Parkinson’s disease (PD) is a progressive neurodegenerative disorder characterised by midbrain dopaminergic neuron degeneration and intracellular a-synuclein (aSyn) accumulation. Here we show that AAV-shRNA-mediated inhibition of HDAC5 protected against aSyn-induced neurodegeneration in vitro and in vivo. We also show that AAV-mediated overexpression of aSyn induces HDAC5 nuclear accumulation in dopaminergic neurons in the rat substantia nigra in vivo, and in SH-SY5Y cells in vitro. To investigate the nuclear HDAC5 complex induced by aSyn, we used a co-immunoprecipitation proteomics approach to identify HDAC5-interacting proteins by overexpressing a nuclear-restricted HDAC5 protein (HDAC5-nuc) in SH-SY5Y cells. We found that HDAC5-nuc specifically interacted with 6 individual proteins. To examine whether inhibition of components of this nuclear HDAC5-protein complex had beneficial effects, we measured the effect of shRNA-mediated inhibition of HDAC5-interacting proteins on neurite growth in SH-SY5Y cells overexpressing HDAC5-nuc or aSyn. The results show a potential HDAC5-mediated pathological mechanism in an in vitro model of PD, rationalising further study of nuclear HDAC5 complex as a potential therapeutic target for neuroprotection in PD.