Inhibition of the deubiquitinase USP5 leads to c-Maf protein degradation and myeloma cell apoptosis

Abstract

The deubiquitinase USP5 stabilizes c-Maf, a key transcription factor in multiple myeloma (MM), but the mechanisms and significance are unclear. In the present study, USP5 was found to interact with c-Maf and prevented it from degradation by decreasing its polyubiquitination level. Specifically, the 308th and 347th lysine residues in c-Maf were critical for USP5-mediated deubiquitination and stability. There are five key domains in the USP5 protein and subsequent studies revealed that the cryptic ZnF domain and the C-box domain interacted with c-Maf but the UBA1/UBA2 domain partly increased its stability. Notably, MafA and MafB are also members of the c-Maf family, however, USP5 failed to deubiquitinate MafA, suggesting its substrate specificity. In the functional studies, USP5 was found to promoted the transcriptional activity of c-Maf. Consistent with the high level of c-Maf protein in MM cells, USP5 was also highly expressed. When USP5 was knocked down, c-Maf underwent degradation. Interestingly, USP5 silence led to apoptosis of MM cells expressing c-Maf but not MM cells lacking c-Maf, indicating c-Maf is a key factor in USP5-mediated MM cell proliferation and survival. Consistent with this finding, WP1130, an inhibitor of several Dubs including USP5, suppressed the transcriptional activity of c-Maf and induced MM cell apoptosis. When c-Maf was overexpressed, WP1130-induced MM cell apoptosis was abolished. Taken together, these findings suggest that USP5 regulates c-Maf stability and MM cell survival. Targeting the USP5/c-Maf axis could be a potential strategy for MM treatment.

RETRACTED: miR‐125a suppresses malignancy of multiple myeloma by reducing the deubiquitinase USP5

“…[19][20][21] As a member of the USPs family, USP5 can play the role of deubiquitination enzyme by hydrolyzing ubiquitin chain. 24,25 Wang et al 26 found that inhibiting the expression of deubiquitinase USP5 could lead to degradation of c-Maf protein and induce apoptosis of MM cells. 24,25 Wang et al 26 found that inhibiting the expression of deubiquitinase USP5 could lead to degradation of c-Maf protein and induce apoptosis of MM cells.…”

The transmembrane protein TMEPAI induces myeloma cell apoptosis by promoting degradation of the c-Maf transcription factor

“…In breast cancer, TMEPAI promotes breast cancer cell proliferation by converting TGF-β from a tumor suppressor to a tumor promoter (5), while in lung cancer cells, TMEPAI sequestrates R-SMAD and downregulates PTEN therefore enhancing the PI3K/AKT signaling pathway (3). In our present study, TMEPAI-induced MM cell apoptosis is probably associated with c-Maf ubiquitination and degradation because c-Maf promotes MM cell survival while silencing c-Maf leads to MM cell apoptosis (19). Moreover, TMEPAI is markedly downregulated in MM cells but highly expressed in bone marrow cells from healthy donors.…”

“…Because c-Maf is a transcription factor, we next evaluated to whether TMEPAI modulated c-Maf transcriptional activity. To this end, a luciferase construct directed by MARE (Maf recognition element) (19) was co-transfected with wild-type or ΔTM-TMEPAI. The luciferase activity analysis revealed that ΔTM-TMEPAI lost its modularity on the stability and transcriptional activity of c-Maf (Fig.…”

“…The NEDD4 gene was first cloned from the genomic DNA of MM cell line RPMI-8226 and it was then subcloned into pcDNA3.1 vector with a Myc tag. The luciferase reporter driven by Maf Recognition Element (MARE, 5'-TGCGAGTGAGGCA-3') (pGL4-MARE.Luci) was cloned as described previously (19). The PY motif mutants of TMEPAI were cloned by site-directed mutagenesis using the following primers: 5'-CCCCCACCCGCCCAGGGCCCCTG CACCCTCCAGCTT-3' (Forward) and 5'-GCCCTGGGCGGGTGGGGGCTCCTCC CCGTCTGACAG-3' (Reverse) for the PY1 mutant, 5'-CCGCCCACCGCCAGCGAGGTCAT CGGCCACTACCCG-3' (forward) and 5'-ACCTCGCTGGCGGTGGGCGGCGG CCCCTCCATGCG-3' (Reverse) for the PY2 mutant.…”

Discovery of small molecule antagonists of the USP5 zinc finger ubiquitin-binding domain

“…USP5 has been associated with tumorigenesis [15][16][17][18][19] , neurodegeneration [20][21][22] , and viral infections 23,24 . In addition, USP5 plays a role in regulation of ubiquitin levels in DNA damage repair 25 and stress granules 26 .…”