Cytoplasmic protein accumulations of the RNA/DNA binding protein TDP-43 are found in affected neurons in almost all instances of amyotrophic lateral sclerosis (ALS) and approximately 50% of frontotemporal dementia (FTD). Nuclear clearance of TDP-43 has been widely observed in affected neurons in sporadic ALS/FTD, evidence strongly supporting the proposal that TDP-43 loss of function is a key aspect of disease mechanism underlying ALS/FTD pathogenesis. We have identified that the mRNA encoding stathmin-2 is 1) an essential factor for axonal regeneration of axotomized iPSC-derived motor neurons and 2) the mRNA most affected by reduction in TDP-43 function, with a striking loss from motor neurons in sporadic ALS and inherited disease from GGGGCC expansion in C9orf72. Stathmin-2 is an abundant, direct binding partner of α/β-tubulin dimers in neuronal perikarya, axons, growth cones, and synapses, including neuromuscular junctions (NMJs). Using genome editing, we will identify the mechanism of TDP-43-dependent premature polyadenylation/cryptic splicing that suppresses stathmin-2 synthesis when TDP-43 levels are lowered. We will use genome editing of induced pluripotent stem cells (iPSCs) derived human motor neurons grown in compartmented chambers (that separate neuronal cell bodies, axons and growth cones) to determine how stathmin-2 functions in a) motor neuron maintenance/repair, b) axonal microtubule stabilization and/or dynamics, c) neuromuscular junction formation and/or stabilization, and d) how palmitoylation of stathmin-2 affects its axonal function(s). Genome wide CRISPR/Cas9 screens using flow cytometry and optical methods will be undertaken to identify factors that control stathmin-2 synthesis or accumulation. Finally, we will determine the consequences in mice of reduction or loss of stathmin-2 on motor neuron function and muscle innervation/denervation and whether reduction in stathmin-2 synergizes with TDP-43 mutation to drive motor neuron disease. Outcomes of these efforts will provide key insights for understanding basic aspects of axonal and synaptic neurobiology and for evaluating whether maintaining or restoring stathmin-2 is an attractive therapeutic option in sporadic ALS/FTD and ALS from its most frequent genetic cause, repeat expansion in C9orf72.