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Transcriptomic and Proteomic Analyses of Mouse Cerebellum Reveals Alterations in RasGRF1 Expression Following In Vivo Chronic Treatment with Delta 9-Tetrahydrocannabinol

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Abstract

We have applied transcriptomic and proteomic techniques to identify changes in the RNA and the protein levels in the mouse cerebellum after chronic treatment with Δ9-tetrahydrocannabinol (THC). Among approximately 14,000 transcripts in a mouse cDNA microarray library, we found 11 genes with altered expression. RasGRF1, a neuron-specific Ras guanine nucleotide exchange factor, showed a reduction both at the RNA and protein levels with a specific decrease of the protein pool associated to cell membranes. In addition, proteomic analysis on cerebellum obtained from chronically THC-treated mice detected quantitative changes of additional 27 spots, mostly in the membranous fraction. We found enrichment of alpha (Gαo, Gαq) and beta subunits (β4/β2 and β5) of guanine nucleotide-binding proteins and of two calcium-binding proteins, calretinin and hippocalcin-like protein-1. In addition, we also detected a significant increase in the membrane fraction of proteins involved in exo–endocytosis such as septins, dynamin-1, and vesicle protein sorting 29. By western blotting, we confirmed increased membrane localization of calretinin and of dynamin-1 isoforms with higher isoelectric point, indicative for an underphosphorylated state of the molecule. In conclusion, our results indicate that chronic THC modulates the expression and subcellular localization of proteins implicated in Ras signaling, calcium-buffering potential, and trafficking.

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Abbreviations

THC:

delta 9-tetrahydrocannabinol

2D-GE:

two-dimensional gel electrophoresis

PF:

parallel fiber

PC:

Purkinje cells

GEF:

guanine nucleotide exchange factor

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Acknowledgments

We thank E. Sturani, R. Brambilla, and R. Tonini for critical comments and useful discussion. This work was supported by a grant from MIUR to R. Zippel and FIRB-Neuroscienze (PRO-NEURO) to E. Martegani.

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Authors and Affiliations

  1. Department of Biomolecular Sciences and Biotechnology, University of Milan, Via Celoria 26, 20133, Milan, Italy

    Graziano Colombo, Francesco Rusconi & Renata Zippel

  2. Department of Structural and Functional Biology, University of Insubria, Varese, Italy

    Tiziana Rubino & Daniela Parolaro

  3. Biological Mass Spectrometry Unit, DIBIT San Raffaele Scientific Institute, Milan, Italy

    Angela Cattaneo & Angela Bachi

  4. Department of Biotechnology and Bioscience, University of Milano-Bicocca, Milan, Italy

    Enzo Martegani

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  1. Graziano Colombo
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  2. Francesco Rusconi
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  3. Tiziana Rubino
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  4. Angela Cattaneo
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  5. Enzo Martegani
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  8. Renata Zippel
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Correspondence to Renata Zippel.

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Colombo, G., Rusconi, F., Rubino, T. et al. Transcriptomic and Proteomic Analyses of Mouse Cerebellum Reveals Alterations in RasGRF1 Expression Following In Vivo Chronic Treatment with Delta 9-Tetrahydrocannabinol. J Mol Neurosci 37, 111–122 (2009). https://doi.org/10.1007/s12031-008-9114-2

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