Regulation of protein abundance in pluripotent cells undergoing commitment to the neural lineage

Abstract

The P19 cell line is a widely studied model of neural differentiation When pluripotent P19 cells are cultured as aggregates in the presence of retinoic acid for 4 days, the cells commit to the neural fate, but have not yet undergone overt differentiation. Two-dimensional polyacrylamide gel electrophoresis was used to analyze cellular protein expression during this induction. Approximately 500 abundant polypeptides were analyzed. Seventeen polypeptides were upregulated during induction; several of these were significantly regulated 48 h after the addition of retinoic acid. No downregulations were observed. Fifteen of the 17 polypeptides continued to be expressed throughout terminal differentiation. The upregulation of 14 of the 17 polypeptides requires both retinoic acid and aggregation, which alone do not induce neural differentiation. Furthermore, these regulated polypeptides are expressed in neural tissue, suggesting they are associated with neural function in vivo. Embryonic stem cells, a totipotent line, also neurally differentiate in response to retinoic acid and aggregation. Comparison of embryonic stem cells to P19 cells shows that the two systems regulate a similar set of polypeptides and are thus likely to utilize a similar pathway. These studies are a step toward determining the full extent of regulation involved in the commitment of pluripotent cells to the neural fate.

A Quantitative Proteomic Analysis of Mitochondrial Participation in P19 Cell Neuronal Differentiation

“…6 A similar 2DE analysis of protein abundance in P19 cells undergoing neuronal development revealed 500 abundant polypeptides, 17 of which were regulated. 7 However, this early study did not identify many of these proteins. A subsequent study of P19 cell neural differentiation identified only 28 differentially expressed proteins.…”

“…A study of the in vitro differentiation of adult rat hippocampal neural stem cells by two-dimensional electrophoresis (2DE), resulted in the identification of 367 regulated spots, of which 128 could be identified, although many of these proteins were isoforms . A similar 2DE analysis of protein abundance in P19 cells undergoing neuronal development revealed 500 abundant polypeptides, 17 of which were regulated . However, this early study did not identify many of these proteins.…”

Early expression of the BM88 antigen during neuronal differentiation of P19 embryonal carcinoma cells

“…This model closely resembles events occurring during mammalian embryogenesis [1,9,26,30,39]. Exposure of P19 EC cells to RA gives rise to a population of dierentiating neurons and, later, to astrocytes that express similar developmental markers as their embryonic counterparts [36]. The polysialylated form of NCAM (PSA-NCAM), the product of an NCAM-speci®c PSA synthase (ST8Sia II/STX), can serve in this in vitro model as an early indicator of neuronal dierentiation [4,17].…”

Comparative Proteomic Analysis of Proteins Involved in Cell Aggregation during Neural Differentiation of P19 Mouse Embryonic Carcinoma Cells

“…The proteomics approach is a powerful tool to find clues to investigate the mechanism of a biological process due to its capability of resolving thousands of proteins at one time. The first proteomic analysis of P19 cell aggregation was carried out by Ray and Gottlieb in 1996, 20 when they tried to find differentially expressed protein spots between aggregated and nonaggregated P19 cells in their 2DE gels, but they failed to reveal any aggregation-related proteins, probably due to technical limitations. Advances in 2DE technology and peptide mass spectrometry offer a promising proteomic approach to study cell aggregation during P19 cell neural differentiation.…”