NEDD8 knockdown via transfection with si‐NEDD8 had similar effects to si‐UCHL3, as well as si‐UCHL3+ si‐NEDD8. Taken together, the results of the present study suggest that UCHL3 knockdown decreases melanoma cell proliferation by increasing cell autophagy through regulating NEDD8 expres‐

It has been reported that ubiquitin C‐terminal hydrolase‐L3 (UCHL3) plays an important role in cancer development; however, the role of UCHL3 in melanoma remains unclear. The present study aimed to investigate the role of ubiquitin C‐terminal hydrolase‐L3 (UCHL3) and determine its underlying molecular mechanisms in melanoma occurrence and development using in vitro studies. Reverse transcription‐quantitative PCR analysis was performed to detect UCHL3 mRNA expression. The MTT assay was performed to assess cell proliferation. Cell apoptosis was analyzed via flow cytometry and the TUNEL assay. Cell ultrastructure was observed via transmission electron micros‐ copy. LC3B protein expression was detected via cellular immunofluorescence, while neural precursor cell‐expressed developmentally downregulated protein 8 (NEDD8) and LC3 protein expression levels, and NEDD8 ubiquitination were detected via western blot analysis. The results demonstrated that transfection with small interfering (si)RNA‐UCHL3 signif‐ icantly suppressed cell proliferation, whereas apoptosis was significantly enhanced, as well as autophagy, autophagosome formation and LC3B protein expression. In addition, NEDD8 protein expression and autophagosome numbers significantly decreased, while the LC3II/LC3I ratio significantly increased. NEDD8 knockdown via transfection with si‐NEDD8 had similar effects to si‐UCHL3, as well as si‐UCHL3+ si‐NEDD8. Taken together, the results of the present study suggest that UCHL3 knockdown decreases melanoma cell proliferation by increasing cell autophagy through regulating NEDD8 expres‐ sion and autophagosome numbers. Introduction Melanoma is the most common malignancy of the skin, with increasing incidence rate (1). Melanoma has a high risk of metastasis. Current treatment options for melanoma primarily consist of surgical resection, radiotherapy and chemotherapy; however, these methods are only effective for tumors at early stages (2). Once metastasis occurs, the median survival time is only 6‐9 months (3). Ubiquitin C‐terminal hydrolase‐L3 (UCHL3) is a member of the ubiquitin carboxyl terminal hydrolase (UCH) family (4). Our previous studies have demonstrated that neural precursor cell‐expressed developmentally downregulated protein (NEDD) is upregulated in melanoma tissues and cells, with associated changes in several regulatory enzymes, among which UCHL3 exhibits the most significant alterations (5,6). Recently, studies have been focusing on the role of the UCH family in cancer (7,8). Unlike other members of this family, UCHL3 not only func‐ tions as a ubiquitin hydrolase, but also regulates NEDD8 hydrolase (9). Furthermore, it acts as a cleavage enzyme of the NEDD8 precursor to promote the maturation of NEDD8 (10). NEDD8 is a ubiquitin‐like protein, and neddylation plays a key role in regulating ubiquitin ligase E3 and promoting the function of the ubiquitin pathway (11). The process of neddylation is similar to ubiquitination, which is maintained by a series of regulatory enzymes. Following transcription, NEDD8 is present in the form of its precursor, which is subsequently converted to mature NEDD8 following hydro‐ lysis by UCHL3 hydrolase (12). Mature NEDD8 is activated by NEDD8 activase and subsequently transferred to NEDD8 ligase (13). NEDD8 carried by E2 is transferred to its substrate and associated with the cullin protein (14). However, NEDD8 hydrolase can cause an isomeric change of NEDD8, dissociating NEDD8 from cullin, which plays a negative regulatory role, a process referred to as de‐neddylation (8). Thus, neddylation and de‐neddylation maintain the balance by simultaneously acting on the NEDD8 protein (15). UCHL3 not only acts on the NEDD8 precursor to convert it to its mature form, but also hydrolyzes the bound NEDD8 (16). Thus, the present study aimed to investigate the effect of UCHL3 knockdown on the biological activities of melanoma cells, and determine its role in downstream NEDD8 signaling via in vitro cell experiments. UCHL3 plays an important role in the occurrence and development of melanoma RUNZHI HE1, YAJING ZHOU2, JIANMIN LIU3, XIAOCHONG ZHANG4, XIAOLING ZHAO5, LIHUI AN6, ZIHAN LI6 and FANG CHENG6 1The Third Department of Neurosurgery, 2Department of Anesthesiology, 3Department of Orthopedics, 4Science and Education Section, 5Tumor Laboratory, 6Department of Dermatology, Xingtai People's Hospital, Xingtai, Hebei 054001, P.R. China Received April 5, 2021; Accepted June 22, 2021 DOI: 10.3892/ol.2021.13017 Correspondence to: Dr Fang Cheng, Department of Dermatology, Xingtai People's Hospital, 16 Hongxing Street, Xingtai, Hebei 054001, P.R. China E‐mail: chengfang0127@163.com