Identifying trafficking signals in the C-terminus of the Drosophila melanogaster ortholog of the Vesicular Monoamine Transporter (DVMAT).

摘要

The Vesicular Monoamine Transporter (VMAT) is a twelve-transmembrane protein that resides on secretory vesicles in neurons and transports monoamines such as serotonin, dopamine, norepinephrine and histamine from the cytosol into vesicles for subsequent exocytotic release. Although it is generally accepted that monoamine neurotransmitters regulate diverse physiological processes in the central nervous system (CNS) and have been implicated in a variety of psychiatric and neurodegenerative diseases, the molecular pathways by which monoamine neurotransmission is regulated are poorly understood. One way that monoamine neurotransmission can be regulated is through the trafficking, localization and expression of VMAT. In this dissertation, we set out to determine whether one isoform of the Drosophila melanogaster ortholog of VMAT (DVMAT-A) contains signals that are important for its trafficking and whether disrupting these signals leads to altered monoamine-linked behaviors. We find that tyrosine at the 600 position (Y600) in the C-terminus of DVMAT-A is necessary for its endocytosis in S2 cells as well as in the Drosophila neuronal cell line DmBG2C6, whereas mutation of a dileucine motif disrupts endocytosis only in DmBG2C6 cells. Furthermore, we show that mutation of Y600 in vivo results in a dramatic reduction in DVMAT-A localization to SVs. Additionally, we find that C-terminal deletion mutant Delta3 DVMAT-A, in which Y600 as well as a potential dileucine motif are deleted, also reduces DVMAT-A localization to SVs and additionally does not rescue female infertility in null dVMAT flies. Based on our findings we propose that certain aminergic circuits and/or behaviors are under tightly regulated synaptic transmission whereas other circuits and/or behaviors remain unaltered even when regulated synaptic monoamine release is disturbed. Future studies will be aimed at determining additional residues that may be important for proper DVMAT-A trafficking as well as which specific circuits/behaviors in the fly require tightly regulated monoamine release. We hope that these and future studies will illuminate some of the molecular aspects that regulate monoamine neurotransmission and may suggest how they are altered during disease.