A New Potential Therapeutic Target for Cancer in Ubiquitin-Like Proteins—UBL3

Ubiquitin-like proteins (Ubls) are involved in a variety of biological processes through the modification of proteins. Dysregulation of Ubl modifications is associated with various diseases, especially cancer. Ubiquitin-like protein 3 (UBL3), a type of Ubl, was revealed to be a key factor in the process of small extracellular vesicle (sEV) protein sorting and major histocompatibility complex class II ubiquitination. A variety of sEV proteins that affects cancer properties has been found to interact with UBL3. An increasing number of studies has implied that UBL3 expression affects cancer cell growth and cancer prognosis. In this review, we provide an overview of the relationship between various Ubls and cancers. We mainly introduce UBL3 and its functions and summarize the current findings of UBL3 and examine its potential as a therapeutic target in cancers

High Expression of Derlin-1 Is Associated with the Malignancy of Bladder Cancer in a Chinese Han Population

<div><p>Derlin-1 is overexpressed in various types of solid tumors and has an important function in cancer progression. However, its expression pattern in and association with the clinicopathological characteristics of human bladder cancer remain unclear. In the present study, 3 pairs of fresh samples of bladder cancer tissue and paracancerous tissue were first detected by liquid chromatography tandem mass spectrometry (UPLC-MS/MS) to screen for differentially expressed proteins. Following bioinformatics analysis and assessments by qRT-PCR and western blotting, Derlin-1 was selected as a candidate protein and was then validated in samples from patients with bladder cancer by immunohistochemistry and western blotting. The results showed that the bladder cancer tissues exhibited higher levels of Derlin-1 expression than the paracancerous tissues (P < 0.05). Positive expression of Derlin-1 was significantly correlated with tumor stage, histological grade, and lymph node metastasis (P < 0.001) but was not correlated with other clinicopathological parameters including patient age (P = 0.758) and gender (P = 0.831). Besides, Derlin-1 was highly expressed in BC cell lines (um-uc-3 and T24), and the interference of Derlin-1 could reverse EMT progression, inhibit the tumor migration and invasion in T24 cells. Further, patients with positive Derlin-1 expression had shorter overall survival than those with negative expression (P < 0.001). Taken together, our results demonstrated that Derlin-1 was overexpressed in bladder cancer and was associated with the malignancy of bladder cancer.</p></div

Expression of Derlin-1 protein in human bladder cell lines.

<p>(A) Expression of Derlin- protein using western blotting in SV-HUC-1 cells, um-uc-3 cells and T24 cells. (B) Interfere of Derlin-1 in T24 cells. Cells were transfected with nontargeting siRNA or Derlin-1 siRNA for 48h, and the protein expression was detected by western blotting. NC, siRNA negative control; siRNA, Derlin-1 siRNA. Data are expressed as the mean ± S.D., **P < 0.01, ***P < 0.001.</p

Effects of Derlin-1 on the migration and invasion in T24 cells.

<p>(A) Effects of Derlin-1 on cellular migration ability. (B) Effects of Derlin-1 on cellular invasion ability. Representative microscopy images of the migration and invasion assay are shown as ×100. NC, siRNA negative control; siRNA, Derlin-1 siRNA. *P < 0.05, **P < 0.01.</p

Effects of Derlin-1 on epithelial–mesenchymal transition (EMT) in T24 cells.

<p>(A-B) Expressions of E-Cadherin and Vimentin by Western blot assay in T24 cells transfected with Derlin-1 siRNA. β-actin was probed as the loading control. ***P<0.001, compared with negative control. (C-D) Expression of Derlin-1, E-Cadherin and Vimentin (<i>red</i>) by immunofluorescence staining analysis using confocal in T24 cells transfected with Derlin-1 siRNA. The nuclei were stained with DAPI (<i>blue</i>). scale bar: 5 μm.</p

Representative images of Derlin-1 protein expression in IHC microarrays.

<p>(A and a), normal mucosa; (B and b), noninvasive cancer; (C and c), invasive cancer; (D and d), negative control; IHC = immunohistochemistry (A, B, C and D, magnification ×40, scale bars: 200 μm; a, b, c and d, magnification ×200, scale bars: 50 μm).</p

Alpha-Synuclein Interaction with UBL3 Is Upregulated by Microsomal Glutathione S-Transferase 3, Leading to Increased Extracellular Transport of the Alpha-Synuclein under Oxidative Stress

Aberrant aggregation of misfolded alpha-synuclein (α-syn), a major pathological hallmark of related neurodegenerative diseases such as Parkinson’s disease (PD), can translocate between cells. Ubiquitin-like 3 (UBL3) is a membrane-anchored ubiquitin-fold protein and post-translational modifier. UBL3 promotes protein sorting into small extracellular vesicles (sEVs) and thereby mediates intercellular communication. Our recent studies have shown that α-syn interacts with UBL3 and that this interaction is downregulated after silencing microsomal glutathione S-transferase 3 (MGST3). However, how MGST3 regulates the interaction of α-syn and UBL3 remains unclear. In the present study, we further explored this by overexpressing MGST3. In the split Gaussia luciferase complementation assay, we found that the interaction between α-syn and UBL3 was upregulated by MGST3. While Western blot and RT-qPCR analyses showed that silencing or overexpression of MGST3 did not significantly alter the expression of α-syn and UBL3, the immunocytochemical staining analysis indicated that MGST3 increased the co-localization of α-syn and UBL3. We suggested roles for the anti-oxidative stress function of MGST3 and found that the effect of MGST3 overexpression on the interaction between α-syn with UBL3 was significantly rescued under excess oxidative stress and promoted intracellular α-syn to extracellular transport. In conclusion, our results demonstrate that MGST3 upregulates the interaction between α-syn with UBL3 and promotes the interaction to translocate intracellular α-syn to the extracellular. Overall, our findings provide new insights and ideas for promoting the modulation of UBL3 as a therapeutic agent for the treatment of synucleinopathy-associated neurodegenerative diseases

Analysis of 5-year overall survival in patients with positive or negative Derlin-1 expression.

<p>Kaplan–Meier survival analysis was used to compare 5-year overall survival between patients with positive or negative Derlin-1 expression. P < 0.001, compared with patients with negative Derlin-1 expression.</p

Derlin-1 was highly expressed in bladder cancer tissue microarrays.

<p>(A) Scatter plot of Derlin-1 protein staining score in the sections which was semiquantitatively scored according to staining intensity and extent. Staining intensity time received a final score of 0–12. (B) The percentage rate of positive Derlin-1 expression in bladder cancer tissues. Data are expressed as the mean±S.D., ***P < 0.001.</p

Derlin-1 protein expression in normal mucosa and bladder cancer.

<p>(A) Representative graphic of western blot analysis of Derlin-1 protein expression in bladder cancer tissues (T) and paracancerous tissues (N) from four patients. (B) Statistical result. *P < 0.05, **P < 0.01.</p