Identification of a human 5-HT6 receptor variant produced by alternative splicing

Area of Science:

• Neuroscience
• Molecular Biology
• Genetics

Background:

• The 5-hydroxytryptamine (5-HT) or serotonin receptor family is known to have complex signaling pathways.
• Previous research identified splice variants for several 5-HT receptor subtypes, including 5-HT2B, 5-HT2C, 5-HT4, and 5-HT7.
• The existence of splice variants for the 5-HT6 receptor subtype in the human brain was previously unconfirmed.

Purpose of the Study:

• To investigate the existence and characteristics of 5-HT6 receptor splice variants in the human brain.
• To determine the molecular basis and cellular localization of any identified 5-HT6 variants.
• To assess the functionality of the variant 5-HT6 receptor protein.

Main Methods:

• Degenerate Polymerase Chain Reaction (PCR) was employed using human caudate cDNA to identify potential 5-HT6 receptor variants.
• Sequence analysis was performed on identified clones to determine the nature of the variant.
• Alternative splicing patterns were analyzed by comparing human, rat, and mouse cDNA.
• Coexpression of variant and full-length transcripts was examined in different human brain regions using PCR.
• Transient transfection of the variant 5-HT6 construct into Cos-7 cells was conducted to evaluate protein localization and function.

Main Results:

• A 5-HT6 receptor splice variant was identified in human brain cDNA, characterized by a 289 bp deletion.
• This deletion spans transmembrane region IV to the third intracellular loop, causing a frameshift and premature stop codon.
• The resulting truncated protein consists of 10 unique amino acids at the carboxyl terminus.
• The variant transcript arises from alternative splicing utilizing an upstream donor site and an intronic acceptor site.
• This specific splicing pattern was not observed in rat or mouse brain cDNA.
• Coexpression of the variant and full-length 5-HT6 transcripts was detected in the caudate and substantia nigra, but not in the hippocampus, cortex, cerebellum, or thalamus.
• Transfected Cos-7 cells showed membrane translocation of the truncated 5-HT6 variant, but it appeared nonfunctional.

Conclusions:

• A novel splice variant of the human 5-HT6 receptor has been identified, resulting from alternative splicing.
• This variant produces a truncated, nonfunctional receptor protein that localizes to the cell membrane.
• The discovery of this variant adds to the complexity of the 5-HT6 receptor system and may have implications for understanding serotonin signaling in specific brain regions.