5HT2CR pre-mRNA can be the subject of
RNA editing. It is the only serotonin receptor as well as the only member of the large family of 7 transmembrane receptors (7TMRs) known to be edited. Different levels of editing result in a variety of effects on receptor function.
Type The type of RNA editing that occurs in the
pre-mRNA of the 5HT2CR is Adenosine to Inosine (A to I) editing. A to I RNA editing is catalyzed by a family of
adenosine deaminases acting on RNA (ADARs) that specifically recognize adenosines within double-stranded regions of pre-mRNAs and deaminate them to
inosine.
Inosines are recognised as
guanosine by the cells translational machinery. There are three members of the ADAR family ADARs 1–3 with
ADAR1 and
ADAR2 being the only enzymatically active members.
ADAR3 is thought to have a regulatory role in the brain. ADAR1 and ADAR2 are widely expressed in tissues while ADAR3 is restricted to the brain. The double stranded regions of RNA are formed by base-pairing between residues in the close to region of the editing site with residues usually in a neighboring intron but can be an exonic sequence. The region that base pairs with the editing region is known as an Editing Complementary Sequence (ECS). ADARs bind interact directly with the dsRNA substrate via their double stranded RNA binding domains. If an editing site occurs within a coding sequence, it can result in a codon change. This can lead to translation of a protein isoform due to a change in its primary protein structure. Therefore, editing can also alter protein function. A to I editing occurs in a non coding RNA sequences such as
introns,
untranslated regions (UTRs),
LINEs, SINEs ( especially Alu repeats) The function of A to I editing in these regions is thought to involve creation of splice sites and retention of RNAs in the nucleus amongst others.
Location Editing occurs in 5 different closely located sites within exon 5, which corresponds to the second intracellular loop of the final protein. The sites are known as A, B, C′ (previously called E), C and D, and are predicted to occur within amino acid positions 156, 158 and 160. Several codon changes can occur due to A-to-I editing at these sites. Thirty-two different mRNA variants can occur leading to 24 different protein isoforms. • An Isoleucine to Valine (I/V) at amino acid position 157,161. • An Isoleucine to a Methionine(I/M) at amino acid position 157 • An Aspartate to a Serine (N/S)at 159 • An Aspartate to Asparagine(N/D) at 159 • An Asparagine to a Glycine(N/G) at 159. These codon changes which can occur due to A to I editing at these sites can lead to a maximum of 32 different mRNA variants leading to 24 different protein isoforms. The number of protein isoforms is less than 32 since some amino acids are encoded by more than one codon. Another editing site, site F has also been located in the exon complementary sequence (ECS) of intron 5. The ECS required for formation of double stranded RNA structure is found within intron 5. The initial demonstration of RNA editing in rat. The editing complementary sequence is known to be conserved across Mammalia.
Regulation The 5-HT2c receptor is the only serotonin receptor edited despite its close sequence similarities to other family members. Mice knock out and other studies have been used to determine which ADAR enzyme are involved in editing. Editing at A and B sites has been demonstrated to be due to ADAR1 editing. Also since ADAR1 expression is increased in response to the presence of interferon α, it was also observed that editing at A and B sites was also increased because of this. The C site has been shown to be mainly edited by ADAR2 but in presence of upregulated expression of ADAR1, there was an increase in editing of this site and the enzymes presence can also result in limited editing in ADAR 2 knock out mice. In contrast, the 5-HT2CR-INI is constitutively internalized and accumulates in endosomes 78.
Structure As mentioned editing results in several codon changes. The editing sites are found in the second intracellular domain of the protein which is also the receptors G protein coupling domain. Therefore, editing of these sites can affect the affinity of the receptor for G protein binding. Editing is also thought to influence splicing. Three different spliced isoforms of the receptor exist. Editing regulates the amount of 5HT2CR mRNA which leads to translation of the full length protein selection of alternative splice sites. t76,77. These splice sites are termed Gu1, Gu2, GU3. Only GU2 site splicing results in translation of the full length receptor while editing at GU1 is known to result in translation of a truncated protein. This is thought to be a regulatory mechanism to decrease the amount of unedited isoform INI to limit serotonin response when editing is inefficient. Most of the pre-mRNAs which are edited are spliced at the GU2 site. There have been several experimental investigations into the effects of alternative editing patterns of the 5HT2CR and these conditions with a wide variability in results especially those relating to schizophrenia. Some studies have noted that there is an increase in RNA editing at site A in depressed suicide victims. In rat models this increase is also observed and can be reversed with fluoxetine with some suggestion that E site editing maybe linked to major depression. == See also ==