Martin Riccomagno - US grants

PROJECT SUMMARY While numerous transgenic tools and approaches exist to enable manipulation of gene expression in many cell types in the healthy brain, tools designed to target and study cells present only in the dis- eased or damaged brain are lacking. Common to virtually all neurodegenerative diseases, brain injuries and infections is a neuroinflammatory and immune response characterized by changes in astrocytes, which become ?reactive?. Astrocytes ordinarily provide critical support for neurons and only turn into reactive astrocytes (RAs) in brain disease and inflammation. A longstanding issue which has remained unknown is whether RAs contribute to or help alleviate disease progression. The objective of this appli- cation is to deliver a new combinatorial transgenic strategy and toolkit to specifically target RAs in dis- ease. This toolkit will enable researchers to selectively alter (eliminate, increase, or decrease) gene ex- pression only in RAs at any point in the progression of brain disease and inflammation. Brain infection by the parasite Toxoplasma gondii will serve as a model of brain inflammation stemming from infection. Three aims are proposed: In Aim 1, we will first characterize the Cre transgenic strategy to selectively manipulate gene expression only in RAs in brain disease. In Aim 2, we will then use the new approach to selectively ablate, prevent, or reprogram RAs back into non-reactive astrocytes at various stages during the acute and chronic inflammatory process. Our work will provide new information on the role of reactive astrocytes in the early vs. sustained stages of brain inflammation. In Aim 3, we will perform ?translatome? analysis to identify genes uniquely altered in reactive astrocytes during brain inflamma- tion for the first time, providing novel targets for future study. Our innovative approach will allow detec- tion of both inductions and reductions in gene expression with unprecedented signal-to-noise over ex- isting approaches. The rationale for the proposed research is that improving understanding of the cellu- lar and molecular mechanisms of brain disease will provide novel insights into the development of more effective treatments. We anticipate that this research will be transformative, as we will introduce to the research community a powerful new strategy to investigate the role of reactive cell types in any disease or disorder of the nervous system.

The proposed studies will identify mechanisms through which Cas adaptor proteins regulate cortical development and function. Accurate cell migration and lamination are indispensable for the development of a functional nervous system. Congenital disruption of any of these processes can result in neurodevelopmental disorders ranging from lissencephaly to autism-spectrum disorders. During multiple developmental steps, neurons are guided by repulsive and attractive cues, while actively interacting with the extracellular matrix (ECM) and other cells. Although much is known about the ligand-receptor pairs required for these cues to regulate cell trajectories, it remains unknown how instructive and adhesive cues are integrated and interpreted within the neuron. The Cas (Crk associated substrate) family of cytosolic adaptor proteins are known to signal downstream of several neural guidance cues, and regulate focal adhesion turnover. Using Cas proteins as a model, we have a unique opportunity to broaden our understanding of how instructive and permissive signaling pathways converge during neural development to regulate cell adhesion. To test the hypothesis that Cas proteins are essential for coordinating cortical lamination and migration, experiments will be organized into three Specific Aims. Guided by strong preliminary data showing that ablation of Cas genes results in cortical defects that resemble cobblestone lissencephaly, Aim 1 will establish, for the first time, the functional requirement for Cas proteins during cortical migration and lamination. Aim 2 will explore the contributions of Cas-mediated cortical scaffold formation to physiology and behavior, using EEG and behavioral approaches. Aim 3 will test how regulation of Cas function affects cortical lamination. These experiments will provide new knowledge on the basic mechanisms underlying the establishment of neural circuits essential for perception and cognition.