TRIM28 Serves As a Key Regulator of Protein Homeostasis and Mediates Proteasome Inhibitor Resistance in Multiple Myeloma

Introduction:

The maintenance of intracellular protein homeostasis is a critical biological process necessary for the survival of multiple myeloma (MM) cells. Tripartite motif (TRIM) proteins, a subfamily of the RING-type E3 ubiquitin ligase family, are integral to protein quality control. In this study, we examined the role of TRIM28, a member of the TRIM family, as a regulator of protein homeostasis in MM.

Methods:

Bioinformatic analysis was utilized to elucidate the association between TRIM28 and genes involved in the proteasome and autophagy mediated protein degradation pathway. ShRNA knockdown (KD) and overexpression (OE) experiments were conducted to investigate the role of TRIM28 in MM pathogenesis. ChIP-seq and IP-MS analyses were performed to identify potential downstream targets of TRIM28.

Results:

Firstly, our survival analysis revealed that MM patients exhibited poor outcomes with high levels of TRIM28 in both the MMRF-CoMMpass and in-house datasets, particularly in the context of bortezomib treatment. Overexpression of TRIM28 promoted cell proliferation and induced drug resistance in MM cell lines, while knock-down of TRIM28 had the opposite effect. Bioinformatic analysis demonstrated a strong correlation between TRIM28 expression and the expression of proteasome subunits and autophagy-related genes in MM datasets. RNA-seq data showed down-regulation of both the proteasome pathway and autophagy pathway after TRIM28 knockdown, leading to decreased proteasome activities and accumulation of ubiquitinated proteins. Immunofluorescence assay confirmed nuclear localization of TRIM28, while ChIP-seq identified its binding to proteasome gene promoters such as PSMB1, PSMD2, and PSMD4. Knock-down of TRIM28 reduced the expression of proteasome subunits, indicating its critical role in transcriptional activation of these genes in MM cells.

Additionally, a decrease in autophagosome formation was observed in TRIM28 KD cells, indicating the involvement of TRIM28 in autophagy activity. Further IP-MS and Co-IP assays demonstrated the interaction between TRIM28 and 14-3-3ζ. A significant reduction in the polyubiquitination level of 14-3-3ζ was noted in TRIM28 KD cells. The 14-3-3ζprotein serves as an adaptor that modulates cellular signaling by binding to a wide array of proteins. Therefore, acting as an E3 ligase, TRIM28 promotes ubiquitin-dependent degradation of 14-3-3ζ, thereby enhancing autophagy activity in MM. These findings underscore the crucial role of TRIM28 in maintaining protein homeostasis by regulating both proteasome and autophagy activities involved in myeloma pathogenesis.

Conclusion:

Our research has provided new and important insights into the crucial role of TRIM28 in maintaining protein homeostasis and its involvement in myeloma pathogenesis. TRIM28 activates the proteasome and autophagy pathways, making it a potential promising therapeutic target in MM.

No relevant conflicts of interest to declare.