¹ Department of Mechanical and Aerospace Engineering, University of California, Los Angeles, CA 90095, US; ² Center for Cell Control, University of California, Los Angeles, CA 90095; US ³ Current address: Institute of Life Sciences, Fuzhou University, Fuzhou 350108, China; ⁴ Department of Ophthalmology and Doheny Eye Institute, University of Southern California, Los Angeles, CA 90033, US

The different cell types in an animal are often considered to be specified by combinations of transcription factors, and defined by marker gene expression. This paradigm is challenged, however, in stem cell research and application. Using a mouse embryonic stem cell (mESC) culture system, here we show that the expression level of many key stem cell marker genes/transcription factors such as Oct4, Sox2 and Nanog failed to monitor cell status transition during mESC differentiation. On the other hand, the response patterns of cell signalling network to external stimuli, as monitored by the dynamics of protein phosphorylation, changed dramatically. Our results also suggest that an irreversible alternation in cell signalling network precedes the adjustment of transcription factor levels. This is consistent with the notion that signal transduction events regulate cell fate specification. We propose that interrogating cell signalling network can assess the cell property more precisely, and provide a sensitive measurement for the early events in cell fate transition. We wish to bring up attention to the potential problem of cell identification using a few marker genes, and suggest a novel methodology to address this issue.