Molecular cloning, nucleotide sequence and presence of multiple functional polyadenylation signals in the 3′-untranslated region of equine dopamine beta-hydroxylase cDNA.

Researchers cloned and sequenced the cDNA of the dopamine beta-hydroxylase (DBH) enzyme in horses, and identified three polyadenylation signals in the 3′-untranslated region, indicating the production of DBH mRNA variants in horse adrenal glands.

Overview of the Research

• The study was focused on the DBH enzyme in equines (horses), an enzyme involved in the creation of neurotransmitters in the nervous system.
• The researchers employed techniques of reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) to generate the cDNA encoding for DBH. The cDNA sequence was determined to be 3842 base pairs (bp) in length.

Analyzing the DBH cDNA Sequence

• The cDNA sequence contained 5 bp of 5′ flanking untranslated sequence, 1833 bp of an open reading frame encoding for 610 amino acids, and 2004 bp of 3′ flanking untranslated sequence.
• The study found that the amino acid sequence of equine DBH was highly similar to DBH sequences in other mammalian species – with 86.3% similarity to bovine (cows), 84.6% to human, 82.2% to rat, and 81.2% to mouse DBH sequences.

Expression of DBH mRNA

• Employing techniques of northern blot analysis and in situ hybridization, the researchers uncovered three different sized expressions of DBH mRNA in equine adrenal medulla tissue.
• Three polyadenylation signal sites, sequences in the DNA that indicate where an mRNA sequence can end, were found in the 3′ flanking untranslated sequence.

Implications of the Study

• The presence of three polyadenylation sites suggested that the horse DBH gene can generate mRNA variants of different sizes, leading to potentially different forms of the DBH enzyme in horses.
• This research adds to the understanding of the DBH enzyme in horses, specifically, and mammalian neurotransmitter production more generally.
• The findings also provide a foundation for further studies into horse specific DBH gene variations and potential impacts on physiology and behavior.