RRC ID 69230
Author Kaida A, Miura M.
Title Differential dependence on oxygen tension during the maturation process between monomeric Kusabira Orange 2 and monomeric Azami Green expressed in HeLa cells.
Journal Biochem Biophys Res Commun
Abstract Although oxygen is required for functional chromophore formation during the maturation process of fluorescent proteins, the effects of hypoxia on their fluorescence have rarely been studied in mammalian cells. We recently reported that severe hypoxia (pO(2)<0.1%) abrogates fluorescence from the fluorescent ubiquitination-based cell cycle indicator (Fucci) expressed in HeLa cells. Fucci is a system for visualizing cell cycle progression in live cells using red (monomeric Kusabira Orange 2, mKO(2)) and green (monomeric Azami Green, mAG) fluorescent proteins. In this study, taking advantage of the system, we attempted to determine the dependence on oxygen tension (pO(2)) of these two fluorescent proteins during the maturation process. The oxygen tension at which the number of fluorescence-positive cells was reduced by 50% (pO(2)·50) was 0.9% and 0.3% for mKO2 and mAG, respectively. Furthermore, we measured fluorescence recovery kinetics after reoxygenation in cells treated at two different pO(2) levels, and observed that mKO2 exhibits slower kinetics of oxidation than mAG. Thus, we demonstrate that mKO2 exhibits a stronger dependence on oxygen tension than mAG, as well as the usefulness of this novel method to produce varying levels of hypoxic conditions.
Volume 421(4)
Pages 855-9
Published 2012-5-18
DOI 10.1016/j.bbrc.2012.04.102
PII S0006-291X(12)00784-X
PMID 22554525
MeSH Cell Hypoxia Fluorescence Green Fluorescent Proteins / chemistry Green Fluorescent Proteins / metabolism* HeLa Cells Humans Kinetics Luminescent Proteins / chemistry Luminescent Proteins / metabolism* Oxygen / chemistry Oxygen / metabolism*
IF 2.985
Human and Animal Cells HeLa/Fucci(RCB2812)