The NEDD4 subfamily, also referred to as C2-WW-HECT-type E3s, typically contains an N-terminal C2 domain (mediating Ca²⁺-dependent membrane targeting), two to four WW domains (recognizing PPxY motifs or phosphorylated sequences), and a C-terminal HECT domain (4,26,27). The C2 domain is a Ca²⁺-dependent phospholipid-binding motif that mediates membrane association and recognition of membrane-associated substrates (4,26). WW domains mediate protein-protein interactions by recognizing PPxY, LPxY, or related sequences, as well as phosphorylated Ser/Thr-Pro motifs (4,27). Essentially, the C2 and WW domains determine the specificity of target protein binding for NEDD4 family members. Known NEDD4 subfamily E3s include NEDD4, NEDD4L, ITCH, SMURF1, SMURF2, WWP1, WWP2, HECW1 and HECW2 (28). The NEDD4 family represents the most extensively studied group of HECT-type E3s, which regulate processes such as plasma membrane protein stability and misfolded protein degradation, and are widely implicated in the mechanisms of liver diseases (29).

HERC subfamily members contain an N-terminal RCC1-like domain (involved in regulating GTPases) and a C-terminal HECT domain. They are subdivided into members such as HERC1-6 based on variations in their intermediate domains. The conserved RCC1 domain mediates GTPase regulation, contributing to pleiotropic roles in cellular homeostasis (30). Their diverse intermediate domains (for example, zinc fingers, ISG15-binding domains) confer distinct functions to individual members, including roles in DNA repair, antiviral immunity and metabolic regulation, potentially exerting protective or repair functions in pathological processes such as liver injury and hepatitis (30,31).

The other group of HECT members includes 13 HECT-type E3s (for example, HECTD1-4, E6AP/UBE3A, HUWE1, TRIP12, UBR5, UBE3B, UBE3C, HACE1, G2E3 and AREL1), all of which possess a C-terminal HECT domain but often contain N-terminal or intermediate domains that are unstructured or lack defining features, precluding their classification into the aforementioned subfamilies (32,33). These E3s have been reported to participate in various disease processes by regulating energy metabolism, ERS, oxidative stress and cell death (34,35).

Dysregulation of lipid homeostasis is a central pathological event in patients with MASLD. Several HECT-type E3 ligases directly modulate key transcriptional regulators and enzymes within lipid metabolic pathways.

The roles of proteins from the NEDD4 subfamily in the pathogenesis of MASLD/MASH have been extensively reported. Research on NEDD4 within this subfamily in MASLD remains relatively limited; current evidence indicates that it is upregulated under conditions of elevated hepatic free fatty acids, where it regulates high-density lipoprotein metabolism by ubiquitinating the scavenger receptor SR-BI (40). Conversely, a study in geese reported opposing findings, showing increased NEDD4 expression in fatty liver that appeared to suppress hepatic steatosis-related injury via ubiquitination of PTEN and IGF-1R (41), suggesting potential species-specific functional differences. NEDD4L has been more extensively studied, revealing a dual role: Wherein reduced AT-rich interactive domain 2 expression in MASLD livers leads to NEDD4L transcriptional downregulation, impairing JAK2 ubiquitination and subsequent degradation, which results in hyperactivation of the JAK-STAT pathway and ultimately exacerbates hepatic steatosis (42), indicating that the normal function of NEDD4L suppresses lipid accumulation. However, the markedly elevated NEDD4L levels observed in MASH serve to promote the K48-linked ubiquitination and degradation of lysosomal-associated protein transmembrane 5 (LAPTM5), activating the cell division cycle 42 (CDC42)-MAPK axis and paradoxically worsening lipid accumulation (43), while NEDD4L-mediated degradation of TMBIM1 has also been confirmed to aggravate steatosis (44). The role of ITCH in lipid metabolism is similarly complex. ITCH knockout mice fed a high-fat diet exhibit reduced hepatic lipid accumulation and inhibited atherosclerosis formation, suggesting that ITCH promotes lipid accumulation, Previous studies indicated that ITCH knockout mice fed a high-fat diet exhibit reduced hepatic lipid accumulation and suppressed atherosclerosis formation. This is associated with ITCH enhancing LDL uptake by mediating the ubiquitination of proteins such as sirtuin 6 (SIRT 6), Sterol regulatory element binding protein 2 (SREBP 2) and liver kinase B1, thereby influencing lipid accumulation. Clinical samples similarly reveal a negative correlation between human ITCH protein abundance and hepatic steatosis severity (45-48). Additionally, ITCH binds to the PPAY motif in Spartin through its WW domain; this interaction recruits ITCH from the cytoplasm to the surface of lipid droplets, leading to the ubiquitination of surface proteins. This process may impair lipid droplet degradation in hepatocytes (49), thereby promoting or exacerbating the onset and progression of MASLD. The regulatory mechanism of SMURF1 in lipid metabolism is particularly unique. Although considered a protective factor that is upregulated in human and mouse MASLD, SMURF1 deficiency alleviates hepatic steatosis. This occurs because SMURF1 interacts with SREBP-1c not to ubiquitinate and degrade it, but to occupy the binding site for its primary E3 ligase, FBW7A, thereby inhibiting SREBP-1c ubiquitination and degradation, which markedly exacerbates hepatic steatosis (50,51). However, another study observed that SMURF1 knockout leads to spontaneous hepatic steatosis and increased susceptibility to HFD-induced MASLD, associated with a sharp increase in PPARγ expression. Research on alcoholic steatohepatitis also suggests that SMURF1 deficiency restricts lipid degradation, thereby exacerbating hepatic lipotoxicity (52,53). These findings indicate that SMURF1 exerts multiple regulatory effects on lipid metabolism, though it is generally considered a protective E3 ligase in MASLD. SMURF2 participates in lipid metabolism by regulating C/EBPβ stability; under high-fat conditions, PRMT1 suppresses SMURF2 expression, thereby inhibiting the ubiquitination and degradation of C/EBPβ, which promotes preadipocyte proliferation and enhances lipid droplet formation induced by PPARγ pathway activation (54). Furthermore, WWP1 knockout mitigates hepatic lipid accumulation by reducing AKT phosphorylation levels (55).

A limited number of studies have reported that other HECT-type E3 ligases outside the NEDD4 subfamily are associated with lipid accumulation in MASLD/MASH. HUWE1, a prominent member of the other HECT family members, promotes lipid accumulation through multiple pathways: it directly binds and ubiquitinates PPARα for degradation, suppressing the fatty acid metabolism-related LXR/RXR/PPARα axis, a process modulated by PAQR3 and PAQR9 through competitive binding and significantly influenced by MCT1 via lactate metabolism (56-59); additionally, HUWE1 is linked to mTORC1 regulation, mediating WIPI2 ubiquitination and degradation (60), with both mechanisms collectively exacerbating hepatic steatosis. Meanwhile, UBE3A overexpression significantly exacerbates hepatic steatosis in MASLD by mediating ubiquitination of PDHA1 and ACAT1, reprogramming hepatic energy metabolism to promote cholesterol and triglyceride accumulation in the liver (61). These findings demonstrate that HECT-type E3 ubiquitin ligases exert precise and complex regulatory effects on lipid accumulation in MASLD by targeting multiple key lipid metabolism pathways, including SREBP, PPAR, JAK-STAT and mTOR.

The inflammatory response is a key driver of the progression from MASLD to MASH, and HECT-type E3 ubiquitin ligases play an important role in shaping the hepatic inflammatory microenvironment by regulating various inflammation-related signaling pathways. Members of the NEDD4 subfamily are involved in the regulation of inflammation. NEDD4 is upregulated under conditions of elevated hepatic free fatty acids and exacerbates liver inflammation and promotes atherosclerosis by ubiquitinating SR-BI and affecting high-density lipoprotein metabolism (40). NEDD4L exhibits significant dual effects in inflammatory regulation: Downregulated NEDD4L expression in MASH leads to impaired ubiquitination of thioredoxin interacting protein (Txnip), consequently enhancing ERS and promoting hepatocyte apoptosis (62); however, another study found markedly elevated NEDD4L levels in MASH that worsen inflammatory responses through LAPTM5 degradation and activation of the CDC42-MAPK axis (43). The association of ITCH with inflammation is equally complex. ITCH knockout mice fed a high-fat diet exhibit suppressed macrophage polarization and show protection against body weight gain and insulin resistance (45); however, further investigation revealed that ITCH knockout leads to elevated branched-chain amino acid (BCAA) levels, which promote inflammatory responses (48). This contradiction between macroscopic protective effects and microscopic pro-inflammatory effects highlights the complexity of ITCH within the global metabolic-immune network. SMURF1 indirectly participates in MASLD-related inflammation and fibrosis by regulating the TGF-β pathway (50-52,63). Notably, SMURF1 and SMURF2 also play a role in hepatic stellate cell (HSC) activation during liver fibrosis by interacting with talin1 (TLN1) and mediating its ubiquitination and degradation, thereby regulating the TLN1/FAK signaling axis and subsequently influencing HSC activation status and downstream inflammatory responses in models of hepatic ischemia-reperfusion injury associated with liver transplantation (64). SMURF2-mediated ubiquitination of AXIN1 attenuates AXIN1-induced Smad7 phosphorylation, inhibiting TGF-β-mTORC1 pathway activation and delaying MASLD-hepatocellular carcinoma (HCC) progression (63). TRIP12, an important member of the other HECT family members, exerts pro-inflammatory effects indirectly through the gut-liver axis: TRIP12 expression is upregulated in the intestines of MASLD and MASH mice, where it promotes K48-linked ubiquitination and degradation of nuclear transcription factor EB, leading to intestinal barrier disruption, increased permeability and worsened MASH severity (65). Additionally, in a proteomic analysis of alcohol-induced hepatic steatosis, TRIP12 was identified as the most significantly altered protein (66), suggesting that it may also participate in local hepatic pathological processes. Although direct studies of HERC subfamily members in MASLD/MASH are limited, their RCC1-like domain-mediated GTPase regulation and known functions in DNA repair, antiviral immunity and metabolic regulation suggest that they may exert protective or reparative functions in pathological processes such as liver injury and hepatitis (30,31). While other HECT members, including HECTD1-4, UBR5 and HACE1, have not been extensively studied in MASLD/MASH, their roles in regulating energy metabolism, ERS, oxidative stress, and cell death in other disease models provide important clues for future exploration of these E3 ligases in the context of MASLD inflammation (34,35).

Synthesizing the aforementioned findings, HECT-type E3 ubiquitin ligases exhibit a distinct dual role in the pathogenesis of MASLD/MASH, wherein the same E3 ligase can both promote and inhibit disease progression. This functional duality reflects the complexity and spatiotemporal specificity of the regulatory networks involving HECT family members. First, members of the same NEDD4 subfamily target different substrates in distinct cellular environments or disease stages, producing diametrically opposite effects. NEDD4L serves as a paradigmatic example: In MASH, NEDD4L downregulation impairs degradation of Txnip and JAK2, activating ERS and the JAK-STAT pathway respectively, thereby promoting apoptosis and steatosis (42,62); however, in advanced MASH, upregulated NEDD4L recognizes the PY motif of LAPTM5 through its WW domain, leading to the ubiquitination and degradation of LAPTM5, and subsequently activates the CDC42-MAPK axis, exacerbating lipid accumulation and inflammation (Fig. 2) (43). Such substrate-switching mechanisms may be determined by cell type, subcellular localization, post-translational modifications, or interactions with accessory proteins, enabling NEDD4L to play both disease-promoting and disease-suppressing roles at different disease stages.

This E3 ligase can exert functions contrary to its classical enzymatic activity through non-canonical mechanisms. The regulatory mechanisms of SMURF1 provide multiple illustrations. Although considered a protective E3 ligase, SMURF1 stabilizes SREBP-1c by occupying the binding site for its primary E3 ligase FBW7A, rather than through classical ubiquitin-mediated degradation, a 'steric hindrance' mechanism that paradoxically promotes steatosis (50,51); simultaneously, SMURF1 exerts protective effects by suppressing PPARγ, while in HSCs, SMURF1 performs protective functions by inhibiting HSC activation and inflammatory responses through ubiquitination and degradation of TLN1. These findings indicate that the effect of SMURF1 ultimately depends on the dynamic balance of different substrates in specific cell types and under specific metabolic stresses (52,64).

ITCH also plays a 'dual role' in MASLD/MASH, revealing the holistic and complex nature of HECT-type E3 ligase function. This is demonstrated by the marked hepatoprotective effect observed in systemic ITCH knockout mice (45), while human clinical samples show a negative correlation between ITCH protein abundance and hepatic steatosis severity (46). Paradoxically, ITCH knockout-induced alterations in BCAA profiles promote inflammation (Fig. 2) (48). This suggests that ITCH simultaneously suppresses lipid accumulation and exerts pro-inflammatory effects within the MASLD/MASH environment, making it difficult to determine whether ITCH possesses therapeutic potential in MASLD/MASH.

These findings suggest that a single E3 ligase may exert opposing effects across the disease spectrum. Specific E3s, notably NEDD4, NEDD4L, and ITCH, engage diverse substrates across different cell types, simultaneously influencing pro-disease and protective outcomes. Their ultimate functional output is determined by the integration of dominant microenvironmental cues, cellular metabolic status and tissue-specific stress responses. This pervasive duality indicates that HECT-type E3 ligases are not mere linear promoters or suppressors of disease but function as dynamic, environmentally responsive molecular hubs. Their activity is precisely calibrated by a confluence of factors, including disease stage, cellular origin, metabolic pressure and local tissue microenvironment. In conclusion, HECT-type E3 ubiquitin ligases act as complex regulators of MASLD/MASH, with functional duality arising from substrate diversity, spatiotemporal expression, non-canonical catalytic activities and trans-organ regulatory networks. Defining a single ligase simply as protective or pathogenic is incomplete.

The involvement of HECT-type E3 ligases in liver fibrosis demonstrates their functional diversity and context-dependent activities. First, distinct family members can exert opposing effects within the same pathological process. It remains challenging to develop targeted therapies by focusing on a specific domain of NEDD4 or a particular HERC subfamily member. For instance, SMURF1/2 exhibits potentially opposing effects on fibrosis compared with NEDD4/4L and WWP2. HERC5/HERC6 also show opposing actions. Second, a single E3 ligase can exhibit functional contradictions, as exemplified by SMURF2, which has been reported to be both profibrotic and antifibrotic, depending on the experimental model and mechanistic context. Collectively, these factors contribute to failure of domain-targeted therapies.

Multiple HECT-type E3 ligases have been implicated in HBV infection and exhibit complex and often opposing functions via distinct mechanisms. Several HECT-type E3 ligases promote HBV replication and release through various strategies. ITCH expression is decreased in HBV-positive human cells, leading to suppressed ubiquitination of Notch1 and subsequent aberrant activation of the Notch pathway, which promotes transcription of covalently closed circular DNA (cccDNA) and enhances HBV replication (82). HERC5 is hijacked by the HBV X protein (HBx), which directly binds HERC5 for ISGylation modification, conferring enhanced replication capability and interferon-α resistance to HBV (83). UBR5 mediates K29-linked ubiquitination of the HBV core protein, which may be involved in regulating HBV transcription, RNA encapsidation, reverse transcription and viral release, although the functional consequences require further elucidation (84).

Conversely, other HECT-type E3 proteins exert protective effects by targeting viral proteins for degradation. NEDD4 directly induces proteasomal degradation of HBx via K48-linked ubiquitination, suggesting an antitumor role in HBV-associated HCC (85). However, NEDD4 exhibits functional duality as it also interacts with and ubiquitinates γ2-Adaptin, promoting HBV nucleocapsid assembly and disrupting the multivesicular body (MVB) pathway, with NEDD4 knockdown significantly reducing HBV release (86-88). Thus, NEDD4 simultaneously facilitates viral production while suppressing viral oncoprotein stability. E6AP forms a p53/E6AP/HBx ternary complex that leads to HBx ubiquitination and degradation, and all-trans retinoic acid activates this pathway to suppress HBV replication (89,90).

HCV infection involves multiple HECT-type E3 ligases acting via diverse mechanisms that collectively influence viral replication, particle release and disease progression. Several HECT-type E3s promote HCV propagation. ITCH plays a proviral role where increased reactive oxygen species (ROS) induce ITCH phosphorylation, leading to ubiquitination of the ATPase VPS4A, a protein necessary for MVB formation, thereby promoting HCV particle release (91). HERC5's role remains controversial, with some studies reporting that HERC5 specifically promotes NS5A ISGylation, thereby enhancing HCV replication (92), while others demonstrate that the inhibitory effect of ISG15 on HCV RNA replication does not require HERC5 binding (93).

By contrast, other HECT-type E3s exert anti-HCV effects. E6AP represents the most extensively studied protective factor, initially characterized as promoting HCV pathogenesis through nonstructural protein 5B (NS5B)-recruited ubiquitination of retinoblastoma tumor suppressor protein (pRb) leading to chromosomal instability (94), but subsequently established as an anti-HCV replication factor through direct binding and ubiquitination of the viral nucleocapsid core protein (95,96). The HCV core protein suppresses E6AP expression via DNA promoter methylation to evade ubiquitin-proteasome system-mediated degradation, while p53 upregulation enhances E6AP expression and recruits E6AP to form a ternary complex with the HCV core, promoting its ubiquitination and degradation (97-99). All-trans retinoic acid activates E6AP by inhibiting its methylation, thereby suppressing HCV replication (100).

SMURF2 has dual functions in HCV infection. It enhances ubiquitination and degradation of the viral protease NS3-4A, inhibiting TGF-β-induced Smad2/3 phosphorylation and alleviating HCV-associated fibrosis (101). However, the SMURF2/NS3-4A complex also interacts with quinolinate phosphoribosyl-transferase (QPRT), leading to QPRT degradation, which lowers the NAD⁺/NADH ratio, potentially shifting lipid metabolism to favor HCV replication (102).

Current evidence for HECT-type E3 involvement in HAV infection primarily focuses on a single mechanism. ITCH facilitates HAV release, and its catalytic activity is essential for the efficient release of quasi-enveloped virions (eHAV). During acute infection, eHAV is released into the blood via the MVB pathway, which involves an interaction between the pX domain and the ESCRT complex and the HECT domain of ITCH, resulting in inhibition of viral activity and release (103).

Synthesizing the findings across different hepatitis viruses reveals that HECT-type E3 ubiquitin ligases exhibit a striking dual role in viral hepatitis, wherein the same E3 ligase can both promote and inhibit viral replication, host pathogenesis, or antiviral responses, depending on the viral context, cellular environment, and target substrate specificity. This functional duality profoundly reflects the evolutionary arms race between host antiviral defense and viral evasion strategies.

The same HECT-type E3 ligases can exert opposing effects on the same virus by targeting different viral or host proteins. NEDD4 in HBV infection provides a paradigmatic example: While NEDD4 promotes HBV nucleocapsid assembly and viral release through γ2-Adaptin ubiquitination and HBc modification (86-88), it simultaneously induces proteasomal degradation of HBx via K48-linked ubiquitination, potentially suppressing HBV-associated hepatocarcinogenesis (85). This substrate-specific duality positions NEDD4 as both a facilitator of viral production and a suppressor of viral oncoprotein stability, suggesting that its net effect may depend on the relative abundance and accessibility of different substrates during different stages of infection.

The same E3 ligase plays different roles in distinct hepatitis viruses, reflecting virus-specific coevolutionary adaptations. ITCH exemplifies this virus-dependent functional diversity: In HAV infection, ITCH facilitates eHAV release through ESCRT pathway interactions (103); in HCV infection, ROS-induced ITCH phosphorylation promotes VPS4A ubiquitination and viral particle release (91); whereas in HBV infection, decreased ITCH expression leads to Notch1 stabilization and enhanced cccDNA transcription (82). This virus-specific engagement suggests that therapeutic targeting of ITCH would require careful consideration of viral context.

E6AP demonstrates how the same E3 ligase can be reinterpreted from pathogenic to protective as mechanistic understanding deepens. Initially characterized as a pro-HCV factor through NS5B-mediated pRb ubiquitination, promoting chromosomal instability (94), E6AP was subsequently identified as an anti-HCV factor through direct HCV core ubiquitination and degradation (95,96). The discovery of the p53/E6AP/viral core axis operating in both HCV and HBV infections unified these observations, establishing E6AP as a host restriction factor whose activity is subverted by viral proteins through epigenetic suppression (97-99).

HERC5 exemplifies the controversy arising from the context-dependent mechanisms. Conflicting studies on HERC5's role in HCV replication and whether it inhibits or promotes viral RNA synthesis through NS5A ISGylation (92,93) may reflect differences in experimental systems, viral strains, or the balance between the direct antiviral effects and proviral consequences of ISGylation.

In conclusion, HECT-type E3 ubiquitin ligases function as critical nodes at the host-virus interface during viral hepatitis, with their dual roles arising from substrate diversity, virus-specific adaptations and the complex interplay between viral life cycle requirements and host antiviral defenses. However, owing to the presence of the aforementioned factors, therapeutic approaches that simply target a specific enzyme or HECT domain cannot be developed. This limitation has constrained the advancement of viral hepatitis treatments.

In studies investigating the function of HECT-type E3 ligases in HCC, multiple studies have confirmed NEDD4 as an oncogene that mediates the ubiquitination and degradation of the classical tumor suppressor PTEN, accelerating HCC progression (106-110). Two clinical studies found that patients with HCC and high NEDD4 expression had shorter survival times, and subsequent experimental validation indicated that NEDD4 upregulation suppresses PTEN expression, thereby promoting PI3K-AKT phosphorylation (106,107). This finding was corroborated by Zhou et al (108), who also suggested that the NEDD4/PTEN/PI3K/AKT axis is modulated by the interaction between craniofacial development protein 1 and NEDD4 (108). Beyond PTEN, some research shows that NEDD4 also promotes HCC invasion and metastasis by ubiquitinating large tumor suppressor kinase 1 (LATS1) in the HIPPO pathway, and TGF-β type I receptor (TGFBR1) in the TGF-β pathway (109,110). Therefore, NEDD4 is currently the most promising HECT-type E3 enzyme to serve as a novel therapeutic target for HCC.

In studies of other HECT-type E3 ligases, HECTD1/2 appear to promote HCC proliferation, although they have been reported to do so through distinct mechanisms. HECTD1 targets growth factor receptor-bound protein 2 (GRB2), and its degradation inhibits MAPK pathway-mediated angiogenesis, although this ubiquitination is competitively inhibited by elevated Golgi protein 73 levels in HCC (111). HECTD2 is highly expressed in HCC and promotes proliferation through KEAP1 ubiquitination and subsequent antioxidant response activation (112). Liu et al (113) suggested that E6AP interacts with G protein-coupled receptor 26 (GPR26) through its HECT domain. By altering the conformation of this domain, E6AP induces ubiquitination at the K286 site of GPR26, thereby promoting tumor proliferation (113). UBR5 interacts with the tumor suppressor esophageal cancer-related gene 4 (ECRG4). ECRG4 overexpression conversely downregulates UBR5 expression, leading to increased p21 levels and anti-proliferative effects (114). Wang et al (115) found that targeting and inhibiting UBR5 with Echinacoside promoted apoptosis and inhibited glycolysis in HepG2 cells (115). Leboeuf et al (116) demonstrated that simultaneous small interfering RNA silencing of UBR1, UBR2, UBR4 and UBR5 inhibited the Arg/N-degron pathway, reducing HCC cell proliferation while increasing spontaneous apoptosis (116). HACE1 was initially found to be downregulated in HCC tissues, and patients with low HACE1 expression had lower overall survival rates (117). Mechanistic studies revealed that HACE1 mediates the ubiquitination of SREBP cleavage-activating protein (SCAP), thereby inhibiting SREBP2-mediated cholesterol biosynthesis, which is crucial for membrane synthesis during rapid proliferation. However, this process is antagonized by the ZDHHC3-mediated S-acylation of SCAP at Cys264 (118). Additionally, LINC00161 recruits EZH2, leading to HACE1 methylation and promotion of HCC progression (119). Therefore, HACE1 is currently recognized as a tumor suppressor that inhibits HCC proliferation. These HECT-type E3 ligases have been extensively studied for their roles in regulating HCC proliferation; however, no significant bidirectional effects have been observed. Consequently, they represent promising novel therapeutic targets for HCC diagnosis and treatment, although they currently lack the level of recognition attained by NEDD4.

The HECT-type E3 ligases described below have demonstrated dual functionality in current research, representing a significantly controversial aspect of HCC proliferation studies, with findings potentially exhibiting both promoting and inhibitory effects. Focus was first addressed on studies of other HECT-type E3 ubiquitin ligases within the NEDD4 subfamily, excluding NEDD4 itself, that regulates HCC proliferation. The role of NEDD4L in HCC proliferation remains controversial. Lee et al (120) demonstrated that NEDD4L expression is significantly upregulated upon activation of the canonical cancer-related Wnt/β-catenin pathway. Conversely, Zhao et al (121) reported that downregulation of NEDD4L in HCC tissues correlates with poor prognosis. The authors proposed that NEDD4L binds to ERK1/2 not to ubiquitinate it but rather to promote its phosphorylation and activation, thereby inducing apoptosis and inhibiting HCC cell proliferation (121). Another study also characterized NEDD4L as a tumor suppressor, attributing this role to its ability to ubiquitinate cyclin-dependent kinase 5 (CDK5). The same study indicated that ubiquitin-specific peptidase 1 (USP1) acts as a deubiquitinating enzyme, counteracting NEDD4L's effect on CDK5 (122). Research on ITCH in HCC proliferation is relatively limited but suggests tumor-suppressive functions. Ge et al (123) found that ITCH ubiquitinates transforming growth factor β-activated kinase 1 (TAK1). TAK1 degradation leads to inhibition of the MAPK and NF-κB pathways, which are critical for cell proliferation and survival. The glycosyltransferase Rab-like GTPase activating protein and GRAM domain-containing protein 4 (GRAMD4) recruits ITCH, thereby promoting TAK1 degradation and inhibiting HCC progression (123). Additionally, ITCH has been linked to programmed cell death ligand 1 (PD-L1) regulation through GLI1 ubiquitination, although this primarily affects immune evasion rather than direct proliferation control (124). Conflicting reports exist regarding the role of WWP1 in HCC proliferation. Cheng et al (125) identified WWP1 as an oncogenic factor and found that its expression was elevated in tissues of patients with HCC and positively correlated with alpha-fetoprotein levels. WWP1 knockout led to HCC cell apoptosis and inhibited proliferation via the upregulation of caspase-3 and P53, although the involvement of ubiquitination was not clarified (125). By contrast, another study reported a tumor-suppressive role for WWP1, associated with its binding to and degradation of pyruvate kinase M2 (PKM2), inhibiting HCC cell proliferation. However, this interaction requires RhoGAP 24 (ARHGAP24) as a scaffolding protein (126). Furthermore, in HCC, WWP1 can be influenced by the deubiquitinating enzyme USP13, leading to activation of AKT and mTOR phosphorylation and accelerated HCC proliferation (127). Similar to NEDD4, WWP2 is also a key regulator of PTEN, cleavage and polyadenylation specificity factor 7 is upregulated, inducing the upregulation of WWP2 and promoting the ubiquitination and degradation of PTEN (128). However, the current body of research on WWP2 remains insufficient, and its potential as a therapeutic target warrants further investigation.

Research on the role of the HERC subfamily in HCC proliferation, although limited, suggests pro-tumorigenic functions. HERC2 is significantly upregulated in primary hepatocytes under inflammatory stimulation, promoting STAT3 activation during the inflammation-to-cancer transition and enhancing cancer stemness (129). HERC4 is elevated in HCC tissues, enhancing cell proliferation through the suppression of Salvador 1 (SAV1) (130,131). HERC5 is also upregulated in HCC, promoting cancer cell proliferation and influencing apoptosis. Wang et al (132) synthesized a disubstituted quinazoline derivative, HZ-6d, which binds to HERC5, inhibits its expression, and activates P53, effectively delaying HCC growth in vivo (132). Interestingly, integrated omics and clinical data analyses identified HERC5 as a prognostic marker for post-liver transplant recurrence in patients with HCC, with low expression associated with poor prognosis because of immune escape mechanisms (133).

HUWE1, TRIP12 and UBE3B/C regulate HCC proliferation. HUWE1 mediates p53 degradation and participates in the regulation of HCC cell apoptosis, which is modulated by the deubiquitinating enzyme USP7 (134). Similar to HUWE1, TRIP12 is also regulated by USP7-mediated de-ubiquitination. Overexpression of USP7 blocks TRIP12-mediated ubiquitination and degradation of p14ARF, promoting HCC proliferation (135). A large-scale population sequencing study found that UBE3C overexpression in HCC tissues was significantly associated with reduced survival, and that UBE3C promoted HCC progression by regulating epithelial-mesenchymal transition (EMT), which indirectly affects proliferation (136). Tao et al (137) also described UBE3C upregulation in HCC and its regulation by the upstream miR-542-3p. Although these genes have also been shown to potentially regulate HCC proliferation, the current body of research remains insufficient. Therefore, it remains unclear whether they exhibit bidirectional effects.

HECT-type E3 ubiquitin ligases play critical roles in HCC invasion and metastasis by regulating cell migration, adhesion, EMT and ECM remodeling. NEDD4 promotes HCC cell invasion and metastasis via multiple mechanisms. Beyond its role in proliferation, research shows that NEDD4 ubiquitinates LATS1 in the HIPPO pathway and TGFBR1 in the TGF-β pathway, thereby relieving the tumor-suppressive effects of these pathways and enhancing the migratory and invasive capacity of HCC cells (109,110). The HIPPO pathway is a key regulator of organ size and metastasis, and its inactivation by NEDD4-mediated LATS1 degradation promotes nuclear translocation of YAP/TAZ, driving EMT and metastasis. Similarly, TGFBR1 ubiquitination by NEDD4 modulates TGF-β signaling, which has dual roles in HCC but often promotes metastasis in advanced stages.

SMURF1 and SMURF2 predominantly exert inhibitory effects on invasion and metastasis in HCC, particularly through negative regulation of the TGF-β signaling pathway, which is a major driver of EMT and metastasis. Liu et al (138) demonstrated that in HCC, SMURF1/2 binds to TGFBR1, inducing its ubiquitination and degradation, which suppresses the TGF-β pathway and its pro-metastatic effects. TGF-β Receptor associated protein 1 (TGFBRAP1) competes with SMURF1/2 for the TGFBR1 binding site, thereby enhancing cancer stemness and metastatic potential. Similarly, Niemann-Pick disease, type C1 protein exerts an effect analogous to TGFBRAP1 (138,139). SMURF1 can also suppress TGF-β pathway activation by binding to and ubiquitinating TGFBR2 (140). Beyond TGF-β signaling, SMURF1 ubiquitinates ultra-violet radiation resistance associated, which inhibits HCC proliferation and metastasis by promoting autophagosome maturation and lysosomal degradation of EGFR, a key driver of metastatic signaling (141). Signal transducer and activator of transcription 1, which can have tumor-suppressive functions in HCC, is also targeted for degradation by SMURF1, but this process depends on the presence of SIRT 7. In addition, hsa_circ_0000235 participates in SMURF1's regulation of HSP60 in a manner similar to that of SIRT 7 (142,143). These findings collectively support the metastasis-suppressive role of SMURF1 in HCC. Therefore, the aforementioned studies conclusively demonstrate the roles of NEDD4 and SMURF1/2 in the regulation of HCC invasion and metastasis. Notably, NEDD4 has emerged as a well-defined target that promotes proliferation, invasion and metastasis and has significant translational potential.

The functions of HECT-type E3 enzymes remain controversial because of limited studies and their bidirectional activity. ITCH indirectly inhibits HCC invasion and metastasis through TAK1 ubiquitination and degradation, which leads to inhibition of the MAPK and NF-κB pathways, both of which can promote EMT and metastasis. GRAMD4 recruits ITCH to promote this process, thereby suppressing HCC progression (123). Additionally, the ITCH/GLI1 axis may indirectly influence HCC metastatic potential by regulating PD-L1 expression and subsequent immune evasion; however, its direct effects on cell migration remain to be explored (124). The role of WWP1 in HCC invasion and metastasis is unclear. One study reported that WWP1, through binding to and degradation of PKM2, inhibits HCC cell migration and invasion, requiring ARHGAP24 as a scaffolding protein (126). However, the oncogenic functions of WWP1 described in other studies suggest context-dependent effects on metastasis, which require further investigation. HERC4, through suppression of SAV1, may influence HCC metastasis, as SAV1 is a core component of the HIPPO pathway that regulates YAP/TAZ activity and subsequent EMT (130,131). UBE3C overexpression promotes HCC progression by regulating EMT in HCC cells, directly implicating it in metastatic spread (136). UBR5 influences HCC cell migration through the regulation of the Arg/N-degron pathway, as simultaneous silencing of UBR1, UBR2, UBR4 and UBR5 reduces HCC cell migration (116).

HECT-type E3 ubiquitin ligases significantly contribute to therapeutic resistance in HCC by affecting the responses to multiple classes of anticancer drugs, including targeted therapies and chemotherapy. NEDD4 plays an important role in drug resistance by regulating the PTEN/PI3K/AKT axis. Inhibition of NEDD4/PTEN axis-mediated autophagy can enhance the sensitivity of HCC animal models and cell lines to sorafenib, a first-line multi-kinase inhibitor used in advanced HCC (107,108,144). Autophagy is a cellular survival mechanism that promotes resistance to therapy, and NEDD4-mediated PTEN degradation activates AKT, which in turn promotes autophagy and survival under stress conditions. In the context of the FOLFOX (5-fluorouracil, oxaliplatin and leucovorin) chemotherapy regimen, glycolysis-enhanced histone lactylation forms an H3K14la/NEDD4/PTEN axis regulating multidrug resistance in HCC (145). This epigenetic mechanism links metabolic reprogramming to drug resistance through NEDD4 upregulation. Notably, this study indicated that NEDD4L is not involved in PTEN regulation in this context (146).

The following enzymes may be underrepresented because of the limited number of studies on drug resistance. SMURF1/2 may indirectly influence drug resistance through their regulation of TGF-β signaling. TGFBRAP1 competes with SMURF1/2 for TGFBR1 binding, thereby enhancing cancer stemness and HCC resistance to regorafenib, another multi-kinase inhibitor used in second-line treatment for HCC (138). HECTD2 is highly expressed in lenvatinib-resistant HCC cell lines, patient tissues, patient-derived organoids and xenografts. Lenvatinib is another multi-kinase inhibitor used as a first-line treatment for advanced HCC. HECTD2 promotes ubiquitination of KEAP1, a negative regulator of antioxidant response. KEAP1 degradation leads to the activation of NRF2 and the subsequent antioxidant response, which reduces oxidative stress-induced cell death and decreases sensitivity to lenvatinib in HCC cells (112). WWP2 contributes to doxorubicin resistance in HCC through its regulation of PTEN. METTL3-mediated m6A modification of WWP2 increases its stability, leading to enhanced PTEN ubiquitination and AKT activation, which counteracts the antitumor effects of doxorubicin, an anthracycline antibiotic used in HCC chemotherapy (147). UBE3B is associated with lenvatinib resistance in HCC. Lobeline, an m6A antagonist, downregulates UBE3B expression, partially reversing lenvatinib resistance. This suggests that UBE3B expression, which is potentially regulated by m6A modifications, contributes to the resistant phenotype (148).

Synthesizing the aforementioned findings, HECT-type E3 ubiquitin ligases exhibit a dual-role effect in the pathogenesis of HCC across all aspects of tumor biology, including proliferation, invasion, metastasis and drug resistance. Different members of the same subfamily, and even the same E3 ligase in different cellular contexts or targeting different substrates, can either promote or suppress HCC progression. This functional duality profoundly reflects the intricate regulation of complex signaling networks in HCC and the functional diversity of the HECT family members.

NEDD4 acts as a definitive oncogene by degrading PTEN to activate PI3K-AKT signaling and promote proliferation (107,108). It also enhances invasion and metastasis by ubiquitinating LATS1 and TGFBR1 (109,110) and contributes to drug resistance through autophagy regulation and epigenetic mechanisms (107,108,144). Although it is now widely recognized that inhibiting NEDD4 significantly suppresses tumor proliferation, invasion and metastasis while improving drug resistance, no studies have described its dual role in HCC treatment, demonstrating its immense translational potential. Unfortunately, no clinical trials have confirmed whether this effect can be effectively applied to HCC therapy.

Unlike NEDD4, the following genes fully demonstrate the complexity of HECT-type E3 ubiquitin ligases in HCC progression. WWP1 illustrates how the same E3 ligase may exhibit contradictory roles owing to context-dependent interactions and substrate availability. WWP1 has been reported both as an oncogene promoting proliferation through caspase-3 and P53 regulation (125), and as a tumor suppressor that inhibits migration and invasion through PKM2 degradation in an ARHGAP24-dependent manner (126). This functional switch may depend on the presence of scaffolding proteins such as ARHGAP24, which direct WWP1 to specific substrates, and on the cellular context that determines which substrates are available or relevant. The involvement of USP13 in the regulation of WWP1 stability and subsequent AKT/mTOR activation further complicates its functional characterization (127). Other HECT members exhibit specialized roles in specific aspects of HCC biology. HECTD1 regulates angiogenesis through GRB2 modulation (111). HECTD2 promotes proliferation and lenvatinib resistance through KEAP1 ubiquitination and NRF2 activation (112). E6AP enhances tumor activity through GPR26 degradation (113). UBR5 affects proliferation, apoptosis, glycolysis and migration through multiple mechanisms (115,116). HACE1 acts as a tumor suppressor by regulating cholesterol metabolism and immune evasion through SCAP ubiquitination, with its activity antagonized by S-acylation (118). HUWE1 and TRIP12 regulate apoptosis through the p53/p14ARF axis and are modulated by USP7 (134,135). UBE3C promotes EMT and metastasis (136), while UBE3B contributes to lenvatinib resistance (148). This functional specialization reflects the diverse roles of different HECT members within the complex signaling network of HCC.

From structural and functional perspectives, the domain architecture of HECT-type E3 ligases determines their substrate recognition specificity and functional outcomes in HCC. The WW domains of NEDD4 and WWP1/2 mediate the recognition of substrates containing PPxY motifs, such as PTEN, enabling these ligases to regulate the PI3K-AKT pathway. The C2 and WW domains of SMURF1/2 enable interaction with membrane-associated proteins such as TGF-β receptors, allowing precise regulation of TGF-β signaling. The RCC1-like domains of the HERC family confer the ability to regulate GTPase-related signaling and respond to inflammatory stimuli. Through their diverse N-terminal domains, other HECT members participate in the regulation of fundamental biological processes including metabolism (HACE1), stress responses (HECTD2) and cell death (HUWE1 and TRIP12). This structure-function relationship explains the molecular basis of the protective or pathogenic roles of different HECT members in HCC. In conclusion, HECT-type E3 ubiquitin ligases function as critical regulatory nodes in HCC, influencing tumor proliferation, invasion, metastasis, and drug resistance through diverse mechanisms. Their functional duality arises from substrate diversity, context-dependent interactions, scaffolding protein involvement, post-translational modifications and tumor microenvironment.