Issue 5, 2009

Proteomic profiling of the stem cellresponse to retinoic acid and synthetic retinoid analogues: identification of major retinoid-inducible proteins

Abstract

The natural retinoid, all-trans retinoic acid (ATRA), is widely used to direct the in vitro differentiation of stem cells. However, substantial degradation and isomerisation of ATRA in response to UV-vis light has serious implications with regard to experimental reproducibility and standardisation. We present the novel application of proteomic biomarker profiling technology to stem cell lysates to rapidly compare the differentiation effects of ATRA with those of two stable synthetic retinoid analogues, EC19 and EC23, which have both been shown to induce differentiation in the embryonal carcinoma cell line TERA2.cl.SP12. MALDI-TOF MS (matrix-assisted laser desorption ionisation time-of-flight mass spectrometry) protein profiles support previous findings into the functional relationships between these compounds in the TERA2.cl.SP12 line. Subsequent analysis of protein peak data enabled the semi-quantitative comparison of individual retinoid-responsive proteins. We have used ion exchange chromatographicprotein separation to enrich for retinoid-inducible proteins, thereby facilitating their identification from SDS-PAGE gels. The cellular retinoid-responsive proteins CRABP-I, CRABP-II, and CRBP-I were up-regulated in response to ATRA and EC23, indicating a bona fideretinoid pathway response to the synthetic compound. In addition, the actin filament regulatory protein profilin-1 and the microtubule regulator stathmin were also elevated following treatment with both ATRA and EC23. The up-regulation of profilin-1 and stathmin associated with retinoid-induced neural differentiation correlates with their known roles in cytoskeletal reorganisation during axonal development. Immunological analysis via western blotting confirmed the identification of CRABP-I, profilin-1 and stathmin, and supported their observed regulation in response to the retinoid treatments.

Graphical abstract: Proteomic profiling of the stem cellresponse to retinoic acid and synthetic retinoid analogues: identification of major retinoid-inducible proteins

Article information

Article type
Paper
Submitted
14 Oct 2008
Accepted
11 Feb 2009
First published
12 Mar 2009

Mol. BioSyst., 2009,5, 458-471

Proteomic profiling of the stem cellresponse to retinoic acid and synthetic retinoid analogues: identification of major retinoid-inducible proteins

D. J. Maltman, V. B. Christie, J. C. Collings, J. H. Barnard, S. Fenyk, T. B. Marder, A. Whiting and S. A. Przyborski, Mol. BioSyst., 2009, 5, 458 DOI: 10.1039/B817912C

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