SERPINB3 enhances NPM1 sumoylation via inhibiting SENP3's activity and promotes lung tumorigenesis.

SERPINB3 or SCCA1, a member of the serine proteinase inhibitor family, is frequently overexpressed in multiple malignancies, including lung adenocarcinoma (LUAD). However, its molecular mechanisms and endogenous substrates in lung cancer remain poorly characterized. Here, we identified SUMO-specific proteinase 3 (SENP3) as a novel direct interaction partner and functional target of SERPINB3 in LUAD. Using proximity-dependent biotin labeling coupled with mass spectrometry, we discovered that SERPINB3 forms a nuclear complex with SENP3 and nucleophosmin (NPM1). This tripartite interaction was validated through endogenous co-immunoprecipitation and bimolecular fluorescence complementation assays. Structural modeling with AlphaFold3 and subsequent mutagenesis studies revealed that SERPINB3 binds specifically to the N-terminal region of SENP3 (amino acids 265-287). Furthermore, our structural and biochemical analyses demonstrate that SENP3 specifically interacts with the C-terminal aromatic domain of NPM1 (amino acids 242-294), while SERPINB3 selectively binds to the pentameric form of NPM1. Mechanistically, SERPINB3 inhibits SENP3's desumoylation activity, leading to enhanced NPM1 sumoylation in the nucleolus. Functional studies demonstrated that SERPINB3 overexpression (but not the RCL-deleted mutant, SERPINB3△6) promotes LUAD cell proliferation and tumor growth, with NPM1 sumoylation being critical for this oncogenic effect. Clinically, SERPINB3 is highly expressed in human LUAD specimens and co-localizes with SENP3 and NPM1. Our work establishes SENP3 as a bona fide endogenous substrate of SERPINB3 and delineates a novel sumoylation-dependent oncogenic axis in LUAD, offering potential therapeutic targets for this malignancy.