Interactome Mapping: Using Protein Microarray Technology to Reconstruct Diverse Protein Networks

Uzoma, Ijeoma; Zhu, Heng

In a follow up study, Li et al. took the inverse approach that employed a herpesvirus EBV protein microarray to assess human-host protein binding events [14] . Small ubiquitin-related modifier (SUMO) is covalently attached to proteins via an enzymatic cascade analogous to the ubiquitin pathway. SUMO is involved in a broad range of cellular processes including signal transduction, regulation of transcription, DNA damage response and mediation of protein-protein interactions [43, 44] . Both latent and lytic EBV proteins interact with components of the SUMO machinery [14, 44] . While covalent modification by SUMO is more commonly understood, noncovalent interactions with SUMO also contribute to SUMO effector signaling [43, 44] . Noncovalent binding to SUMO is often mediated through SUMO-interaction motif (SIM) domains on target proteins [43, 44] . To comprehensively identify the EBV proteins that bind to the SUMO moiety, the authors fabricated a protein microarray of full length proteins from EBV and KSHV individually purified from yeast. The array was used to perform a protein-protein binding assay using the SUMO2 paralog. They identified 11 EBV proteins as potential SUMO partners, including BGLF4, a conserved kinase [14] . As BGLF4 is known to play a multitude of roles in EBV, the authors pursued the importance of the cellular PTM in BGLF4 function. The BGLF4 SIM domains were mapped and when mutated at both the N-and C-terminal SIMs, the intracellular localization of the kinase shifted from nuclear to cytoplasmic. A mutation in the N-terminal SIM showed largely nuclear localization, whereas the C-terminal SIM mutation generated an intermediate phenotype with nuclear and cytoplasmic expression. The authors found that BGLF4 inhibits SUMOylation of lytic cycle transactivator ZTA and demonstrated that the SIM domains as well as kinase activity are required for inhibition [14] . SIM domains of BLGF4 were also shown to be necessary for suppressing global SUMOylation, inducing cellular DDR and promoting EBV lytic replication.