Understanding Binding Partner Association Interactions with Unphosphorylated STAT3

Abstract

The transcription factor Signal transducer and activator of transcription 3 (STAT3) enters the cell nucleus to regulate critical immune and metabolic functions. STAT3 exists in two distinct forms: phosphorylated STAT3 (pSTAT3) and unphosphorylated STAT3 (uSTAT3). pSTAT3 mediates normal cytokine signaling, while uSTAT3 promotes persistent inflammation linked to various cancers. Therefore, selectively inhibiting nuclear import of uSTAT3 while sparing pSTAT3 represents a promising therapeutic strategy. Unfortunately, uSTAT3 continually switches between a monomeric and moderately uncharacterized dimeric state. Furthermore, the interface between STAT3 and its nuclear transporters remain poorly understood. We used AlphaFold3 to predict the structures of uSTAT3β monomers and dimers, both free in solution and bound to their primary transporter complex (importin-α3+β). Our models revealed a novel architecture for unbound dimeric uSTAT3 featuring an antiparallel orientation of its DNA-binding domains (DBDs) away from the dimer interface, in contrast to existing structural models. Additionally, we propose that the monomeric and dimeric structures share an identical binding interface, with contacts occurring at the beta-sheets within the DBD and/or other signaling domains. Our models enable the design of peptides that can mimic the interface of importin-α3, inhibiting uSTAT3’s ability to bind the nuclear transport complex. Identification of active peptides offers a novel approach to treat diseases driven by aberrant uSTAT3 activity while minimizing disruption of canonical pSTAT3 signaling.