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Drug Addiction: Mechanism Illuminated

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Dr. Judith A. Potashkin, Ph.D. and her colleagues at Rosalind Franklin University of Medicine and Science recently completed a study investigating one of the changes in gene expression that occurs when individuals take addictive drugs. 

Dr. Potashkin, Associate Professor and Vice Chair of the Department of Cellular and Molecular Pharmacology, is an expert in gene expression. She commented, “Addiction is a brain disorder that manifests itself by repetitive behaviors despite negative consequences.  Currently, there is an abundance of information known about the cellular and behavioral changes that occur during addiction, but little is understood concerning the changes that occur at the molecular level with regards to gene expression. Understanding the changes that occur between transcription and protein synthesis is key to revealing the mechanism that leads to addiction.”

Dr. Potashkin’s studies focus on how the primary RNA transcript is processed by splicing to produce a mature transcript. The fidelity of splicing must be maintained since errors may lead to the development of disease.

One immediate and prominent alteration that occurs with administration of amphetamine or cocaine is the accumulation in one region of the brain of very stable truncated isoform of the transcription factor FosB termed DFosB that is produced by alternative splicing of the transcript. DFosB mediates some of the neural and behavioral modifications that occur with drug addiction.

The results from the study identified a splicing factor, polypyrimidine tract binding protein, as a key factor in regulating the switch in splicing that result in the truncated form of FosB being produc ed instead of the less stable full-length protein.

The study also provided clues about the signaling pathway that is activated that leads to splicing regulation. This information provides several potential therapeutic targets for drug addiction.

PLoS One 2(9): e828.doi:10.1371/journal.pone.0000828

Regulation of Retention of FosB Intron 4 by PTB

Victor Marinescu#, Patricia A. Loomis#, Svetlana Ehmann, Mitchell Beales, Judith A. Potashkin*

Department of Cellular and Molecular Pharmacology, The Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, Illinois, United States of America

One effect of stressors such as chronic drug administration is that sequence within the terminal exon of the transcription factor FosB is recognized as intronic and removed by alternative splicing. This results in an open-reading-frame shift that produces a translation stop codon and ultimately a truncated protein, termed ΔFosB. In vitro splicing assays with control and mutated transcripts generated from a fosB mini-gene construct indicated a CU-rich sequence at the 3′ end of intron 4 (I4) plays an important role in regulating fosB pre-mRNA splicing due to its binding of polypyrimidine tract binding protein (PTB). PTB binding to this sequence is dependent upon phosphorylation by protein kinase A and is blocked if the CU-rich sequence is mutated to a U-rich region. When this mutated fosB minigene is expressed in HeLa cells, the splicing efficiency of its product is increased compared to wild type. Moreover, transient transfection of PTB-1 in HeLa cells decreased the splicing efficiency of a wild type fosB minigene transcript. Depletion of PTB from nuclear extracts facilitated U2AF65 binding to wild type sequence in vitro, suggesting these proteins function in a dynamic equilibrium to modulate fosB pre-mRNA alternative splicing. These results demonstrate for the first time that phosphorylated PTB promotes intron retention and thereby silences the splicing of fosB I4.

Written by huehueteotl

September 12, 2007 at 12:08 pm

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