Methods for addressing the protein-protein interaction between histone deacetylase 6 and ubiquitin

Histone deacetylase 6 (HDAC6) is a cytoplasmic HDAC isoform able to remove acetyl groups from cellular substrates such as α-tubulin. In addition to the two deacetylase domains, HDAC6 has a C-terminal zinc-finger ubiquitin (Ub)-binding domain (ZnF-UBP) able to recognize free Ub. HDAC6-Ub interaction is key in regulating the elimination of misfolded proteins during stress response through the aggresome pathway. Small molecules inhibiting deacetylation by HDAC6 were shown to reduce aggresomes, but the interplay between HDAC6 catalytic activity and Ub-binding function is not fully understood. Here we describe two methods to measure HDAC6-Ub interaction in vitro using full-length HDAC6. Our results suggest a potential role for the HDAC6 deacetylase domains in modulating HDAC6-Ub interaction. Both methods were effective for screening inhibitors of the HDAC6-Ub protein-protein interaction devoid of catalytic activity. This new HDAC6 modulation mechanism can be targeted to address the function of HDAC6-Ub interaction in normal and disease conditions.