RhoGDIα is a key regulator of Rho proteins, coordinating their GTP/GDP and 
membrane/cytosol cycle. Recently, it was demonstrated by quantitative mass 
spectrometry that RhoGDIα is heavily targeted by post-translational lysine 
acetylation. For one site in its N-terminal domain, namely K52, we reported 
earlier that acetylation completely switches off RhoGDIα function. Herein we 
show that K52-acetylated RhoGDIα is specifically deacetylated by the sirtuin 
deacetylase Sirt2. We show that acetylation at K52 decelerates cervical cancer 
cell proliferation, suggesting RhoGDIα acetylation to be a promising therapeutic 
target. We demonstrate that treatment of cervical cancer cells with a 
RhoGDIα-derived K52-trifluoroacetylated, substrate-derived peptidic sirtuin 
inhibitor severely impairs cell proliferation. Finally, we conclude that the 
potency of substrate-derived sirtuin inhibitors depends on structural features, 
the substrate-derived amino acid sequence as a determinant for selectivity, as 
well as the presence of an acetyl-lysine analogue to increase its potency. These 
data reveal a prospective therapeutic potential for novel substrate-derived 
sirtuin inhibitors.