PROJECT SUMMARY/ABSTRACT
Damage to the peripheral nervous system (PNS) can be incurred through a wide spectrum of conditions
including trauma, metabolic imbalances, and chemotherapy. Neuronal injuries occur when their long and
fragile processes (i.e. axons) are damaged, for example by stretching or severing. Compared to PNS neurons,
the regenerative capacity of injured central nervous system (CNS) neurons is extremely limited, typically
resulting in life-long disability. Regenerative growth of axons can occur in the injured peripheral nervous
system, however, restoration of function is often incomplete, resulting in loss of motor and sensory function
and frequently the development of neuropathic pain.
Regeneration of an injured axon is thought to involve changes in microtubule dynamics and post-
translational modifications (PTMs). Microtubules are dynamic polymers of a,ß-tubulin and are the core
structural element of axons and dendrites. Microtubules also serve as tracks for molecular motor proteins
(kinesin, cytoplasmic dynein) that transport various cargos between synaptic terminals and the neuronal cell
soma. Following injury, microtubules undergo rapid fragmentation, a process known as Wallerian
degeneration. In developing neurons and during axon regeneration, microtubules in the neuronal growth cone
and axon shaft play key roles in axon guidance and axon elongation.
Regulation of MT function is largely achieved by the differential expression of a- and ß-tubulin isotypes,
and by PTMs of tubulin subunits within the microtubule polymer. Studies of microtubule isotypes and PTMs
inherently rely on antibodies that require fixation and permeabilization of the cell or tissue, which severely limits
our understanding of the spatiotemporal component of tubulin PTMs. We have developed a pipeline that allows
us to generate live-cell probes for tubulin PTMs. Here, we will generate probes for tubulin isotypes and PTMs
relevant to neuronal degeneration and regeneration and use them to map the spatiotemporal patterns of
changes to the microtubule cytoskeleton following injury to the PNS and CNS.
