RT Journal Article
SR Electronic
T1 An optogenetic method for interrogating YAP1 and TAZ nuclear-cytoplasmic shuttling
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 2020.06.08.140228
DO 10.1101/2020.06.08.140228
A1 Anna M. Dowbaj
A1 Robert P. Jenkins
A1 Klaus Hahn
A1 Marco Montagner
A1 Erik Sahai
YR 2020
UL http://biorxiv.org/content/early/2020/06/09/2020.06.08.140228.abstract
AB The shuttling of transcription factors and transcriptional regulators in and out of the nucleus is central to the regulation of many biological processes. Here we describe a new method for studying the rates of nuclear entry and exit of transcriptional regulators. A photo-responsive AsLOV (Avena sativa Light Oxygen Voltage) domain is used to sequester the transcriptional regulators, YAP1 and TAZ/WWTR1, on the surface of mitochondria. Illumination with blue light is used to release fluorophore-tagged YAP1 and TAZ from mitochondria and their entry into the nucleus can be observed. Cessation of blue light illumination leads to re-sequestration on the surface of mitochondria. This method generates three distinct curves that are then used to fit ordinary differential equations for the rates of nuclear entry and exit. Using this method, we demonstrate that phosphorylation of YAP1 on sites of canonical regulation by LATS1/2 enhances its rate of nuclear export. Moreover, we provide evidences that YAP1 import and export rates, despite high intercellular variability, are correlated within the same cell. Interestingly, the ratio of import and export rates correlated with nuclear-cytoplasmic (NC) distribution of YAP1 and TAZ proteins at steady state.Competing Interest StatementThe authors have declared no competing interest.