title: Real-time imaging of bHLH transcription factors reveals their dynamic control in the multipotency and fate choice of neural stem cells.
abstract: The basic-helix-loop-helix (bHLH) transcription factors Ascl1/Mash1, Hes1, and Olig2 regulate the fate choice of neurons, astrocytes, and oligodendrocytes, respectively; however, these factors are coexpressed in self-renewing multipotent neural stem cells (NSCs) even before cell fate determination. This fact raises the possibility that these fate determination factors are differentially expressed between self-renewing and differentiating NSCs with unique expression dynamics. Real-time imaging analysis utilizing fluorescent proteins is a powerful strategy for monitoring expression dynamics. Fusion with fluorescent reporters makes it possible to analyze the dynamic behavior of specific proteins in living cells. However, it is technically challenging to conduct long-term imaging of proteins, particularly those with low expression levels, because a high-sensitivity and low-noise imaging system is required, and very often bleaching of fluorescent proteins and cell toxicity by prolonged laser exposure are problematic. Furthermore, to analyze the functional roles of the dynamic expression of cellular proteins, it is essential to image reporter fusion proteins that are expressed at comparable levels to their endogenous expression. In this review, we introduce our recent reports about the dynamic control of bHLH transcription factors in multipotency and fate choice of NSCs, focusing on real-time imaging of fluorescent reporters fused with bHLH transcription factors. Our imaging results indicate that bHLH transcription factors are expressed in an oscillatory manner by NSCs, and that one of them becomes dominant during fate choice. We propose that the multipotent state of NSCs correlates with the oscillatory expression of several bHLH transcription factors, whereas the differentiated state correlates with the sustained expression of a single bHLH transcription factor.
authors: The Hakubi Center, Kyoto University Kyoto, Japan ; Laboratory of Growth Regulation, Institute for Virus Research, Kyoto University Kyoto, Japan ; World Premier International Research Initiative-Institute for Integrated Cell-Material Sciences, Kyoto University Kyoto, Japan ; Precursory Research for Embryonic Science and Technology, Japan Science and Technology Agency Saitama, Japan.
authors: Itaru
authors: I
authors: Imayoshi
authors: World Premier International Research Initiative-Institute for Integrated Cell-Material Sciences, Kyoto University Kyoto, Japan.
authors: Fumiyoshi
authors: F
authors: Ishidate
authors: Laboratory of Growth Regulation, Institute for Virus Research, Kyoto University Kyoto, Japan ; World Premier International Research Initiative-Institute for Integrated Cell-Material Sciences, Kyoto University Kyoto, Japan ; Core Research for Evolutional Science and Technology, Japan Science and Technology Agency Saitama, Japan.
authors: Ryoichiro
authors: R
authors: Kageyama
keywords: bHLH; fluorescent protein; imaging; neural stem cells; oscillation