By Marisa Sanchez, Molecular and Cellular Biology ’15

SV2A is a synaptic vesicle protein, which participates in the regulation of neurotransmitter release in humans. SV2A is expressed in neurons and endocrine cells. The exact function of SV2A is still unknown, but it has been identified that SV2A is the binding site for the antiepileptic drug, levetiracetam. Levetiracetam reduces presynaptic glutamate release, especially in neurons with high frequency firing. Abnormally enhanced glutamatergic neurotransmission with high frequency neural firing is found in epilepsy and several neurodegenerative disorders, such as Parkinson’s disease, Alzheimer’s disease, and Huntington’s disease.

In hopes to discover more about the role of SV2A in neurogenetic disorders, human SV2A was studied in yeast cells. In the study, many copies Human SV2A was made using PCR amplification.The SV2A gene was then ligated in-frame into a yeast centromeric expression vector. Standard yeast media was used in the initial growth experiments. After the yeast colonies grew and colonies containing SV2A were identified, the cells were transferred to different media containing different carbon sources. These yeast cells were deficient in 6-carbon sugar transport, which means they lack proteins required for 6-carbon sugar uptake, and these yeast colonies will die on media containing 6-carbon sugars as a nutrient source.  However, yeast colonies containing SV2A were able to grow on media that contained galactose, but not media containing other 6-carbon sugar sources, which proves SV2A is a galactose transporter, which means that galactose transport capability of SV2A may play an important role in regulating synaptic function.

To study the effects of the antiepileptic drug levetiracetam on galactose uptake, this drug was added to media that contained galactose. Yeast cells containing SV2A were then transferred to this media from the standard growth media. These yeast cells did not grow, which indicates that levetiracetam inhibits the galactose-dependent growth of hexose transport-deficient yeast cells expressing SV2A. This shows that levetiracetam inhibits galactose uptake and transport by SV2A. These results further support the findings that SV2A is a galactose transporter, and that galactose has an important role in neuronal physiology.

To see if SV2A-mediated galactose uptake was coupled to proton transport, CCCP, which is a protonophore (a proton translocator, which allows protons to cross lipid bilayers, which does not normally occur without presence of a transport protein) was added to a fixed time assay. CCCP completely blocked the transport of galactose by SV2A. This indicates that SV2A is a proton-coupled symporter, which means that SV2A, as a transporter, has dual functions in neurons. These experiments have shown that SV2A is a galactose transporter, but further studies still need to be done to determine the exact function of SV2A in neurons and endocrine cells.

Reference:

Madeo, Marianna, Attila D. Kovacs, and David A. Pearce. “The Human Synaptic Vesicle

Protein, SV2A, Functions as a Galactose Transporter in Saccharomyces Cerevisiae.” The Journal of Biological Chemistry 289.48 (2014): 33066-3071.