Proteomic Analysis of Neural Epigenetic Mechanisms and Immune System Underlying Long-Lasting Effects of Developmental Cannabis Exposure

While developmental marijuana exposure has been shown to result in life-long vulnerability to reward, motivation and cognitive impairments in the offspring, the cellular and molecular mechanisms that mediate impairments of neuronal development and synaptogenesis remain largely elusive. Recently, epigenetic regulatory mechanisms of gene expression emerged as prime biological candidates to establish and maintain persistent aberrant neuronal processing as a result of developmental drug exposure. Since these mechanisms are highly dynamic and readily influenced by environmental agents including drugs of abuse, the developing brain might be particularly sensitive to epigenetic influences, given the dynamic neuroplasticity characteristic of this period. We have observed that in utero or adolescent exposure to THC leads to epigenetic and transcriptional changes of genes linked to dopamine and glutamate receptor disturbances in the nucleus accumbens that appear to have relevance to specific striatal pathways. Moreover, these patterns persist into adulthood. In this project (DA030359), we will study THC-related changes in epigenetic regulatory mechanisms at affected genes that might underlie the persistent alterations in gene expression. Our preliminary results indicated that post-translational modifications of histone H3 (i.e. H3K4me3) and specific histone modifying enzymes (Kmt2a) are affected by developmental THC exposure. However, histone modifications are known to influence each other and to act in combination. Moreover, several different enzyme complexes might be involved in establishing disease-specific anomalies of these marks. Using a proteomic approach, our goal is to explore the combinatorial histone modification landscape at specific loci, as well as the associated protein complexes that mediate these modifications.

A second important use of the proteomic approach in our research program relates to studying placental specimens from women who used cannabis while pregnant since we have observed significant alterations of the transcriptome (assessed with RNA-sequencing) in which there was a marked alteration of the immune gene expression signature. It is important to determine whether these gene expression changes relate to functional proteins. As such we will use proteomic approaches to determine the protein profile within the human cannabis-exposed placenta as well as placenta from our rat prenatal THC model. This project is a focus of our competitive renewal grant application (DA030359).

Technologies and consultations available from the Yale/NIDA Neuroproteomics Center’s Discovery and Targeted Proteomics Cores will significantly advance our efforts to characterize the complex landscape of post-translational histone modifications regulating exposure to drugs of abuse, as well as our work to explore protein complexes regulating these modifications. In addition, we will work in close collaboration with the Bioinformatics and Biostatistics Core of the Center.

Our laboratory has previously focused on gene expression, thus being part of the Yale/NIDA Neuroprotemics Center will be a unique opportunity for my postdoctoral fellows as well as PhD and MD/PhD students to advance their knowledge of proteomic approaches with respect to neurobiological aspects of addiction. I plan to have a number of my trainees not only learn proteomics, but participate actively in data analysis, which will be critical for their future development. As such my post-doctoral fellows, such as Anissa Bara PhD, who has already begun to learn about proteomics will receive MS/proteomics training at the Center and will attend the Center’s Research in Progress Meetings.