Homeostatic control of neuronal activity, which maintains activity levels within a functional range, is critical for the stability of neuronal circuits and normal brain functions. Yet the underlying mechanisms governing homeostatic control of neuronal activity levels remain largely unknown. Emerging evidence suggests that proteostasis, especially the synthesis and degradation of activity-induced newly-synthesized proteins, may be involved in the regulation of activity homeostasis in neurons. The recent discovery and initial characterization of a neuronal membrane specific proteasome (NMP) suggests that NMPs preferentially degrade activity-induced newly-synthesized proteins and that NMP function influences neuronal activity. Together, these in vitro studies suggest a potential protein-degradation based regulatory mechanism for neuronal homeostasis, however evidence for in vivo functions of the NMP are lacking, particularly whether the NMP degrades newly-synthesized proteins, whether NMP activity affects neuronal activity levels and circuit function and whether in vivo NMP substrates can be identified. Here, we propose to investigate the function of the NMP in vivo in the visuomotor circuit of Xenopus laevis tadpoles. First, we will determine whether NMPs are present in optic tectal neurons of tadpoles and assess NMPs proteolytic activity for activity-induced newly-synthesized or pre-existing proteins in vivo. Then we will examine the interaction between NMP activity and neuronal activity using in vivo functional Ca imaging, and the influence of NMP function on visuomotor behavioral plasticity in intact animals. Finally, we will quantify changes of individual proteins to identify NMP substrates under basal and stimulated conditions. Results of these studies will begin to establish NMP functions in vivo and to assess the potential function of NMP-mediated protein degradation in the maintenance of neuronal and circuit homeostasis.
Protein degradation is required for healthy function of brain cells. A thorough understanding of the cellular and molecular machinery neurons employ for protein degradation will further our basic knowledge of the mechanisms ensuring normal brain function and facilitate studies of their malfunction under diseased conditions.
