In vivo and in vitro tools to identify and study transcriptional regulation of USF-1 target genes.
In response to environmental stress, cells trigger a number of molecular mechanisms to control their survival and growth. These include changes in gene expression with corresponding Post-translational modifications to critical transcriptional-control proteins. Transcription is a highly-regulated process that is impacted by a large number of ubiquitous and specific factors. In order to determine how gene expression is regulated in response to environmental cues, it is necessary to correlate modifications to specific transcription proteins with an accurate assessment of the transcriptional response. This chapter details quantitative Real Time PCR (qPCR) and Luciferase assay protocols to illustrate, both in vivo and in vitro, the role of the USF-1 transcription factor in the UV-dependant regulation of pigmentation genes (POMC and MC1R). The procedures have been optimized for the USF-1 transcription factor and the regulation of specific target genes in response to physiological UV doses.
Galibert
Marie-Dominique
MD
Genetic and Development Institute of Rennes, Transcriptional Regulation and Oncogenesis Team, Rennes University, Rennes, France. mgaliber@univ-rennes1.fr
Corre
Sébastien
S
D045744:Cell Line, Tumor; D004789:Enzyme Activation / Q000528:radiation effects; D006801:Humans; D008156:Luciferases / Q000235:genetics; D010858:Pigmentation / Q000235:genetics / Q000528:radiation effects; D011333:Pro-Opiomelanocortin / Q000235:genetics; D044101:Receptors, Melanocortin / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014158:Transcription, Genetic; D014466:Ultraviolet Rays; D051720:Upstream Stimulatory Factors / Q000378:metabolism*; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D044101:Receptors, Melanocortin; D051720:Upstream Stimulatory Factors; D011333:Pro-Opiomelanocortin; D008156:Luciferases; D048051:p38 Mitogen-Activated Protein Kinases