UC San Diego

Central nervous system neurons, like retinal ganglion cells, often fail to regenerate their axons and die following axonal injury. A complete understanding of cellular dynamics in injured neurons is necessary to identify critical regulators of degeneration and to develop therapies for survival and regeneration. Although extensive efforts have been made to dissect the transcriptomic and genetic changes in these neurons, less headway has been made into understanding protein dynamics, the effectors of cellular responses.

In this dissertation, I begin by reviewing the literature of regulators of neuronal regeneration and mechanisms underlying regenerative failure. Insights from disease point towards axonal transport failure as a unifying hypothesis underlying a portion of degenerative disease, yet methods of detecting protein transport are unsatisfactory. In fact, physiological protein transport in central nervous system in vivo was not well understood, surprising given the importance of protein transport for pre-synaptic regulation. Therefore we first developed a mass spectrometry-compatible methodology for detecting axonally-transported proteins (the “transportome”), using it to first catalogue the RGC transportome and then to probe why different sub-cortical RGC targets transduce different signal across their synapses. I next adapt this methodology to quantify changes in protein transport to the optic nerve after injury, identifying a significant reduction in transport of Kif5a, a protein necessary for RGC survival. I next adapt a method for quantifying new protein synthesis to the retina in two time points following optic nerve injury, generating candidate proteins for neurite outgrowth in vitro and RGC survival after injury in vivo. I conclude that the successful integration of multiple modalities of cellular response across time points after injury, including ChIP-sequencing, RNA-sequencing, protein synthesis, degradation, transport, and maintenance, will give a more complete understanding of dynamic cellular regulation in degeneration and provide key therapeutic targets for survival and axon regeneration.