Aberrant aggregation of the prion-like RNA-binding protein TDP-43 drives several fatal neurodegenerative proteinopathies, including amyotrophic lateral sclerosis (ALS). Here, we define how short, specific RNAs solubilize TDP-43. These short RNAs engage and stabilize the TDP-43 RNA-recognition motifs, which allosterically destabilizes a conserved helical region in the prion-like domain, thereby promoting aggregation-resistant conformers. Sequence-space mining identified short RNA chaperones with enhanced activity against TDP-43 and disease-linked variants. Enhanced short RNA chaperones mitigate aberrant TDP-43 phenotypes in optogenetic models and in ALS patient–derived and control motor neurons. In mice experiencing cytoplasmic TDP-43 aggregation and motor neuron loss, an enhanced short RNA chaperone reduces pathological aggregation, restores TDP-43 function, and confers neuroprotection. These findings define a mechanistic and therapeutic framework for RNA-based strategies to counter TDP-43 proteinopathies.