Alternative processing of thyroid hormone receptor mRNA: Role of cis-acting sequences and antisense RNA

Presentation

Stephen Munroe
Department of Biology
Wehr Life Sciences Building
Marquette University
PO Box 1881
Milwaukee, WI 53201
telephone: 414-288-1485
fax: 414-288-7357
email: stephen.munroe@marquette.edu
prestype: Platform
presenter: Munroe
http://mendel.biol.mu.edu/faculty/munroe.html

Stephen H. Munroe, Michelle L. Hastings and Valerie K. Salato
Department of Biology, Marquette University

In mammals mRNA for the a-type thyroid hormone receptor (TRa1) is transcribed from a complex locus that encodes several nuclear receptors. Alternative processing give rise to two mRNAs encoding two functionally antagonistic proteins, TRa1 and TRa2. The latter is a variant receptor that lacks the C-terminal domain required for hormone binding and transcriptional activation. A third nuclear receptor, Rev-ErbAa (RevErb), is encoded on the opposite strand and overlaps TRa2 but not TRa1 mRNA. Cells expressing high levels of RevErb generally have higher amounts of TRa1 relative to TRa2. In vivo overexpression of RevErb leads to a shift in the TRa1/TRa2 ratio, and in vitro RevErb RNA efficiently blocks TRa2 splicing. These results support an antisense mechanism in which RevErb selectively blocks TRa2 mRNA splicing. TRa2 mRNA is formed by splicing out a portion of the 3' terminal exon of TRa1 mRNA and utilizing a downstream 3' exon specific to TRa2. In common with other regulated alternative processing sites, those specific to TRa1 or TRa2 mRNA are suboptimal. Point mutations within the TRa2-specific splice sites that increase the match to the consensus splice site sequences greatly increase the level of TRa2 mRNA. Similarly, replacing the TRa1-specific polyA site with a strong viral polyA site increases the level of TRa1 mRNA to the exclusion of TRa2 splicing. Recent work has focused on a highly conserved 80 splicing enhancer located near the 5' end of the TRa2-specific intron, immediately downstream of the TRa1 stop codon. This element includes a conserved pseudo-5'ss sequence that is critical for TRa2 mRNA splicing. A number of single nucleotide replacements within the pseudo-5'ss reduce TRa2 splicing. However, point mutation that improve the match to the 5'ss consensus sequence activate cryptic splicing. Several proteins, including hnRNP H and SF2/ASF, crosslink specifically to the enhancer element. Mutations within the pseudo-5'ss that inactivate TRa2 splicing also disrupt the crosslinking of these proteins, strongly indicating that these proteins play a role in splicing. Further experiments examine the role of a 3' intronic splicing enhancer, the TRa1 polyadenylation site and the role of the complementary RevErb mRNA in modulating the balance between TRa1 and TRa2 expression.

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CONTACT:
stephen.munroe@marquette.edu
http://mendel.biol.mu.edu/faculty/Munroe.html