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RNA editing of the glutamate receptor subunits GluR2 and GluR6 in human brain tissue
W Paschen1, J C Hedreen, C A Ross
1Department of Psychiatry and Neuroscience, Johns Hopkins University, School of Medicine, Baltimore.
Abstract:
Editing of mRNA in the coding region of the second transmembrane domain of glutamate receptor subunits GluR2, GluR5, and GluR6 involves a change of the base A in genomic DNA to the base G in mRNA as described in rat brain. To determine whether this reaction occurs in humans as well as rats, we studied RNA editing of GluR2 and GluR6 in human brain. We compared the extent of editing in controls and cases with Huntington's disease. To assay the extent of editing in brain RNA, first strand cDNA was amplified using the polymerase chain reaction yielding a product across the region of the second transmembrane spanning segment in which editing takes place in rats. The PCR product was incubated with the restriction enzyme BbvI, which recognizes the sequence GCAGC present in the nonedited sequence of the mRNA in subunits GluR2 and GluR6. Thus, BbvI cuts the nonedited version but leaves the edited version intact. As in the rat, the GluR2 subunit mRNA was completely edited in human brain. The GluR6 subunit was nearly completely edited in all gray matter structures investigated including cortex, striatum, thalamus, hippocampus, amygdala, and cerebellum with extent of editing ranging from 89% in the cerebellum to 95% in the cortex and striatum. No significant differences in the extent of RNA editing were apparent in control versus Huntington's disease brains. To compare the extent of editing in neurons and glia in the brain, editing in cerebral cortex (predominantly gray matter and thus neurons) was compared with editing in corpus callosum (white matter and thus nearly completely glial cells).(ABSTRACT TRUNCATED AT 250 WORDS)
Insights
RNA editing of glutamate receptor subunits GluR2 and GluR6 occurs in the human brain, similar to rats. This process was fully efficient for GluR2 and nearly complete for GluR6 across various brain regions, with no changes observed in Huntington
Area of Science:
- Neuroscience
- Molecular Biology
- Genetics
Background:
- RNA editing, specifically base A-to-G conversion in mRNA, occurs in the coding region of glutamate receptor subunits.
- This editing is known in rat brains for GluR2, GluR5, and GluR6 subunits.
Purpose of the Study:
- To investigate RNA editing of glutamate receptor subunits GluR2 and GluR6 in the human brain.
- To compare RNA editing levels between control and Huntington's disease brains.
- To examine differences in RNA editing between neurons and glial cells.
Main Methods:
- RNA was extracted from human brain samples.
- First-strand cDNA was synthesized and amplified using polymerase chain reaction (PCR) across the editing site.
- PCR products were digested with the restriction enzyme BbvI, which distinguishes edited from unedited mRNA.
Main Results:
- GluR2 mRNA was completely edited in the human brain.
- GluR6 mRNA editing was nearly complete (89-95%) across all gray matter regions studied.
- No significant differences in RNA editing were found between control and Huntington's disease brains.
- RNA editing levels were comparable in neurons (cerebral cortex) and glial cells (corpus callosum).
Conclusions:
- RNA editing of GluR2 and GluR6 subunits is conserved between rats and humans.
- RNA editing is highly efficient in various human brain regions and cell types.
- Huntington's disease does not appear to affect the extent of RNA editing in these glutamate receptor subunits.