The Dependency Map (DepMap) is a genome-wide pooled
CRISPR-Cas9 knockout proliferation screen conducted in more than 700 cancer cell lines spanning many
different tumor lineages. Each cell line in the DepMap contains a unique barcode, and each gene
knockout is assigned a “dependency score” on a per cell-line basis which quantifies the rate of
CRISPR-Cas9 guide drop. It has been found that proteins with similar DepMap scores across cell
lines, a phenomenon known as co-dependent genes, have closely related biological functions. This can
include activity in the same or parallel pathways or membership in the same protein complex or the
same pathway.
We identified the strongest seven co-dependent genes (“Symbol”) for DUBs and ran GO enrichment
analysis. We used Biogrid, IntAct, and Pathway Commons PPIDs, and the NURSA protein-protein
interaction databases (PPIDs) to determine whether co-dependent genes interact with one another. The
“Evidence” column contains the PPIDs in which the interaction appears as well as whether there is
support for the association by an INDRA statement. As another approach to identify potential
interactors, we looked at proteomics data from the Broad Institute's Cancer Cell Line Encyclopedia (CCLE) for
proteins whose expression across ~375 cell lines strongly correlated with the abundance of each DUB;
it has previously been observed that proteins in the same complex are frequently significantly
co-expressed. The correlations and associated p-values in the CCLE proteomics dataset are provided.
And, we determined whether co-dependent genes yield similar transcriptomic signatures
in the Broad Institute's Connectivity
Map (CMap). A CMap score greater than 90 is considered significantly similar.
Using the biological processes and other Gene Ontology terms from well characterized DUBs as a
positive control, several gene set enrichment analyses were considered. Threshold-less methods
like GSEA had relatively poor results.
Over-representation analysis with a threshold of of the top 7 highest absolute value Dependency Map
correlations yielded the best results and is reported below.
There were too few differentially expressed genes to run a meaningful GSEA.
Literature Mining
INDRA was used to automatically assemble known mechanisms
related to USP22 from literature and knowledge bases.
The first section shows only DUB activity and the second shows all other results.
Deubiquitinase Activity
psp
cbn
pc
bel_lc
signor
biogrid
lincs_drug
tas
hprd
trrust
ctd
vhn
pe
drugbank
omnipath
conib
crog
dgi
|
rlimsp
isi
tees
geneways
eidos
trips
medscan
sparser
reach
As we discussed previously, USP22 is a component of a transcriptional activator complex SAGA and can deubiquitinate histones H2A and H2B, as well as several other substrates (Zhang et al., 2008a, b).
However, de-ubiquitylation of H2B by Usp22, the human homolog of yeast Ubp8, inhibits heterochromatic silencing and promotes gene activation [XREF_BIBR, XREF_BIBR].
The requirement of ATXN7L3 for H2B deubiquitination by USP22, USP27x, and USP51 suggests that all three use the ATXN7L3 zinc finger to dock the H2A and H2B acidic patch in a manner similar to that shown in the structure of the yeast DUB module bound to ubiquitinated nucleosomes.
As a subunit of hSAGA, USP22 participates in the deubiquitination of histones H2A and H2B and the acetylation of histone H4 to regulate gene transcription and expression.
Ubiquitin specific protease 22 (USP22) reduced Sirt1 ubiquitination and degradation and decreased FN and TGF-beta1 expression in GMCs under both basal and AGEs treated conditions.
For instance, HULC can upregulate the expression of the ubiquitin specific peptidase 22 (USP22) protein by suppressing miR-6825-5p, miR-6845-5p, and miR-6886-3p at the epigenetic or transcriptional level in HCC cells; USP22 enhances the HULC induced deubiquitination of Sirt1 and stabilizes it, and Sirt1 stability induces the autophagy of HCC cells, thus increasing the resistance of HCC cells to oxaliplatin [XREF_BIBR].
In addition, USP22 knockdown prevented c-MYC-mediated reduction of SIRT1 ubiquitination (XREF_FIG) and increase in SIRT1 expression (XREF_SUPPLEMENTARY).
Collectively, USP22 might deubiquitinate SIRT1 and subsequently activate the AKT pathway, increasing the expression of MRP1 to induce MDR in HCC cells.
Given that SIRT1 is deubiquitinated by USP22 and stabilized at the protein level (Armour etal., 2013; Lin etal., 2012) and previous studies have reported that SIRT1 could negatively influence the chemosensitivity of HCC cells (Chen etal., 2012), our results supported the notion that USP22 increases SIRT1 protein levels in HCC cells.
USP22 can deubiquitinate Sirt1 and enhance its stability through c-MYC-related network, leading to FLT3 tyrosine kinase inhibitors (TKIs) resistance in acute myeloid leukemia (AML).
Moreover, using in vitro deubiquitination assay, we found that KDM1A ubiquitination was decreased by incubating with recombinant USP22, suggesting that USP22 deubiquitinates KDM1A directly (XREF_FIG).
Given the known regulation of TRF1 by ubiquitination, we hypothesized that the SAGA complex might facilitate Usp22 dependent deubiquitination of TRF1 and that loss of Gcn5 might alter this activity by compromising complex integrity.
Additionally, Usp22 directly deubiquitinates TRF1 (TBP (TATA box binding protein)-related factor 1) to regulate the transcription of cell cycle and apoptosis genes [XREF_BIBR] and inhibits the transcriptional activity of p53 by deubiquitinating SIRT1 histone deacetylase [XREF_BIBR] and by regulating MDMX stability [XREF_BIBR].
GCN5 and USP22 also protect telomeres from DNA damage response through the stabilization of a shelterin component called TRF1, and interestingly, this regulation is not transcriptional but involves USP22 mediated deubiquitination of TRF1 [XREF_BIBR].
As a subunit of hSAGA, USP22 participates in the deubiquitination of histones H2A and H2B and the acetylation of histone H4 to regulate gene transcription and expression.
Several DUBs have been implicated in histone deubiquitination, including USP3, USP12, USP22, and USP46, which deubiquitinate both histones H2A and histones H2B [XREF_BIBR].
As we discussed previously, USP22 is a component of a transcriptional activator complex SAGA and can deubiquitinate histones H2A and H2B, as well as several other substrates (Zhang et al., 2008a, b).
The deubiquitinase catalytic activity of USP22 is required for CCNB1 deubiquitination because the catalytically inactive USP22 (USP22 and C185A) mutant failed to suppress CCNB1 ubiquitination without affecting its interaction with CCNB1 (XREF_SUPPLEMENTARY).
Mechanistically, USP22 deubiquitinates EGFR localized on late endosomes, prevents ubiquitination mediated EGFR degradation and enhances recycling of EGFR after EGF stimulation.
Additionally, USP22 sustained the activation of multiple EGFR downstream signaling pathways, including STAT3, AKT/mTOR and MEK/ERK pathways, in lung ADC cell lines H1975 and PC9.
In this model, late endosome localized USP22 deubiquitinates EGFR and impedes sorting of EGFR to the lysosome, thus sustaining the trafficking of EGFR to the plasma membrane (Figs. 6 and 9).
To test whether the endogenous BMI1 is also deubiquitinated by USP22 in glioma cells, we knocked down endogenous USP22 in U251 cells pretreated with CHX and found that endogenous BMI1 also became unstable and degraded rapidly.
USP22 also deubiquitinates non histone proteins, including telomeric repeat binding factor 1 (TRF1), sirtuin 1 (SIRT1), cyclin B1 and others, leading to protein stabilization by preventing proteasome mediated degradation.
It could be through a direct mechanism where USP22 deubiquitylates c-MYC inducing its stabilization and activation, or indirectly through ubiquitin removal from histones at c-MYC target genes, recruitment of other transcriptional machinery or deubiquitylation of proteins important for c-MYC activity.
Gao et al found that USP22 interacts with and deubiquitinates NFATc2, and also stabilizes NFATc2 protein and promotes NFATc2 function to facilitate IL-2 expression in T cells.
Additionally, Usp22 directly deubiquitinates TRF1 (TBP (TATA box binding protein)-related factor 1) to regulate the transcription of cell cycle and apoptosis genes [XREF_BIBR] and inhibits the transcriptional activity of p53 by deubiquitinating SIRT1 histone deacetylase [XREF_BIBR] and by regulating MDMX stability [XREF_BIBR].
Deubiquitination of H2B by Ubp8 and USP22 results in the recruitment of Ctk1 leading to Ser2 phosphorylation, a modification associated with elongation.
These findings confirm that USP22 could deubiquitinate H2AX and promote its phosphorylation, thus contributing to DNA damage repair and inducing cisplatin resistance.
In non-small cell lung cancer (NSCLC), aberrant expression of USP22 is a predictor of poor survival, as is high expression of cyclooxygenase-2 (COX-2).
Several studies demonstrating that USP22 can deubiquitinate uH2B in vitro and might affect the transcriptional elongation on the IRF1 gene (interferon regulatory factor 1) suggested a possible mechani[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]
A recent study showed that USP22‐ mediated deubiquitination of Hairy and Enhancer of split 1 (Hes1) is important for neuronal differentiation in the developing brain
USP22 was found to hydrolyze monoubiquitin tagged to uH2A and to antagonize PcG or hydrolyze monoubiquitin from uH2B to regulate MLL-trithorax-mediated trimethylation of histone H3 lysine-4
Deubiquitination of H2B by Ubp8 and USP22 results in the recruitment of Ctk1 leading to Ser2 phosphorylation, a modification associated with elongation.
In diabetic nephropathy, the increased expression of USP22 reduced the Sirt1 ubiquitination and degradation, and decreased fibronectin and TGF-beta1 expression in glomerular mesangial cells under both[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]
To determine whether the six EZH2 interacting DUBs affect EZH2 ubiquitination and protein levels, we transfected them individually into HEK293T cells, and we found that USP22 and ZRANB1 decreased the polyubiquitination of EZH2 (XREF_SUPPLEMENTARY); however, only ZRANB1 upregulated endogenous EZH2 protein (XREF_SUPPLEMENTARY).
While the role of altered STAGA and USP22 deubiquitination complex function in SCA7 disease pathogenesis is unclear, recent studies of the related polyQ disorder SCA1 indicate that the polyQ expansion in ataxin-1 attenuates the formation and function of the Capicua transcription factor complex, contributing to SCA1 disease pathogenesis through a partial loss-of-function mechanism.
Loss of Usp22 in Treg reduces Foxp3 transcript levels, increases FOXP3 ubiquitination and degradation, and reduces suppressive activity in vivo in mice.
Knockdown of USP22 significantly inhibited the viability of CAL-62 and 8505C cells, and impaired proliferation as revealed by reduced EdU incorporation in CAL-62 and 8505C cells.
Silencing of USP22 downregulated COX-2, decreased its half-life, and inhibited lung carcinoma cell proliferation by directly interacting with and modulating the stability and activity of COX-2 through the regulation of its ubiquitination status.
XREF_BIBR - XREF_BIBR Zhang and colleagues have demonstrated that ectopic overexpression of USP22 promotes cell proliferation and that suppression of USP22 expression by small hairpin RNA induces cell cycle arrest in human lung cancer cells.
iv ) The downregulation of USP22 not only suppresses multidrug resistance-associated protein 1 ( MRP1 ) , enhances the intracellular accumulation of sorafenib , and finally inhibits cell proliferation and cancer angiogenesis , but also inhibits glycolysis in hepatocellular carcinoma ( HCC ) cells , enhancing sorafenib chemosensitivity and impairing cell metabolism .
Silencing USP22 in chemoresistant HCC Bel/Fu cells dramatically inhibited proliferation, migration, invasion and epithelial-mesenchymal transition in vitro; suppressed tumorigenic and metastatic capacities in vivo; and inhibited drug resistance related proteins (MDR1, LRP, MRP1).
A recent study suggested that USP22 could increase TGF-β expression and promote the epithelial–mesenchymal transition (EMT). xref In addition, another study demonstrated that silencing USP22 could inhibit proliferation and induce cell cycle arrest in bladder cancer cells. xref Furthermore, USP22 was also reported with high expression level in different malignancies, such as breast cancer xref and colorectal cancer. xref Thus, these lines of evidence strongly suggested that the oncogenic role of the USP22 might contribute to progression and predict the prognosis and become an attractive therapeutic target in cancers.
The assay results showed that USP22 downregulation significantly inhibited the proliferation (XREF_FIG) and invasion (XREF_FIG) of U2OS and MG-63 cells.
Additionally, investigation into the underlying mechanism, using small interfering RNA, revealed that the downregulation of USP22 inhibited proliferation and promoted apoptosis though the phosphoinositide 3-kinase/protein kinase B signaling pathway.
As a novel de-ubiquitinating enzyme with ubiquitin hydrolase activity, USP22 might inhibit apoptosis in HCC by activating the BMI-1-mediated PcG stem cell pathway [ xref ].
As a consequence, gain of USP22 functions promotes cell cycle progression and inhibits cell apoptosis, leading to cancer cell hyper-proliferation and tumorigenesis.
In the attempt to explore the mechanisms by which USP22 silencing leads to ATC cell apoptosis, we found that the proapoptotic members of Bcl-2 family proteins, Bid and Bax, were upregulated in response to USP22 knockdown, consistent with increased activation of caspase-3.
Our previous studies have confirmed that defects in USP22 can not only cause cell cycle arrest in G0/G1 phase, but also inhibit apoptosis and promote tumor cell proliferation [XREF_BIBR].
Further experimental verification showed that overexpression of USP22 reversed the suppression effects of miR-132-3p overexpression on the proliferation, migration and invasion of SW480 and SW620 cells, as well as the acceleration effect of apoptosis (XREF_FIG -G), further confirming that miR-132-3p regulated the progression of CRC by targeting USP22.
Silencing of USP22 in podocytes attenuated high d-glucose-induced apoptosis and inflammatory responses, evidenced by increases in proliferation and MMP levels and decreases in the apoptotic rate, ROS production, the Bax and Bcl -2 ratio, caspase-3 expression and secretion of TNF-alpha, IL-1beta, IL-6 and TGF-beta1.
Further analysis demonstrated that USP22 downregulation activated mitochondrial apoptosis by regulating several apoptosis related proteins, including Bcl-2, Bax, cytochrome c and caspase-3.
To investigate the molecular mechanism underlying USP22 silencing induced ATC cell apoptosis, we examined the cellular levels of documented apoptosis regulators or executioners.
Additionally, investigation into the underlying mechanism, using small interfering RNA, revealed that the downregulation of USP22 inhibited proliferation and promoted apoptosis though the phosphoinositide 3-kinase/protein kinase B signaling pathway.
The overexpression of USP22 significantly enhanced cell proliferation potency and telomerase activity, elevated TERT expression level, inhibited p53 expression and cell aging, as well as decreased cell apoptosis or DNA damage.
USP22 can promote lung cancer cell invasion via epithelial-mesenchymal transition (EMT), which participates in the metastasis of primary tumors by activating TGF-beta1 [XREF_BIBR].
We drew a conclusion that USP22 could increase the abilities of proliferation, migration and invasion of glioma cells, and promote the growth and development of glioma.
Through measuring the proliferation, migration and invasion of SW480 and SW620 cells, we uncovered that USP22 knockdown could suppress the proliferation, migration and invasion of CRC cells, while USP22 overexpression had an opposite effect (Supplement Figure 2).
Silencing USP22 decreased the migration and invasion of Bel/Fu cells, and combination with 5-Fu further decreased the rate of migration and number of invading cells.
Silencing USP22 in chemoresistant HCC Bel/Fu cells dramatically inhibited proliferation, migration, invasion and epithelial-mesenchymal transition in vitro; suppressed tumorigenic and metastatic capacities in vivo; and inhibited drug resistance related proteins (MDR1, LRP, MRP1).
Taken together, these results indicate that USP22 increases cell migration and invasion by inducing EMT by binding to the promoter of AP4 to activate its transcription.
We report that deletion of USP22 in pancreatic tumor cells reduced the infiltration of myeloid cells and promoted the infiltration of T cells and NK cells, leading to an improved response to combination immunotherapy.
Ablation of USP22 in liver tumor cells has been shown to increase tumor immunogenicity and promote infiltration of T cells into the resulting liver tumors.
The results of in vitro and in vivo studies confirmed USP22 depletion reduced the growth and invasion of ATC cells by regulating the expression and activation of a series of pro tumorigenesis molecules.
The assay results showed that USP22 downregulation significantly inhibited the proliferation (XREF_FIG) and invasion (XREF_FIG) of U2OS and MG-63 cells.
We report that deletion of USP22 in pancreatic tumor cells reduced the infiltration of myeloid cells and promoted the infiltration of T cells and NK cells, leading to an improved response to combination immunotherapy.
Further experimental verification showed that overexpression of USP22 reversed the suppression effects of miR-132-3p overexpression on the proliferation, migration and invasion of SW480 and SW620 cells, as well as the acceleration effect of apoptosis (XREF_FIG -G), further confirming that miR-132-3p regulated the progression of CRC by targeting USP22.
Given the ability of USP22 to enhance AR accumulation, AR activity, and CRPC, the biological impact of a model of tetracycline inducible shUSP22 was developed in therapy sensitive PCa cells.
Together, these data demonstrate that USP22 regulates ligand independent AR residence at target gene loci and promotes AR driven CRPC gene profiles, which may have specificity for USP22 perturbation, further implicating USP22 as an independent effector of aggressive tumor phenotypes.
These data suggest that USP22 functions to enhance AR stability and promote inappropriate castration resistant AR signaling through proteasome dependent regulation of AR levels.
Strikingly, USP22 deregulation significantly increased AR occupancy at known ARORs (1.15-2.6% input; XREF_FIG), but not in control regions of the KLK3 and PSA (' EF ' region, XREF_SUPPLEMENTARY).
Based on the data above, suppression of USP22 in models of ADT sensitive PCa and aggressive CRPC decreased the AR signaling axis (XREF_FIG and XREF_FIG).
By contrast, USP22 deregulation induced marked enhancement of ligand independent AR activity, determined by analyses of multiple, clinically relevant AR target genes (XREF_FIG).
As shown, in CRPC cells, USP22 depletion suppressed AR protein accumulation in both the androgen stimulated and deprived conditions (XREF_FIG, compare lanes 1,2 and 3,4).
Conversely, depletion of USP22 dramatically down-regulates AR protein levels and abrogates basal and DHT stimulated AR activity in both ADT sensitive and CRPC cells.
In contrast, USP22 depletion reduced AR protein levels in the absence of androgen, and inhibited DHT induced AR expression, within in models of therapy sensitive disease (XREF_FIG, compare lanes 2, 4).
Schrecengost et al XREF_BIBR previously showed that USP22 depletion dramatically downregulated androgen receptor protein levels and abolished androgen receptor activity in both androgen deprivation therapy and castration resistant prostate adenocarcinoma cells.
The observations that USP22 upregulation is sufficient to promote ligand independent AR expression and activity, and induce Casodex resistance are clinically relevant, as these attributes reflect key biochemical characteristics of CRPC.
Further, USP22 mediates AR expression through a proteasome dependent mechanism, since modeling clinically-relevant USP22 upregulation results in an increased AR protein half-life and proteasome inhibition rescued the decreased AR expression following USP22 depletion.
Studies have shown that overexpression of USP22 can enhance the inhibitory effect of cell cycle inhibitors such as p21 and enhance the proliferation of tumor cells, thus promoting the occurrence and development of tumors [XREF_BIBR].
We found that inhibition of USP22 suppressed cell proliferation by inducing G1 phase cell cycle arrest through synergy with oncogenic transforming growth factor-beta1 (TGFB1).
Gain- and loss-of-function assays showed that USP22 promoted gastric cancer cell growth and cell cycle transition while suppressing apoptosis in vitro.
XREF_BIBR In addition, another study demonstrated that silencing USP22 could inhibit proliferation and induce cell cycle arrest in bladder cancer cells.
First, using hormone-proficient conditions, USP22 enhanced the rate of cell cycle progression, evidenced through increased BrdU incorporation (XREF_FIG, left).
While it is clear that USP22 is overexpressed in various cancer types and may promote oncogenesis by altering gene expression, cell death and cell cycle progression, emerging evidence suggests that USP22 also harbors tumor suppressor like properties.
As shown, doxycycline (Dox) decreased USP22, resulting in marked loss of AR (XREF_FIG, top), attenuated cell cycle progression (determined by BrdU incorporation XREF_FIG, top right, middle), and significantly suppressed of cell doubling (XREF_FIG, bottom).
Knockdown of USP22 was found to suppress cell proliferation in vitro and tumour growth in vivo by inducing G1 phase cell cycle arrest through synergy with TGF-beta1 (Ji et al., 2015).
However, Ling et al reported that knockdown of USP22 by siRNA induced cells G0/G1 cell cycle arrest via the c-Myc and cyclin D2 pathway in HepG2 cells.
XREF_BIBR - XREF_BIBR Zhang and colleagues have demonstrated that ectopic overexpression of USP22 promotes cell proliferation and that suppression of USP22 expression by small hairpin RNA induces cell cycle arrest in human lung cancer cells.
Functionally, this overexpression of USP22 actively contributes to tumorigenesis, as USP22 depletion blocks cancer cell cycle progression in vitro, and inhibits tumor progression in animal models of lung, breast, bladder, ovarian, and liver cancer, among others.
In stomach cancer, USP22 abundance increased from normal tissue to primary carcinoma to lymph node metastasis and was also associated with shorter patient survival (26 vs. 59 months disease specific survival for USP22 positive vs. -negative primary carcinoma) [XREF_BIBR].
These results suggest that USP22 promotes tumor development and metastasis, and highlight USP22 as a novel prognostic marker and potential therapeutic target in ATC.
These results lend further support to the results of our vitro experimental studies, as they showed that USP22 promotes CRC cell metastasis by activating AP4 to induce EMT.
Moreover, USP22 overexpression can promote EMT and TGF-beta expression, whereas depletion of USP22 can reverse EMT and reduce metastasis of lung adenocarcinomas.
Similarly, Zhao et al. reported that USP22 depletion suppressed cell survival and proliferation as well as tumor growth and lung metastasis of anaplastic thyroid carcinoma cells XREF_BIBR.
Moreover, it has been shown that USP22 promotes cell growth by regulating the far upstream element (FUSE)-binding protein 1 (FBP1), a transcriptional regulator of p21 [XREF_BIBR].
Collectively, our results show that USP22 silencing in HepG2 cells suppressed cell growth through mitochondrial apoptosis and that this suppression was dependent on caspase activation.
Taken together, our results suggest that USP22 promotes NSCLC cell growth in vitro and NSCLC tumorigenesis in vivo, and these effects are through MDMX up-regulation and subsequent p53 inhibition.
Furthermore, USP22 small interfering RNA inhibited cell growth and reduced the expression levels of Aurora-B, Survivin and Cyclin B, together with the upregulation of cyclin dependent kinase inhibitor 1A (p21).
These results suggested that overexpression USP22 induced EMT in lung adenocarcinoma cells.Moreover it is well known that TGF-beta1 stimulates the EMT in lung cancer cells [43].
Subsequent experiments showed that USP22 knockdown resulting from up-regulation of miR-30e-5p could inhibit proliferation, invasion, migration, and EMT in 5-8F cells.
According to the results of previous studies, USP22 can induce EMT by regulating TGF-beta1 in lung cancer cells [XREF_BIBR], and elevated USP22 expression can promote EMT by up-regulating ZEB1 and Snail in pancreatic cancer cells though a process that involves focal adhesion kinase (FAK) signaling [XREF_BIBR].
Moreover, USP22 overexpression can promote EMT and TGF-beta expression, whereas depletion of USP22 can reverse EMT and reduce metastasis of lung adenocarcinomas.
USP22 Inhibits SIRT1 to Regulate Ferroptosis-Induced Cardiomyocyte Death A previous study has shown that USP22 possesses the ability to stabilize SIRT1 by the process of deubiquitination ( Lin et al ., 2012 ) .
Additionally, SIRT1 instability caused by loss of USP22, a deubiquitinating enzyme that stabilizes SIRT1, is associated with the defective embryogenesis in USP22 null mice [XREF_BIBR].
Given that SIRT1 is deubiquitinated by USP22 and stabilized at the protein level (Armour etal., 2013; Lin etal., 2012) and previous studies have reported that SIRT1 could negatively influence the chemosensitivity of HCC cells (Chen etal., 2012), our results supported the notion that USP22 increases SIRT1 protein levels in HCC cells.
USP22 knockdown reduced SIRT1 expression in FLT3-ITD AML cells, whereas USP22 overexpression increased SIRT1 levels by increasing protein stability, indicating that USP22 is an important positive regulator of SIRT1 in FLT3-ITD cells.
In addition, USP22 knockdown prevented c-MYC-mediated reduction of SIRT1 ubiquitination (XREF_FIG) and increase in SIRT1 expression (XREF_SUPPLEMENTARY).
In addition, USP22 could also decrease the acetylation of Ku70 by stabilizing the expression of Sirt1, thus inhibiting Bax mediated apoptosis and contributing to cisplatin resistance.
USP22 knockdown reduced SIRT1 expression in FLT3-ITD AML cells, whereas USP22 overexpression increased SIRT1 levels by increasing protein stability, indicating that USP22 is an important positive regulator of SIRT1 in FLT3-ITD cells.
USP22 also increases SIRT1 protein stability, which leads to the suppression of p53 transcriptional activity and inhibition of cell death [XREF_BIBR, XREF_BIBR].
Intriguingly, we found that USP22 silencing activates the p53 pathway in human NSCLC cells and tumor tissues along with downregulation of MDMX protein, a major negative regulator of p53.
On the bases of these results, we speculated that USP22 silencing activates the p53 pathway in NSCLC cells by post-transcriptional down-regulation of MDMX.
Hence USP22 silencing reduces the capacity of FBP1 to repress p21 (independently of TP53 status), which in turn inhibits CDKs to prevent the G1/S transition and resulting in G1 accumulation [XREF_BIBR].
Several studies demonstrated that USP22 silencing could activate p53. xref , xref Because USP22, Ube2d4, and Ube3b were important downstream genes of YWHAZ, we infer that YWHAZ downregulates p53 by activating USP22, Ube2d4, and Ube3b, which make P53 ubiquitination and lead to its proteasomal degradation.
USP22 also play roles in cell cycle regulation, where depletion of USP22 increases the expression of p53 and p21, inhibits proliferation, and induces cell cycle arrest at G1 phase.
We found that USP22 silencing in A549 and NCI-H460 cells increased the protein expression of p53, p21 and Bax, the key p53 signal molecules (XREF_FIG A), suggesting that p53 activation plays a role in USP22 silencing induced growth inhibition.
According to that model, USP22 enhances DNA damage repair and cisplatin resistance by deubiquitinating histone H2A, which in turn facilitates the phosphorylation of histone H2AX.
Combining with our previous results, we concluded that both USP22 and Sirt1 can induce cisplatin resistance, but Sirt1 overexpression ca n't phenocopy USP22 mediated cisplatin resistance.
The study reveal the dual mechanism of USP22 involvement in cisplatin resistance : (1) USP22 enhances DNA damage repair and induce cisplatin resistance by promoting the phosphorylation of histone H2AX via deubiquitinating histone H2A.
These results confirm that USP22 is involved in the cisplatin resistance of A549 and CDDP cells and H2AX, gammaH2AX, and Sirt1 may be responsible for USP22 mediated cisplatin resistance.
To verify whether inhibition of USP22 expression could reverse the cisplatin resistance of A549 and CDDP cells, CCK8 assays showed that, after the inhibition of USP22 expression, the 48h IC50 of A549 and CDDP decreased from 0.925 +/- 0.04 muM to 0.337 +/- 0.03 muM.
To verify whether inhibition of USP22 expression could reverse the cisplatin resistance of A549 and CDDP cells, CCK8 assays showed that, after the inhibition of USP22 expression, the 48h IC50 of A549 and CDDP decreased from 0.925 +/- 0.04 muM to 0.337 +/- 0.03 muM.
These results suggest that inhibiting USP22 expression enhanced cisplatin sensitivity in lung adenocarcinoma by downregulation of Sirt1 and gammaH2AX in vivo.
Furthermore, USP22 knockout significantly impaired non homologous DNA damage repair capacity, enhanced cisplatin and irradiation induced apoptosis in these cells.
Western blot analysis showed that USP22 overexpression also induced activation of the RAS and ERK and PI3K and AKT pathways in SGC7901 cells and xenograft tumor tissues.
Previous studies have confirmed that USP22 can promote the biological process of NSCLC cells by regulating BMI-1 and AKT signaling pathway [XREF_BIBR].
Moreover, SIRT1 deficiency attenuated USP22 induced activation of the AKT and MRP1 pathway in BEL7402 cells, suggesting that USP22 regulated the AKT and MRP1 pathway in a SIRT1 dependent manner.
Furthermore, our results showed that USP22 deletion also caused down-regulation of Akt and GSK3beta activity, which can also contribute to the reduction of cyclin D2.
We have reported that USP22 mediates cell survival and proliferation by promoting the expression of BMI-1 and upregulation of activated AKT pathway in colon cancer cells.
The results showed that USP22 downregulation remarkably decreased the protein expression of p-PI3K and p-Akt without change in the total protein levels of PI3K and Akt.
The downregulation of USP22 suppresses multidrug resistance-associated protein 1 ( MRP1 ) to induce intracellular sorafenib accumulation and hampers glycolysis of HCC cells .
] Our study provided strong evidence that the downregulation of USP22 by Gal-SLPs suppressed the expression of MRP1 and caused high intracellular sorafenib accumulation .
In our previous study , USP22 bound to SIRT1 and subsequently activated the AKT pathway , increasing the expression of MRP1 to induce 5-FU resistance in HCC cells [ 15 ] .
iv ) The downregulation of USP22 not only suppresses multidrug resistance-associated protein 1 ( MRP1 ) , enhances the intracellular accumulation of sorafenib , and finally inhibits cell proliferation and cancer angiogenesis , but also inhibits glycolysis in hepatocellular carcinoma ( HCC ) cells , enhancing sorafenib chemosensitivity and impairing cell metabolism .
Subsequently, using qPCR array analysis, we found that knockdown of USP22 could drastically inhibit the expression of MRP1, but not P-gp, in BEL/FU cells.
Collectively, USP22 might deubiquitinate SIRT1 and subsequently activate the AKT pathway, increasing the expression of MRP1 to induce MDR in HCC cells.
The downregulation of USP22 suppresses multidrug resistance associated protein 1 (MRP1) to induce intracellular sorafenib accumulation and hampers glycolysis of HCC cells.
Studies herein suggest that USP22 mediated MYC regulation in PCa is likely gene selective, based on USP22 increasing gene expression of AR/MYC co-regulated target (ODC) but not of multiple MYC targets.
While these outcomes could occur via USP22 mediated MYC and CyclinD2 and/or BMI-1-mediated modulation, AR activity promotes tumorigenesis in several of these tumor types, including bladder, HCC and breast carcinoma.
C-Myc abundance is also indirectly increased by the activity of USP22 on its substrate SIRT1, a NAD dependent protein deacetylase [XREF_BIBR] (see Section 7), but this is not observed in all cell types [XREF_BIBR].
Decreased levels of USP22 reduce the ability of c-MYC, which can stimulate activation of AP4 to directly or indirectly induce EMT [XREF_BIBR], activating the transcription of its targets [XREF_BIBR, XREF_BIBR].
In conclusion, the present study reveals that USP22 in breast cancer cell lines increases c-Myc stability through c-Myc deubiquitination, which is closely correlated with breast cancer progression.
Because the Myc protein controls the expression of thousands of other genes, the depletion of cellular USP22 can inhibit Myc function, inhibiting the invasive growth of cancer cells.
Above, we demonstrated that USP22 silencing predominantly decreases the BMI1 protein levels rather than mRNA expression, and further alters GC cell proliferation, gastric CSC formation and maintenance of stem cell stemness, indicating post-transcriptional regulation of BMI1.
We have reported that USP22 mediates cell survival and proliferation by promoting the expression of BMI-1 and upregulation of activated AKT pathway in colon cancer cells.
An in vitro study showed that the upregulation of USP22 mediated the enhanced expression of BMI1 and Cyclin D2, and was responsible for increased cell proliferation and the metastatic behavior of colon cancer cells .
In this study, we demonstrated that USP22 mediated protein stabilization of BMI1 promotes gastric CSC stemness maintenance and GC progression, thereby providing a rationale for USP22 targeting as a potential therapeutic approach against GC.
Previous studies have confirmed that USP22 can promote the biological process of NSCLC cells by regulating BMI-1 and AKT signaling pathway [XREF_BIBR].
In this study, we showed that USP22 depletion significantly decreased the expressions of BMI-1, vimentin, and snail and increased E-cadherin expression in ATC cells.
GSK3beta- and USP22 dependent KDM1A stabilization is required for the demethylation of histone H3K4, thereby repressing BMP2, CDKN1A and GATA6 transcription, which results in cancer stem cell self-renewal and glioblastoma tumorigenesis.
In MGC-803 cells and SGC-7901 cells, knockdown of USP22 and BMI1 both increased P21 expression and reduced the expression of CSC stemness genes of CD133 and SOX2.
USP22 also suppresses CDKN1A and p21 -locus expression by deubiquitinating its transcriptional repressor FBP1 (Atanassov and Dent, 2011), and acts as a negative regulator of the tumor suppressor p53 b[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]
USP22 has also been demonstrated to inhibit transcription of the p21 gene by deubiquitinating the transcriptional regulator, FBP1, leading to cell proliferation and tumorigenesis.
We found that USP22 silencing in A549 and NCI-H460 cells increased the protein expression of p53, p21 and Bax, the key p53 signal molecules (XREF_FIG A), suggesting that p53 activation plays a role in USP22 silencing induced growth inhibition.
We found that USP22 depletion inhibited the proliferation of the human GC cell lines MGC-803 and SGC-7901 cells and increased expression of P21, indicating cell cycle arrest [XREF_BIBR].
Recent studies have demonstrated that USP22 can inhibit the transcription of the p21 gene by de-ubiquitinating the transcriptional regulator FBP1, leading to cell proliferation and tumorigenesis [XREF_BIBR].
Hence USP22 silencing reduces the capacity of FBP1 to repress p21 (independently of TP53 status), which in turn inhibits CDKs to prevent the G1/S transition and resulting in G1 accumulation [XREF_BIBR].
USP22 knockdown, using lentivirus delivered siRNA, increased the expression levels of cell cycle proteins P21 and P27, but reduced the levels of phosphorylated retinoblastoma protein, resulting in the inhibition of FaDu cell growth and proliferation.
USP22 also play roles in cell cycle regulation, where depletion of USP22 increases the expression of p53 and p21, inhibits proliferation, and induces cell cycle arrest at G1 phase.
Ubiquitin specific protease 22 (USP22) reduces the degradation of sirtuin-1 and the expression of FN and TGF-beta1 in AGE treated GMCs, whereas depletion of USP22 promotes sirtuin-1 degradation and the expression of FN and TGF-beta1 in this cell model.
Ubiquitin specific protease 22 (USP22) reduced Sirt1 ubiquitination and degradation and decreased FN and TGF-beta1 expression in GMCs under both basal and AGEs treated conditions.
Furthermore, we have found overexpression of USP22 correlated with high TGF-beta1 expression in the lung ADC tissues.To determine whether USP22 expression in lung ADC cells induced EMT by changing the[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]
In diabetic nephropathy, the increased expression of USP22 reduced the Sirt1 ubiquitination and degradation, and decreased fibronectin and TGF-beta1 expression in glomerular mesangial cells under both[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]
Ubiquitin specific protease 22 (USP22) reduced Sirt1 ubiquitination and degradation and decreased FN and TGF-beta1 expression in GMCs under both basal and AGEs treated conditions.
Ubiquitin specific protease 22 (USP22) reduces the degradation of sirtuin-1 and the expression of FN and TGF-beta1 in AGE treated GMCs, whereas depletion of USP22 promotes sirtuin-1 degradation and the expression of FN and TGF-beta1 in this cell model.
In diabetic nephropathy, the increased expression of USP22 reduced the Sirt1 ubiquitination and degradation, and decreased fibronectin and TGF-beta1 expression in glomerular mesangial cells under both[MISSING/INVALID CREDENTIALS: limited to 200 char for Elsevier]