Prosapip1-a novel target of alcohol drinking behaviors

We previously reported that excessive consumption of alcohol activates the H-Ras/PI3K/AKT signaling in the nucleus accumbens (NAc) of rodents, which ultimately leads to the activation of the mechanistic target of rapamycin complex 1 (mTORC1) [1-3]. mTORC1 is localized in dendrites, and plays an important role in synaptic plasticity by promoting the translation of synaptic proteins [4]. We therefore hypothesized that mTORC1-dependent mRNA translation contributes to neuroadaptations that underlie the development and/or maintenance of excessive alcohol consumption. To do so, we utilized the high throughput RNA sequencing (RNA seq) approach to identify novel gene products whose mTORC1-dependent translation is induced in the NAc in response to excessive alcohol consumption. Specifically, mice underwent an intermittent access to 20% alcohol or water only using 2-bottle choice paradigm for 8 weeks, and were then treated with rapamycin or vehicle. The NAc was dissected, polysomes (RNA undergoing translation) were isolated, and RNA seq was performed. Among the 12 identified candidates whose translation was dependent of mTORC1 was ProSAP-interacting protein 1 (Prosapip1), a synaptic protein [5] that we found to be highly expressed in the striatum, whose function is not well understood. First, we confirmed the RNAseq data and showed that the translation of Prosapip1 was induced in response to excessive alcohol drinking in an mTORC1-dependent manner. We further showed that the increase in mRNA of Prosapip1 was due solely to translation and not transcription. The corresponding increase in Prosapip1 protein in the NAc of mice in response to a binge drinking session was localized to the synaptic fraction and was maintained even after 24 hours of withdrawal. Prosapip1 levels were not altered in other brain regions where mTORC1 is not activated by alcohol. To test for potential cellular consequences of Prosapip1 increases by alcohol, we overexpressed the protein in a neuroblastoma cell line and observed an increased filamentous F-actin content. Conversely, knockdown of Prosapip1 by shRNAs decreased the cell’s F-actin content. Next, we tested the behavioral outcomes of alcohol-mediated increase in Prosapip1 levels by overexpressing the gene in NAc neurons. We found that lenti-virus-mediated upregulation of Prosapip1 levels in the NAc of mice dramatically increased operant self-administration of 20% alcohol when a fixed-ratio 2 schedule (i.e. two presses on the active lever resulted in the delivery of one drink of alcohol) was applied. Furthermore, ProSAPIP1 overexpression in the NAc exacerbated the motivation of mice to obtain alcohol as assessed by measuring lever presses in a single operant session using a progressive ratio procedure (i.e. the number of active lever presses required to obtain each successive drink of alcohol was progressively increased). The enhancement of alcohol intake by ProSAPIP1 was not due to non-specific locomotion or cognitive changes. Together, our data suggest that alcohol-induced activation of mTORC1 in the NAc leads to translation of the synaptic protein Prosapip1 that in turn drives the motivation to obtain and consume alcohol.

1. Neasta, J., Proc Natl Acad Sci U S A (2010)
2. Neasta, J., Biol Psychiatry (2011)
3. Ben Hamida, S., J Neurosci (2012)
4. Lipton, J.O. and M. Sahin, Neuron (2014)
5. Wendholt, D., J Biol Chem (2006)

Supported by NIH P50 AA017072 (DR)