BACKGROUND: The firefly luciferase reporter protein is a crucial tool for studies
targeting a broad range of biological questions. Importantly, luciferase assays are
also widely used to explore mechanisms underlying thyroid hormone dependent regulation
of gene expression. However, it was demonstrated that the firefly luciferase reporter
is subject to triiodothyronine (T3)-evoked, promoter independent downregulation that
is mediated by the thyroid hormone receptor. Since this effect can interfere with
readout accuracy, the study aimed to find luciferase reporters that are not susceptible
to this phenomenon. METHODS: Luciferase reporter constructs were generated under the
control of a minimal thymidine kinase (TK) promoter and transiently transfected into
JEG-3 cells to test their activity upon T3 treatment. RESULTS: Activity of the TK-(dCpG)Luc
encoding a synthetic (dCpG)Luciferase and TK-NanoLuc expressing the NanoLuc reporter
was not significantly changed by T3 treatment while the firefly luciferase control
was suppressed by approximately 2.6-fold. T3 also downregulated the activity of Renilla
luciferase by approximately 30%. CONCLUSIONS: Novel types of luciferase reporters,
especially the synthetic (dCpG)Luciferase, can be more accurate to study T3-regulated
gene expression than the classical firefly luciferase reporter. Renilla luciferase,
a popular transfection control of dual luciferase assays, should be used with caution
in conditions with T3 treatment.
