Summer Research
* Student(s): Scott Myer , Scott Alpert
Advisor: Herman Lehman
Title: Identification of the Tyramine b-Hydroxylase gene in Manduca sexta
Abstract:
Our studies have focused on octopamine, a neurotransmitter commonly found in the nervous systems of insects and other arthropods. Previously, members of this laboratory have demonstrated that tyramine b-hydroxylase (TbH) catalyzes the formation of octopamine and is functionally similar to dopamine b-hydroxylase (DbH), a mammalian enzyme required for the hydroxylation of dopamine to form norepinephrine. In addition, octopamine levels and TbH activity are regulated developmentally during insect metamorphosis and the steroid hormone, 20-hydroxyecdysone (20-HE) is necessary and sufficient for the developmental regulation of these molecules in the insect nervous system.
To unravel the molecular mechanisms of action of steroid hormones, we have focused on the molecular cloning and identification of the gene that encodes TbH. Using PCR technology and degenerate primers based upon conversed regions of the DbH a 578 bp cDNA fragment that codes for TbH has been cloned and sequenced. This cDNA fragment was labeled with 32P dCTP and used to screen three different nervous system cDNA libraries. More than 3.5 million individual plaques were screened for the presence of the TbH gene and five potential positives have been identified. These positive plaques are currently being re-screened and the phagemid excised for the positive identification of the TbH gene. Once the presence of the TbH gene has been confirmed the sequencing of the gene will commence.
Stipend support for Scott Meyer came from the Sergei Zlinkoff Fund and Scott Albert was supported by New York Science Education Program (NYSEP).