HDAC6-dependent deacetylation of SAE2 enhances SUMO1 conjugation for mitotic integrity.

Mammalian cells express three conjugatable SUMO variants: SUMO1 and the closely related SUMO2 and SUMO3 (together referred to as SUMO2/3). While some substrates are modified by both, others show a clear preference, though the basis for this selectivity remains unclear. Here, we examine a modification of the catalytic component of the human SUMO activation enzyme, SAE2. We find that lysine 164 of SAE2 undergoes HDAC6-dependent deacetylation during mitosis. A non-deacetylatable acetyl-mimetic mutant, SAE2-K164Q, selectively enhances SUMO2 over SUMO1 activation and conjugation, and distinguishes between SUMO1 and SUMO2/3 based on differences in their C-terminal tails. Complementation of SAE2-deficient or inhibited cells with SAE2-K164Q suppresses mitotic SUMO1 conjugation and promotes multipolar spindle formation. We identify NuMA as a SUMO E1-dependent substrate and demonstrate that mitotic defects caused by SAE2-K164Q or HDAC6 inhibition are rescued by SUMO1 overexpression or expression of a GFP-SUMO1-NuMA-K1766R fusion. These results support a model in which SAE1:SAE2 deacetylation during early mitosis promotes SUMO1 conjugation to ensure mitotic fidelity, highlighting a regulatory role for the SUMO-activating enzyme in the selection of SUMO proteins.