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Differentiation. 2000 May;65(5):271-9.

Spontaneous retinoic acid receptor beta 2 expression during mesoderm differentiation of P19 murine embryonal carcinoma cells.

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1
Department of Cellular and Molecular Medicine, University of Ottawa, Ontario, Canada. cpratt@uottawa.ca

Abstract

Exposure of aggregated murine P19 embryonal carcinoma cells to dimethylsulfoxide (DMSO) induces mesoderm and both embryonic cardiac and skeletal muscle differentiation, while retinoic acid (RA) is an inducer of neuroectodermal differentiation. P19 cells constitutively express the retinoic acid receptor alpha (RAR alpha) and RAR gamma mRNAs while RAR beta expression is induced by RA through a consensus RA-response element in the RAR beta promoter. In the present study we show that the RAR beta transcript is strongly expressed in both P19 cells and in a RA-nonresponsive derivative of P19 cells, called RAC65, during DMSO-induced mesoderm and muscle differentiation. Reverse transcriptase-polymerase chain reaction analysis indicated that RAR beta 2 is the predominant isoform expressed in DMSO-differentiated cells, providing the first evidence for RA-independent regulation of RAR beta 2 transcript levels. Immunoblot analysis showed a 3-fold increase in the RAR beta protein expression over basal levels in differentiated cells, and immunohistochemistry indicated that all cells in the culture including muscle reacted positively for the RAR beta protein. RAR beta 2 transcript expression was differentiation-dependent and occurred without transactivation of a transfected RARE beta 2 reporter gene. Little transcription of the RAR beta gene was detected in nuclear run-off assays of undifferentiated P19 cells and only a small increase in transcription was observed in nuclei from DMSO-treated cells. RA treatment of P19 cells stably transfected with the RA-responsive element from the RAR beta gene showed that RAR beta 2 mRNA expression during DMSO differentiation was associated with increased sensitivity to RA. Together these data show that RAR beta 2 is expressed spontaneously in an apparently RA-independent manner in differentiating mesoderm and mesoderm derivatives, resulting in increased sensitivity to RA in these cells.

[Indexed for MEDLINE]

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