Mantle cell lymphoma (MCL) is considered one of the most aggressive B-cell lymphoid neoplasms. The transcription factor SOX11 is aberrantly expressed in conventional aggressive MCL but is either negative or very weakly expressed in the leukemic non-nodal MCL subtype, which has a predominantly indolent clinical evolution. SOX11 is a key MCL driver through the regulation of several oncogenic mechanisms, suggesting that it forms additional protein complexes to exert multiple actions.
To understand how SOX11 regulates different specific target genes and signaling pathways in MCL, we employed a proteomic strategy to identify SOX11 interactions. By over-expressing SOX11 fused to the modified biotin ligase BioID2 and performing streptavidin-based biotinylated-protein purification coupled with LC-MS/MS, we identified a SOX11-specific interactome of 92 proteins in MCL, mainly involved in nucleosome disassembly, chromatin organization, and transcriptional regulation. Among the highest confident SOX11-partner proteins, several components of the SWItch/Sucrose NonFermentable (SWI/SNF) chromatin remodeling complex were identified.
Using co-immunoprecipitation and in situ fluorescence proximity labeling assays, we validated the physical interaction of endogenous SOX11 with SMARCA4, the catalytic subunit of the SWI/SNF chromatin remodeling complex, in MCL cell lines and primary tumors. Moreover, the superimposition of the three-dimensional structures of SOX11 and SMARCA4 within the human nucleosome core complex indicated that SOX11 binds to nucleosome-DNA near the SMARCA4-ATPase domain, suggesting they physically interact within the nucleosome, enabling access to chromatin to modulate transcription.
We observed significantly higher expression of SMARCA4 in SOX11-positive cMCL than in SOX11-negative nnMCL tumors, with a significant positive correlation between SMARCA4 and SOX11 expression in MCL primary samples. Additionally, SOX11 bound to SMARCA4 promoter only in SOX11-positive MCL cell lines. SOX11 knockout downregulated SMARCA4 expression, suggesting it directly regulates the transcription of SMARCA4. Moreover, high SMARCA4 expression was significantly associated with shorter overall survival in MCL patients, independently of other high-risk factors, such as SOX11 expression, high copy number alterations, or TP53 alterations, suggesting its involvement in the progression of this disease.
Integration of genome-wide SOX11-specific and SMARCA4-specific DNA-binding sites, identified by Cleavage Under Targets & Release Using Nuclease (CUT&RUN) experiments, with transcriptomic profiles by RNA-sequencing revealed that 58.84% of the SOX11-specific binding regions are co-occupied by SMARCA4. The SOX11:SMARCA4 complex binds mainly to open chromatin regions enriched in active promoters, directly regulating the expression of essential genes for MCL progression and aggressiveness.
The BCR-signaling emerged as the most significant pathway regulated by the SOX11:SMARCA4 complex. The AU-15330, a PROTAC-degrader of SMARCA4, significantly reduced the binding of SOX11 to its specific regulatory regions, reduced BCR-signaling activation and cell proliferation, and induced apoptosis in SOX11-positive/SMARCA4-high cells, but not in the JVM2 SOX11-negative MCL cell line, which also express low levels of SMARCA4. AU-15330 treatment also reduced the expression of genes of other oncogenic pathways, such as the alternative NF-kB pathway and the anti-apoptotic BCL2-signaling of both Ibrutinib-resistant and -sensitive SOX11-positive cell lines, but not in JVM2. These results suggest that AU-15330 may offer a promising therapeutic strategy for MCL patients relapsing from Ibrutinib and potentially other therapies.
Overall, we have identified the SOX11-specific interactome and demonstrated the physical interaction of SOX11 and SMARCA4 in MCL cells. SMARCA4 upregulation, directly regulated by SOX11, is associated with poor MCL patient outcomes, highlighting its role in MCL pathogenesis. SOX11:SMARCA4 complex commonly regulates the expression of critical genes for MCL, and the disruption of this complex could represent an innovative therapeutic strategy to overcome resistance and relapse to current therapies in MCL patients.
No relevant conflicts of interest to declare.
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