The Effects of Induced Expression of Rev-erb? mRNA on 
Complementary TR?2 mRNA Expression

Anina C. Tollett
Howard University
Mentor:  Dr. Stephen Munroe

Antisense regulation involves the control of expression of a RNA target through direct base pairing with a complementary antisense RNA.  Prokaryotic antisense regulators are very small, non-coding RNAs, many of which affect gene expression by blocking translation.  Many examples of antisense regulation have been proposed in eukaryotic systems, but a definitive display of regulation by endogenous antisense RNAs has only been demonstrated in a few instances.

Alternative splicing of transcripts of the TR? gene yields two mRNAs: TR?1 and TR?2.  TR?1 mRNA encodes an ?-thyroid hormone receptor that is able to bind to the thyroid hormone, T3, subsequently activating gene expression.  TR?2 mRNA encodes a variant nuclear receptor that lacks a C-terminus, therefore, compromising its ability to bind T3. However, it does inhibit transcription of a T3-regulated gene by binding DNA.  TR?2 antagonizes T3 action by competing with TR?1 for specific DNA binding sites.  In addition, a complementary mRNA encoding Rev-erb? overlaps TR?2 mRNA, but not TR?1 mRNA.  Rev-erb? and TR?2 are cis-encoded antisense RNA pairs.  Some evidence suggests that expression of the complementary Rev-erb? mRNA may regulate levels of TR?2 mRNA through base-pairing with the overlapping sequence, resulting in a decrease of TRa2 and an increase in the TR?1/TR?2 ratio.  We have induced Rev-erb? mRNA expression from a chimeric gene that includes both Rev-erb? and GFP reporter sequences regulated by a tetracycline responsive promoter.  This induction provides a system from which we can determine if TR?2 mRNA expression is affected by over expression of Rev-erb? mRNA.

In our studies, we have found nine cell lines that display a large increase in RevGFP transgene expression upon the removal of doxycycline.  Although no strong correlation has been observed between changes of Rev-erb? mRNA levels and TR?2 mRNA levels, the ratio of Rev-erb? mRNA to TR?2 mRNA varies widely.  Thus future investigations must be undertaken to distinguish between endogenous TR?1 and TR?2 mRNAs and possible expression of transgene RNA in these cell lines.

<Summer Research Program Home