Aberrant function of pathogenic STAT3 mutant proteins is linked to altered stability of monomers and homodimers

Abstract

STAT3 mutations, predominantly in the DNA-binding domain (DBD) and Src-homology 2 domain (SH2D), cause rare cases of immunodeficiency, malignancy, and autoimmunity. The exact mechanisms by which these mutations abrogate or enhance STAT3 function are not completely understood. Here, we examined how loss-of-function (LOF) and gain-of-function (GOF) STAT3 mutations within the DBD and SH2D affect monomer and homodimer protein stability, as well as their effect on key STAT3 activation events, including recruitment to phosphotyrosine (pY) sites within peptide hormone receptors, tyrosine phosphorylation at Y705, dimerization, nuclear translocation, and DNA binding. DBD LOF mutants showed reduced DNA binding when homodimerized, while the DBD GOF mutants showed increased DNA binding; DBD LOF and GOF mutants showed minimal changes in other STAT3 functions or in monomer or homodimer protein stability. However, SH2D LOF mutants demonstrated reduced conformational stability as either monomers or homodimers, leading to decreased pY-peptide recruitment, tyrosine phosphorylation, dimerization, nuclear localization, and DNA binding. In contrast, cancer-causing SH2D GOF mutants showed increased STAT3 homodimer stability, which increased their DNA binding. Of note, a small-molecule inhibitor of STAT3 that targets the pY-peptide binding pocket within the STAT3 SH2D potently inhibited cell proliferation driven by STAT3 SH2D GOF mutants. These findings indicate that stability of STAT3 protein monomer and homodimer is critical for the pathogenesis of diseases caused by SH2D LOF and GOF mutations and suggest that agents that modulate STAT3 monomer and/or homodimer protein stability may have therapeutic value in diseases caused by these mutations.

Fluorinated and N-Acryloyl-Modified 3,5-Di[(E)-benzylidene]piperidin-4-one Curcuminoids for the Treatment of Pancreatic Carcinoma

“…This discovery has the potential to pave the way for the development of two different groups of anticancer-active curcuminoids depending on their phenyl substitution pattern. In addition, mutant STAT3 proteins might be addressed more efficiently, leading to an improved curcuminoid response in various cancer diseases [ 49 ]. EF24 and 2d also suppressed phospho-STAT3 levels in pancreatic cancer cells, corroborating the docking results obtained for these two curcuminoids.…”

“…Piperidin-4-one monohydrate hydrochloride (153 mg, 1.0 mmol) and 3-fluoro-4methoxydehyde (308 mg, 2.0 mmol) were dissolved in MeOH (10 mL) and NaOH (40 mg) in H 2 O (1 mL) was added. The reaction mixture was stirred at room temperature for 2 h. 16) [M + ], 355 (49), 210 (31), 169 (100).…”

Signal Transducer and Activator of Transcription Proteins at the Nexus of Immunodeficiency, Autoimmunity and Cancer

“…Several of the STAT inhibitors have shown effectiveness in vitro and in mouse tumor models [99]. A small molecule inhibitor that targets the phosphotyrosine-binding pocket of the STAT3 SH2 domain was able to block cell proliferation mediated by STAT3 GOF mutants [115]. However, the clinical reality has so far been that multiple therapeutics are typically required [41], or alternative strategies need to be employed, such as hematopoietic stem cell transplantation, including for patients harboring STAT1 GOF [57] or STAT3 GOF [69] mutations.…”

Asymmetric post-translational modifications regulate the nuclear translocation of STAT3 homodimers in response to leukemia inhibitory factor

“…In the context of STAT3, there are several situations where STAT3 dimers could be formed by two slightly different STAT3 monomers with functional consequences, just as it happens with asymmetric PTMs. Several gain-of-function and dominant negative mutations in only one STAT3 allele were associated to human pathologies, suggesting that normal and mutant forms of STAT3 can coexist, dimerize and function in many instances [ 21 , 22 ]. STAT3β is a non-pathogenic splicing isoform that coexist with the main STAT3α isoform (the one used in this study) with variable stoichiometry in different cells and tissues, also having consequences for human pathologies [ 23 , 24 ].…”