Numerous carcinogenic or tumor-suppressive proteins involved in tumorigenesis, metastasis, or drug resistance are regulated by S-palmitoylation. Studies have shown that zDHHC9-mediated S-palmitoylation of oncogenic NRAS is essential for its localization on the plasma membranes and downstream signaling activation, promoting the occurrence of leukemia (34, 35). APT1 has been found to depalmitoylate H-Ras and N-Ras, reducing their affinity to the plasma membrane and releasing them from it, allowing diffusion back into the Golgi apparatus (36, 37). Therefore, the S-palmitoylation-depalmitoylation cycle of N-Ras and H-Ras regulates the transport of Ras between the Golgi apparatus and the plasma membrane ( Figure 2 ). EGFR, a growth factor receptor critical for cell proliferation and differentiation (38), S-palmitoylation of EGFR by zDHHC1, zDHHC2, and/or zDHHC21 on Cys797 is essential for the stability, membrane localization, dimerization, and activation of this receptor and increases cell migration and anchor-independent growth in lung cancer (39). Fatty acid synthase (FASN), a major lipogenic enzyme that synthesizes palmitic acid (40), induces an oncogenic phenotype when overexpressed in normal cells. This transformation involves enhanced lipid synthesis and an increase in phosphorylation and expression of EGFR (41). EGFR activation is involved in FASN-dependent palmitoylation, which is required for both ligand-dependent and ligand-independent activation of EGFR and occurs within the cell. Treatment of prostate and lung cancer cells with the FASN inhibitor cerulenin or the palmitoylation inhibitor 2-bromopalmitate (2-BP) reduces EGFR levels on the plasma membrane while increasing lysosome levels (42). A recent published study also reported the lipid-rich microenvironment activates palmitate biosynthesis in metastatic colorectal cancer cells by up-regulating FASN. The increased intracellular palmitate promotes EGFR palmitoylation, thereby activating EGFR signaling. The FASN inhibitor orlistat inhibits EGFR palmitoylation and inhibits the stem cell properties of colorectal cancer (CRC) cells (43). EGF stimulates the S-palmitoylation of the oncogenic protein CDCP1, thereby blocking the degradation of CDCP1 (44). This study establishes a connection between S-palmitoylation events and EGFR signaling. However, further research is required to fully understand the specific mechanisms involved.

The membrane protein claudin-3 (CLDN3) is crucial for the formation and maintenance of tight junctions and is highly expressed in various cancers (45). In ovarian cancer, zDHHC12 mediates S-palmitoylation of CLDN3 at C181, C182, and C184, which contributes to its proper localization and stabilization on the plasma membrane. zDHHC22 is significantly upregulated in renal cancer cells (RCC) resistant to tyrosine kinase inhibitors (46). In RCC, zDHHC2 mediates AGK S-palmitoylation, facilitating AGK translocation to the plasma membrane and activating the PI3K-AKT-mTOR signaling pathway (47). These findings indicate the existence of a zDHHC2-AGK signaling axis and suggest that targeting zDHHC2 could enhance the effectiveness of sunitinib in RCC treatment (47). zDHHC22 expression is significantly reduced in estrogen receptor-negative breast cancer tissues, and its expression is positively correlated with the clinical prognosis of breast cancer patients (46). Through S-palmitoylation, zDHHC22 decreases mTOR stability and suppresses AKT signaling pathway activation. Restoring zDHHC22 expression sensitizes tamoxifen-resistant MCF-7 cells to tamoxifen treatment (46). Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a crucial role in cholesterol metabolism and antitumor immune responses (48). Abnormal upregulation of PCSK9 promotes cell proliferation and sorafenib resistance in HCC. zDHHC16-mediated S-palmitoylation of PCSK9 at cysteine 600 enhances the binding affinity between PCSK9 and tensin homolog (PTEN) (49). S-palmitoylation of PCSK9 leads to lysosome-mediated PTEN degradation and subsequent AKT activation (50). Inhibiting PCSK9 S-palmitoylation suppresses AKT phosphorylation and enhances the antitumor effects of sorafenib (49).

P53 is a well-known tumor suppressor that regulates cell proliferation and apoptosis. The S-palmitoylation of p53 at the C135, C176, and C275 residues, mediated by zDHHC1, has been reported to play necessary roles in p53 nuclear trafficking and subsequent pathway activation ( Figure 1A ) (51). Conversely, p53 epigenetically regulates zDHHC1 expression by recruiting DNMT3A and HDAC1 to the promoter region (51). This regulatory feedback loop could help explain the function of zDHHC family members and may be utilized for targeted therapy.

Apoptosis, a cell-intrinsic mechanism for suicide, plays a pivotal role in inflammatory diseases and tumors (52). Recent studies have identified dysregulated S-palmitoylation of apoptosis-related proteins in many human cancers. For instance, the TNF family receptor Fas can undergo S-palmitoylation, as revealed by metabolic labeling of [3H] palmitate in HEK293 cells (53, 54). zDHHC7-mediated S-palmitoylation of Fas leads to an increase in the active form of caspase-8 (55). Decreased expression of zDHHC7 inhibits the expression of Fas in the plasma membrane of SW480 cells, thereby suppressing FASL-induced apoptosis. While Fas is located on various cell surfaces, Fas ligand (FasL) is primarily found on activated T cells and natural killer (NK) cells (56). Dysregulation of Fas/FasL signaling compromises immune function. FasL can be palmitoylated at residue cysteine 82, and mutants at this site show significantly lower cell death compared to controls (57). S-palmitoylation of the Fas signaling member Lck is essential for Fas-mediated apoptosis (58). The binding of the Fas receptor to its ligand Fas results in a rapid and substantial increase in the S-palmitoylation level of the tyrosine kinase Lck. Inhibition of Lck S-palmitoylation not only disrupts proximal Fas signal transduction events but also confers resistance to Fas-mediated apoptosis. zDHHC21, a palmitoyltransferase, functions as a PAT that controls Lck activation. Knockout of zDHHC21 eliminates the activation and downstream signaling of Lck following Fas receptor stimulation (59).

Death receptor-6 (DR6) is a newly discovered member of the Death receptor subfamily that can induce apoptosis and/or activate NF-κB and JNK/SAPK family stress kinases in specific cell types (60). The S-palmitoylation of DR6 occurs at Cys368, but the relationship between DR6 S-palmitoylation and apoptosis requires further investigation. In COS7 cells, the increase in caspase-6 S-palmitoylation at the C264 and C277 sites, mediated by zDHHC17, leads to a decrease in caspase-6 activation (61). Additionally, S-palmitoylation can also occur in Bcl-2 family proapoptotic proteins. S-S-palmitoylation of BAX at Cys-126 is crucial for the apoptosis process as it affects its localization to the mitochondria (62). The mutation C126 significantly reduces the number of apoptotic cells and the activation of caspase-3. These findings collectively indicate that S-palmitoylation plays an essential role in the apoptosis process of immune cells and tumor cells.

Aberrant S-palmitoylation has been shown to contribute to tumor initiation and growth, and its potential role in tumor metastasis has been investigated. Tetraspanins, which are small membrane proteins, have been found to promote the adhesion and metastasis of tumor cells (63). The S-palmitoylation of integrin subunits beta-4 (ITGB4), alpha-3 (ITGA3), and alpha-6 (ITGA6) has an impact on the formation of integrin tetraspanin complexes (64). Additionally, ITGA6 and ITGB4 collaborate with EGFR and ERBB2 to promote various aspects of tumor progression and metastasis (65). It is crucial to conduct studies to determine how different S-palmitoylation of proteins interferes with EGF receptor-integrin dynamics within tetraspanin complexes. Ras-related protein RAB27A is significantly overexpressed in oral squamous cell carcinoma (66). RAB27A regulates the S-palmitoylation of EGFR via zDHHC13 in oral squamous cell carcinoma (66). The laminin-binding integrin α6β4 plays a key role during tumor cell metastasis (67).

Several studies have indicated that the S-palmitoylation of α6β4 protein by zDHHC3 inhibits the degradation of α6β4. This inhibition is likely due to a decrease in endosomal exposure to cathepsin D (68). Flotillin-1 is a membrane-associated protein that plays a role in multi-cellular signaling events in cells (69). It has been observed that Flotillin-1 is overexpressed in numerous solid tumors, and its S-palmitoylation contributes to its stability and subsequent metastatic capabilities in breast cancer cells and experimental metastasis models (70, 71). Therefore, targeting flotillin-1 S-palmitoylation could be a promising approach for addressing breast cancer metastasis. MUC1, a mucin-like protein located on the apical membrane of epithelial cells, is also associated with tumor metastasis (72). S-palmitoylation of MUC1 modulates its recycling from endosomes to the plasma membrane (73), and the intracellular localization of MUC1 is altered during breast cancer metastasis (74). CCR5 is a CC chemokine receptor expressed on immune cells such as memory lymphocytes, macrophages, and dendritic cells. Weinberg and colleagues have discovered that CCR5 undergoes S-palmitoylation at three cysteine residues within its C-terminal region (75). Palmitoylated cysteines are known to have a significant role in the intracellular trafficking of CCR5. When CCR5 S-palmitoylation is eliminated, it reduces the surface expression of CCR5 by trapping the receptor in organelles, leading to its degradation. This is particularly relevant in breast cancer metastasis to bone, as CCR5 S-palmitoylation facilitates tumor metastatic spread through CCL5-CCR5 chemokine signaling. Another important factor in tumor metastasis is SMAD3, which is involved in the activation of the transforming growth factor-beta (TGF-β) signaling pathway (76). S-palmitoylation of SMAD3, mediated by palmitoyltransferase zDHHC19, promotes its activation. Furthermore, the interaction between SMAD3 and EP300 promotes the expression of mesenchymal markers in the mesenchymal subtype of glioblastoma multiforme (GBM). Therefore, targeting Smad3 S-palmitoylation could be a crucial molecular approach in combating tumor metastasis (77).

The relationship between cellular metabolic reprogramming and protein S-palmitoylation is inherently connected, as S-palmitoylation involves the covalent attachment of palmitic acid to proteins. In cancer cells, the addition of the saturated fatty acid palmitate enhances the synthesis of palmitoyl-CoA, thereby upregulating protein S-palmitoylation (78). This direct involvement of central metabolites in post-translational modifications allows tumor cells to integrate information from the microenvironment and effectively regulate cellular processes. Consequently, these palmitoylated metabolic proteins are believed to influence cancer cell metabolism. For instance, glucose transporter (GLUT1), a transmembrane protein responsible for glucose uptake, is frequently upregulated in various cancer types (79). The localization of GLUT1 to the plasma membrane is crucial for its function in glucose uptake. Studies have shown that zDHHC9 mediates the S-palmitoylation of GLUT1 at C207, promoting its plasma membrane localization and resulting in heightened glycolysis in glioblastoma (80). Malate Dehydrogenase (MDH2), a key enzyme in the TCA cycle, catalyzes the reversible conversion of malate to oxaloacetate. S-palmitoylation of MDH2 by zDHHC18 at C138 leads to increased MDH2 activity (81). This S-palmitoylation event activates mitochondrial respiration and promotes the malignancy of ovarian cancer. Additionally, zDHHC21 has recently been identified as a critical regulator of oxidative phosphorylation activation in acute myeloid leukemia cells. zDHHC21 specifically catalyzes the S-palmitoylation of mitochondrial adenylate kinase 2 (AK2) and activates OXPHOS in these leukemia cells (82). Depletion of ZDHHC21 induces the differentiation of acute myeloid leukemia cells and weakens their stemness potential (83).

Activated lipid metabolism has been found to promote cancer development both in vitro and in vivo (11, 84). Treatment with exogenous palmitate enhances Src-dependent mitochondrial β-oxidation, increases the level of Src kinase localized in the cell membrane, and activates Src-mediated downstream MAPK and FAK signaling (78). These results reveal that dietary palmitate, in collaboration with elevated Src kinase, accelerates prostate tumor progression (78). CD36, a fatty acid transporter, is widely expressed in immune cells and tumor cells (85). CD36 can be palmitoylated by zDHHC4 and zDHHC5 at both N-terminal and C-terminal cysteine residues (86). zDHHC4 and zDHHC5 function at different subcellular localizations and regulate CD36 S-palmitoylation, targeting it to the plasma membrane for fatty acid uptake (87). Inhibiting CD36 S-palmitoylation reduces the hydrophobicity of the CD36 protein and its localization in the plasma membrane of hepatocytes. Depletion of either zDHHC4 or zDHHC5 disrupts the capability of adipose tissues to uptake fatty acids (88). These findings demonstrate the critical role of zDHHC4 and zDHHC5 in regulating CD36 S-palmitoylation and fatty acid uptake.

IFN-γ is a cytokine secreted by various activated immune cells, including NK and CD8⁺ T cells (90). Binding of IFNγ to its receptor activates the JAK-STAT pathway, inducing the expression of classical interferon-stimulated genes that play a critical role in antitumor responses (90). Recent research has discovered that S-palmitoylation of IFN-γ receptor 1 (IFNGR1) at cysteine 122 acts as a sorting signal for IFNGR1 lysosomal degradation, mediated by AP3D1 in CRC (91, 92). OPTN, which is lost in early-stage human CRC, interacts with AP3D1 to hinder its recognition of IFNGR1, thereby maintaining IFNGR1 stability and the integrity of downstream MHC-I signaling. This promotes antigen presentation to T cells and inhibits CRC progression (92).

NOD1/2 are intracellular pattern-recognition proteins that recognize peptidoglycan associated with microorganisms. The activation of NOD signaling is crucial for host defense against infections (93). Studies have demonstrated that zDHHC5 palmitoylates NOD1/2, which is necessary for its membrane localization and induction of NF-κB signaling in response to peptidoglycan (94). Toll-like receptor 4 (TLR4) is a pattern recognition receptor expressed by various innate immune cells, including macrophages, dendritic cells, and neutrophils (95). TLR4 participates in inflammasome activation by triggering downstream myeloid differentiation primary response protein (MYD88) signaling, leading to the release of inflammatory factors TNF-α, IL-1β, and IL-18 (96). The binding of IRAK4 to the MYD88 intermediate domain and subsequent signal activation require MYD88 S-palmitoylation at cysteine 113 (97). The S-palmitoylation of MYD88 is facilitated by de novo fatty acid synthesis and CD36-mediated incorporation of exogenous fatty acids. zDHHC6 is responsible for palmitoylating MYD88, and knockdown of zDHHC6 reduces MYD88 S-palmitoylation and impairs TLR/MYD88 activation upon lipopolysaccharide stimulus in neutrophils (97).

STING plays a crucial role in the innate immune response during infection (98). Research has shown that STING expression is elevated in patients with RCC and controls tumor growth through non-standard innate immune signaling involving the maintenance of mitochondrial ROS and calcium homeostasis (99). STING interacts with mitochondrial voltage-dependent anion channel (VDAC2) via STING-C88/C91 palmitoylation. zDHHC3, zDHHC7, and zDHHC15 palmitoylate STING in the Golgi apparatus of C89/91 (99). Depletion of STING enhances the formation of mitochondria-ER contacts mediated by VDAC2/grp75, leading to increased levels of mitochondrial ROS/calcium and damage to mitochondrial function. Recently, covalently bound small molecule inhibitors that target STING palmitoylation have been developed, further confirming the critical role of STING palmitoylation in its assembly into polymeric complexes in the Golgi apparatus and the activation of downstream STING-triggered inflammatory signals (100). These studies reveal the significance of S-palmitoylation of STING in regulating mitochondrial function and growth in RCC, providing a rationale for targeting STING palmitoylation in the treatment of RCC.

T cells play a crucial role in the adaptive immune response. Recent evaluation of the palmitome in primary T cells and Jurkat T cells revealed a pool of 120 palmitoylated proteins (101). This pool includes both well-known palmitoylated proteins and 92 previously unidentified palmitoylated proteins in vivo. Among the known palmitoylated proteins are ERBB2IP, FASN, CD44, and products of 52 genes associated with breast cancer metastasis to bone, lung, or brain (102). The study also demonstrated that certain PAT enzymes are themselves palmitoylated, suggesting the presence of a feedback mechanism. Furthermore, differential S-palmitoylation of T-cell surface antigens (mono- versus dual-lipidation) was observed, which likely contributes to increased functional flexibility. Previous studies have shown that S-palmitoylation regulates various components of the T-cell receptor (TCR) signaling pathway (59). S-palmitoylation has also been found to impact the T-cell co-receptors CD4 and CD8, as well as associated signaling molecules such as SRC family kinases LCK and FYN (103). Specifically, S-palmitoylation of CD4 on cysteine residues Cys396 and Cys399 near the membrane is associated with LCK, and together they enhance the enrichment and localization of CD4 in lipid rafts (3). Similarly, S-palmitoylation of CD8b is essential for the effective functioning of the CD8 co-receptor, as it increases the association of CD8 with p56lck and enhances p56lck activation in lipid rafts (104). In addition to N-terminal glycine myristoylation, LCK is palmitoylated on Cys3 and Cys5 (105). Both zDHHC2 and zDHHC21 have been identified as enzymes responsible for palmitoylating LCK (89, 106). S-palmitoylation plays a critical role in the plasma membrane targeting of LCK and the phosphorylation of its substrates, such as CD8. Knockdown of zDHHC21 leads to decreased LCK S-palmitoylation and subsequently reduced T-cell activation.

Calcium signaling plays a crucial role in T cell signaling, with calcium increases primarily mediated by 1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase gamma-1 (PLC-γ1). PLC-γ1 activates the production of inositol triphosphate, which then binds to the inositol triphosphate receptor, triggering an increase in calcium levels within the cell membrane (107). zDHHC21 palmitoylates PLC-γ1, and its activity is dynamically regulated by TCR signaling (59). Furthermore, the T cell activation adapter LAT undergoes S-palmitoylation at Cys26 and Cys29. This S-palmitoylation helps LAT efficiently distribute to membrane microdomains and enables its tyrosine phosphorylation. The plasma membrane targeting function of S-palmitoylation is crucial in TCR signaling, and the absence of LAT S-palmitoylation results in T cell inactivity (108).

JAK2/STAT3 is a crucial signaling pathway that plays a significant role in promoting inflammation and tumor progression (109). The activation of JAK2 occurs when extracellular IL-6 binds to its receptor on the cell membrane, subsequently leading to the activation of STAT3 signaling. Once phosphorylated, STAT3 translocates to the nucleus and functions as a transcription factor, facilitating the expression of downstream genes. In this context, zDHHC7 plays a noteworthy role by palmitoylating STAT3 at cysteine 108, which enhances its membrane recruitment and phosphorylation (110). Interestingly, phosphorylated STAT3 is depalmitoylated by APT2, allowing it to translocate to the nucleus. This palmitoylation-depalmitoylation cycle augments STAT3 activation, thereby promoting Th17 cell differentiation (110). The excessive activation of TH17 cells has been linked to various inflammatory diseases, including inflammatory bowel disease (IBD). In a mouse model, the inhibition of APT2 through pharmacological means or the knockout of zDHHC7 has been found to alleviate the symptoms associated with IBD (110). These findings not only propose a potential therapeutic strategy for treating IBD but also provide insights into the role of S-palmitoylation in regulating cell signaling, which could have broader implications for understanding the functional significance of various S-palmitoylation events.

Immune checkpoint inhibitors have been widely utilized in tumor immunotherapy. PD-L1, a transmembrane protein, is highly expressed on various types of cancer cells. By binding to its receptor PD-1 on T cells, PD-L1 significantly inhibits T cell activation and activity, thereby playing a crucial role in enabling tumor cells to evade immune surveillance (111). Antibodies that block PD-L1/PD-1 interactions have revolutionized cancer treatment, showing promising clinical outcomes in melanoma, lung, bladder, colorectal, and renal cell carcinoma. However, the response rate to PD-L1/PD-1 antibody therapy is less satisfactory in other cancers such as prostate, ovarian, and breast cancer. In breast cancer, PD-L1 has been found to be palmitoylated by zDHHC9 at the C272 site, enhancing its stability (33). Mutation of Cys272 to Ala dramatically abolishes mPD-L1 palmitoylation, or inhibiting the expression of zDHHC9 sensitizes breast cancer cells to T-cell killing, thereby repressing tumor growth (33). Another study discovered that S-palmitoylation of PD-L1 at Cys272 inhibits its ubiquitination-mediated lysosomal degradation. zDHHC3 is the primary acyltransferase responsible for PD-L1 S-palmitoylation, and inhibiting PD-L1 S-palmitoylation using 2-BP or silencing DHHC3 activates anti-tumor immunity in CRC (112). The same group further elucidated the regulatory role of S-palmitoylation in PD-1 stability in tumor cells. Mechanistically, zDHHC9 mediates PD-1 palmitoylation at C192, promoting its binding with recycling endosomes and preventing lysosome-dependent degradation (113). They demonstrated that palmitoylation of PD-1, but not PD-L1, promotes mTOR signaling and tumor cell proliferation.

S-palmitoylation of PD-L1 plays a crucial role in regulating its stability. It inhibits the monoubiquitination of PD-L1, thereby preventing its passage through the endosomal sorting complex required for transport (ESCRT). Consequently, the degradation of PD-L1 via the lysosomal pathway is blocked, leading to increased expression of PD-L1 on the cell surface. This increased expression of PD-L1 inhibits the cytotoxicity of T cells. In order to enhance the lethality of cytotoxic T cells against cancer cells, researchers developed a competitive peptide that specifically inhibits PD-L1, surpassing the commonly used universal palmitoylation inhibitor 2-BP in terms of specificity. As a result, the inhibitor reduced the expression of PD-L1 in tumor cells. These findings highlight the significant antitumor effects of targeted S-palmitoylation in vitro and offer promising opportunities for the development of new approaches in cancer immunotherapy.