-3007. doi: 10.1073/pnas.1420024112.
CRISPR-Cas9 nuclear dynamics and target recognition in living cells.
Ma H, Tu LC, Naseri A, Huisman M, Zhang S, Grunwald D, Pederson T.
J. Cell Biol. 2016. 214:529-537. doi: 10.1083/jcb.201604115.
Multiplexed labeling of genomic loci with dCas9 and engineered sgRNAs using CRISPRainbow.
Ma H, Tu LC, Naseri A, Huisman M, Zhang S, Grunwald D, Pederson T.
Nat. Biotechnol. 2016. 34:528-530. doi: 10.1038/nbt.3526
CRISPR-Based DNA Imaging in Living Cells Reveals Cell Cycle-Dependent Chromosome Dynamics
Hanhui Ma, Li-Chun Tu, Ardalan Naseri, Yu-Chieh Chung, David Grunwald, Shaojie Zhang, Thoru Pederson. bioRxiv 195966; doi: https://doi.org/10.1101/195966
The nucleolus as a staging center for microRNAs and messenger RNAs
Our discovery that subsets of microRNAs and messenger RNAs are localized in nucleoli, and that these have base-pairing complementarity (Reyes-Gutierrez et al., 2014) has led us to postulate that this nuclear body, the nucleolus, is a staging center for the assembly of certain (but not all) microRNA:mRNA complexes prior to their export to the cytoplasm in a “pre-regulated” state. This intriguing idea suggests several experimental approaches for confirmation.
A mRNA and cognate microRNAs localize in the nucleolus.
Reyes-Gutierrez P, Ritland Politz JC, Pederson T.
Nucleus. 2014. 5:636-642. doi: 10.4161/19491034.2014.990864.
Sensing the shape of DNA in live cells
Our extensive studies with CRISPR-based labeling of genomic DNA (vide supra) have given us reason to ponder if the cellular DNA surveillance machinery can sense changes in DNA helix shape. We of course use nuclease-dead Cas9 in our labeling platform so there is no DNA cleavage, and yet the DNA helix is distorted. We are working with a collaborating laboratory to pursue the question of whether the eukaryotic cell’s genome surveillance systems can detect the DNA shape changes imposed by this archae-/eubacterial “visitor”.