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Molecules 2015, 20(5), 8304-8315; doi:10.3390/molecules20058304

Culture Conditions Affect Expression of DUX4 in FSHD Myoblasts

1
Research Center for Genetic Medicine, Children's National Medical Center, Washington, DC 20010, USA
2
Department of Integrative Systems Biology and Department of Pediatrics, George Washington University, Washington, DC 20037, USA
*
Author to whom correspondence should be addressed.
Academic Editor: Leonidas A. Phylactou
Received: 24 February 2015 / Revised: 28 April 2015 / Accepted: 29 April 2015 / Published: 8 May 2015
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Abstract

Facioscapulohumeral muscular dystrophy (FSHD) is believed to be caused by aberrant expression of double homeobox 4 (DUX4) due to epigenetic changes of the D4Z4 region at chromosome 4q35. Detecting DUX4 is challenging due to its stochastic expression pattern and low transcription level. In this study, we examined different cDNA synthesis strategies and the sensitivity for DUX4 detection. In addition, we investigated the effects of dexamethasone and knockout serum replacement (KOSR) on DUX4 expression in culture. Our data showed that DUX4 was consistently detected in cDNA samples synthesized using Superscript III. The sensitivity of DUX4 detection was higher in the samples synthesized using oligo(dT) primers compared to random hexamers. Adding dexamethasone to the culture media significantly suppressed DUX4 expression in immortalized (1.3 fold, p < 0.01) and primary (4.7 fold, p < 0.01) FSHD myoblasts, respectively. Culture medium with KOSR increased DUX4 expression and the response is concentration dependent. The findings suggest that detection strategies and culture conditions should be carefully considered when studying DUX4 in cultured cells. View Full-Text
Keywords: DUX4; Superscript; FSHD; dexamethasone; KOSR DUX4; Superscript; FSHD; dexamethasone; KOSR
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Pandey, S.N.; Khawaja, H.; Chen, Y.-W. Culture Conditions Affect Expression of DUX4 in FSHD Myoblasts. Molecules 2015, 20, 8304-8315.

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