Transmembrane 4 L Six Family Member 5 Research Articles

Glucose-mediated mitochondrial reprogramming by cholesterol export at TM4SF5-enriched mitochondria-lysosome contact sites.

Transmembrane 4 L six family member 5 (TM4SF5) translocates subcellularly and functions metabolically, although it is unclear how intracellular TM4SF5 translocation is linked to metabolic contexts. It is thus of interests to understand how the traffic dynamics of TM4SF5 to subcellular endosomal membranes are correlated to regulatory roles of metabolisms. Here, we explored the metabolic significance of TM4SF5 localization at mitochondria-lysosome contact sites (MLCSs), using in vitro cells and in vivo animal systems, via approaches by immunofluorescence, proximity labelling based proteomics analysis, organelle reconstitution etc. RESULTS: Upon extracellular glucose repletion following depletion, TM4SF5 became enriched at MLCSs via an interaction between mitochondrial FK506-binding protein 8 (FKBP8) and lysosomal TM4SF5. Proximity labeling showed molecular clustering of phospho-dynamic-related protein I (DRP1) and certain mitophagy receptors at TM4SF5-enriched MLCSs, leading to mitochondrial fission and autophagy. TM4SF5 bound NPC intracellular cholesterol transporter 1 (NPC1) and free cholesterol, and mediated export of lysosomal cholesterol to mitochondria, leading to impaired oxidative phosphorylation but intact tricarboxylic acid (TCA) cycle and β-oxidation. In mouse models, hepatocyte Tm4sf5 promoted mitophagy and cholesterol transport to mitochondria, both with positive relations to liver malignancy. Our findings suggested that TM4SF5-enriched MLCSs regulate glucose catabolism by facilitating cholesterol export for mitochondrial reprogramming, presumably while hepatocellular carcinogenesis, recapitulating aspects for hepatocellular carcinoma metabolism with mitochondrial reprogramming to support biomolecule synthesis in addition to glycolytic energetics.

TM4SF5-mediated abnormal food-intake behavior and apelin expression facilitate non-alcoholic fatty liver disease features

Transmembrane 4L six family member 5 (TM4SF5) engages in non-alcoholic steatohepatitis (NASH), although its mechanistic roles are unclear. Genetically engineered Tm4sf5 mice fed ad libitum normal chow or high-fat diet for either an entire day or a daytime-feeding (DF) pattern were analyzed for metabolic parameters. Compared to wild-type and Tm4sf5-/- knockout mice, hepatocyte-specific TM4SF5-overexpressing Alb-TGTm4sf5-Flag (TG) mice showed abnormal food-intake behavior during the mouse-inactive daytime, increased apelin expression, increased food intake, and higher levels of NASH features. DF or exogenous apelin injection of TG mice caused severe hepatic pathology. TM4SF5-mediated abnormal food intake was correlated with peroxisomal β-oxidation, mTOR activation, and autophagy inhibition, with triggering NASH phenotypes. Non-alcoholic fatty liver disease (NAFLD) patients' samples revealed a correlation between serum apelin and NAFLD activity score. Altogether, these observations suggest that hepatic TM4SF5 may cause abnormal food-intake behaviors to trigger steatohepatitic features via the regulation of peroxisomal β-oxidation, mTOR, and autophagy.

Systemic TM4SF5 overexpression in ApcMin/+ mice promotes hepatic portal hypertension associated with fibrosis

Mutation of the gene for adenomatous polyposis coli (APC), as seen in ApcMin/+ mice, leads to intestinal adenomas and carcinomas via stabilization of β-catenin. Transmembrane 4 L six family member 5 (TM4SF5) is involved in the development of non-alcoholic fatty liver disease, fibrosis, and cancer. However, the functional linkage between TM4SF5 and APC or β-catenin has not been investigated for pathological outcomes. After interbreeding ApcMin/+ with TM4SF5-overexpressing transgenic (TgTM4SF5) mice, we explored pathological outcomes in the intestines and livers of the offspring. The intestines of 26-week-old dual-transgenic mice (ApcMin/+:TgTM4SF5) had intramucosal adenocarcinomas beyond the single-crypt adenomas in ApcMin/+ mice. Additional TM4SF5 overexpression increased the stabilization of β-catenin via reduced glycogen synthase kinase 3β (GSK3β) phosphorylation on Ser9. Additionally, the livers of the dualtransgenic mice showed distinct sinusoidal dilatation and features of hepatic portal hypertension associated with fibrosis, more than did the relatively normal livers in ApcMin/+ mice. Interestingly, TM4SF5 overexpression in the liver was positively linked to increased GSK3β phosphorylation (opposite to that seen in the colon), β-catenin level, and extracellular matrix (ECM) protein expression, indicating fibrotic phenotypes. Consistent with these results, 78-week-old TgTM4SF5 mice similarly had sinusoidal dilatation, immune cell infiltration, and fibrosis. Altogether, systemic overexpression of TM4SF5 aggravates pathological abnormalities in both the colon and the liver. [BMB Reports 2022; 55(12): 609-614].

Liver-originated small extracellular vesicles with TM4SF5 target brown adipose tissue for homeostatic glucose clearance.

Transmembrane 4 L six family member 5 (TM4SF5) is involved in chronic liver disease, although its role in glucose homeostasis remains unknown. TM4SF5 deficiency caused age‐dependent glucose (in)tolerance with no link to insulin sensitivity. Further, hepatic TM4SF5 binding to GLUT1 promoted glucose uptake and glycolysis. Excessive glucose repletion caused hepatocytes to secrete small extracellular vesicles (sEVs) loaded with TM4SF5 (hep‐sEVTm4sf5), suggesting a role for sEVTm4sf5 in glucose metabolism and homeostasis. Hep‐sEVTm4sf5 were smaller than sEVControl and recruit proteins for efficient organ tropism. Liver‐derived sEVs, via a liver‐closed vein circuit (LCVC) using hepatic TM4SF5‐overexpressing (Alb‐Tm4sf5 TG) mice (liv‐sEVTm4sf5), improved glucose tolerance in Tm4sf5 −/− KO mice and targeted brown adipose tissues (BATs), possibly allowing the clearance of blood glucose as heat independent of UCP1. Taken together, hep‐sEVTm4sf5 might clear high extracellular glucose levels more efficiently by targeting BAT compared with hep‐sEVControl, suggesting an insulin‐like role for sEV™4SF5 in affecting age‐related metabolic status and thus body weight (BW).

TM4SF5-Mediated Regulation of Hepatocyte Transporters during Metabolic Liver Diseases.

Nonalcoholic fatty liver disease (NAFLD) is found in up to 30% of the world’s population and can lead to hepatocellular carcinoma (HCC), which has a poor 5-year relative survival rate of less than 40%. Clinical therapeutic strategies are not very successful. The co-occurrence of metabolic disorders and inflammatory environments during the development of steatohepatitis thus needs to be more specifically diagnosed and treated to prevent fatal HCC development. To improve diagnostic and therapeutic strategies, the identification of molecules and/or pathways responsible for the initiation and progression of chronic liver disease has been explored in many studies, but further study is still required. Transmembrane 4 L six family member 5 (TM4SF5) has been observed to play roles in the regulation of metabolic functions and activities in hepatocytes using in vitro cell and in vivo animal models without or with TM4SF5 expression in addition to clinical liver tissue samples. TM4SF5 is present on the membranes of different organelles or vesicles and cooperates with transporters for fatty acids, amino acids, and monocarbohydrates, thus regulating nutrient uptake into hepatocytes and metabolism and leading to phenotypes of chronic liver diseases. In addition, TM4SF5 can remodel the immune environment by interacting with immune cells during TM4SF5-mediated chronic liver diseases. Because TM4SF5 may act as an NAFLD biomarker, this review summarizes crosstalk between TM4SF5 and nutrient transporters in hepatocytes, which is related to chronic liver diseases.

Therapeutic effects of TM4SF5-targeting chimeric and humanized monoclonal antibodies in hepatocellular and colon cancer models

The transmembrane 4 L six family member 5 (TM4SF5) is aberrantly expressed in hepatocellular and colorectal cancers, and has been implicated in tumor progression, suggesting that it could serve as a novel therapeutic target. Previously, we screened a murine antibody phage-display library to generate a novel monoclonal antibody, Ab27, that is specific to the extracellular loop 2 of TM4SF5. In this study, we evaluated the effects of chimeric Ab27 using cancer cells expressing endogenous TM4SF5 or stably overexpressing TM4SF5 in vivo and in vitro. Monotherapy with Ab27 significantly decreased tumor growth in liver and colon cancer xenograft models, including a sorafenib-resistant model, and decreased the phosphorylation of focal adhesion kinase (FAK), p27Kip1, and signal transducer and activator of transcription 3 (STAT3). No general Ab27 toxicity was observed in vivo. Combination treatment with Ab27 and sorafenib or doxorubicin exerted higher antitumor activity than monotherapy. In addition, we humanized the Ab27 sequence by the complementarity-determining region (CDR) grafting method. The humanized antibody Ab27-hz9 had reduced immunogenicity but exhibited target recognition and antitumor activity comparable with those of Ab27. Both Ab27 and Ab27-hz9 efficiently targeted tumor cells expressing TM4SF5 in vivo. These observations strongly support the further development of Ab27-hz9 as a novel therapeutic agent against liver and colorectal cancers.

TM4SF5-mediated liver malignancy involves NK cell exhaustion-like phenotypes

Aberrant extracellular matrix and immune cell alterations within the tumor microenvironment promote the pathological progression of liver carcinogenesis. Although transmembrane 4 L six family member 5 (TM4SF5) is involved in liver fibrosis and cancer, its mechanism avoiding immune surveillance during carcinogenesis remains unknown. We investigated how TM4SF5-mediated signaling caused immune evasion using in vitro primary cells and in vivo liver tissues from genetic or chemically induced mouse models. TM4SF5-transgenic and diethylnitrosamine (DEN)-induced liver cancer mouse models exhibited fibrotic and cancerous livers, respectively, with enhanced TM4SF5, pY705STAT3, collagen I, and laminin γ2 levels. These TM4SF5-mediated effects were abolished by TM4SF5 inhibitor, 4′-(p-toluenesulfonylamido)-4-hydroxychalcone (TSAHC). TM4SF5-dependent tumorigenesis involved natural killer (NK) cell exhaustion-like phenotypes including the reduction of NK cell number or function, which were blocked with TSAHC treatment. TM4SF5 expression in cancer cells downregulated stimulatory ligands and receptors for NK cell cytotoxicity, including SLAMF6, SLAMF7, MICA/B, and others. TM4SF5 suppression or inhibition reduced STAT3 signaling activity and recovered the receptor levels and NK cell surveillance, leading to reduced fibrotic and cancerous phenotypes, and longer survival. Altogether, these findings suggest that TM4SF5-mediated STAT3 activity for extracellular matrix modulation is involved in the progression of liver disease to HCC and that TM4SF5 appears to suppress NK cells during liver carcinogenesis.

Tetraspanin TM4SF5 in hepatocytes negatively modulates SLC27A transporters during acute fatty acid supply

Transmembrane 4 L six family member 5 (TM4SF5) is involved in nonalcoholic steatosis and further aggravation of liver disease. However, its mechanism for regulating FA accumulation is unknown. We investigated how TM4SF5 in hepatocytes affected FA accumulation during acute FA supply. TM4SF5-expressing hepatocytes and mouse livers accumulated less FAs, compared with those of TM4SF5 deficiency or inactivation. Binding of TM4SF5 to SLC27A2 increased gradually upon acute FA treatment, whereas TM4SF5 constitutively bound SLC27A5. Suppression of either SLC27A2 or SLC27A5 in hepatocytes expressing TM4SF5 differentially modulated initial and maximal FA uptake levels for a fast turnover of fatty acid. Altogether, TM4SF5 negatively modulates FA accumulation into hepatocytes via association with the transporters for an energy homeostasis, when FA are supplied acutely.

TM4SF5 Knockout Protects Mice From Diet-Induced Obesity Partly by Regulating Autophagy in Adipose Tissue.

Transmembrane 4 L six family member 5 (TM4SF5) functions as a sensor for lysosomal arginine levels and activates the mammalian target of rapamycin complex 1 (mTORC1). While the mTORC1 signaling pathway plays a key role in adipose tissue metabolism, the regulatory function of TM4SF5 in adipocytes remains unclear. In this study we aimed to establish a TM4SF5 knockout (KO) mouse model and investigated the effects of TM4SF5 KO on mTORC1 signaling–mediated autophagy and mitochondrial metabolism in adipose tissue. TM4SF5 expression was higher in inguinal white adipose tissue (iWAT) than in brown adipose tissue and significantly upregulated by a high-fat diet (HFD). TM4SF5 KO reduced mTORC1 activation and enhanced autophagy and lipolysis in adipocytes. RNA sequencing analysis of TM4SF5 KO mouse iWAT showed that the expression of genes involved in peroxisome proliferator–activated receptor α signaling pathways and mitochondrial oxidative metabolism was upregulated. Consequently, TM4SF5 KO reduced adiposity and increased energy expenditure and mitochondrial oxidative metabolism. TM4SF5 KO prevented HFD-induced glucose intolerance and inflammation in adipose tissue. Collectively, the results of our study demonstrate that TM4SF5 regulates autophagy and lipid catabolism in adipose tissue and suggest that TM4SF5 could be therapeutically targeted for the treatment of obesity-related metabolic diseases.

SLAC2B-dependent microtubule acetylation regulates extracellular matrix-mediated intracellular TM4SF5 traffic to the plasma membranes.

Transmembrane 4 L six family member 5 (TM4SF5) translocates intracellularly and promotes cell migration, but how subcellular TM4SF5 traffic is regulated to guide cellular migration is unknown. We investigated the influences of the extracellular environment and intracellular signaling on the TM4SF5 traffic with regard to migration directionality. Cell adhesion to fibronectin (FN) but not poly-l-lysine enhanced the traffic velocity and straightness of the TM4SF5WT (but not palmitoylation-deficient mutant ) toward the leading edges, depending on tubulin acetylation. Acetylated-microtubules in SLAC2B-positive cells reached mostly the juxtanuclear regions, but reached-out toward the leading edges upon SLAC2B suppression. TM4SF5 expression caused SLAC2B not to be localized at the leading edges. TM4SF5 colocalization with HDAC6 depended on paxillin expression. The trimeric complex consisting of TM4SF5, HDAC6, and SLAC2B might, thus, be enriched at the perinuclear cytosols toward the leading edges. More TM4SF5WT translocation to the leading edges was possible when acetylated-microtubules reached the frontal edges following HDAC6 inhibition by paxillin presumably at new cell-FN adhesions, leading to persistent cell migration. Collectively, this study revealed that cell-FN adhesion and microtubule acetylation could control intracellular traffic of TM4SF5 vesicles to the leading edges via coordinated actions of paxillin, SLAC2B, and HDAC6, leading to TM4SF5-dependent cell migration.

TM4SF5-mediated CD44v8-10 splicing variant promotes survival of type II alveolar epithelial cells during idiopathic pulmonary fibrosis

Reactive oxygen species (ROS) regulate cell fate, although signaling molecules that regulate ROS hormesis remain unclear. Here we show that transmembrane 4 L six family member 5 (TM4SF5) in lung epithelial cells induced the alternatively spliced CD44v8-10 variant via an inverse ZEB2/epithelial splicing regulatory proteins (ESRPs) linkage. TM4SF5 formed complexes with the cystine/glutamate antiporter system via TM4SF5- and CD44v8-10-dependent CD98hc plasma-membrane enrichment. Dynamic TM4SF5 binding to CD98hc required CD44v8-10 under ROS-generating inflammatory conditions. TM4SF5 and CD44v8-10 upregulated cystine/glutamate antiporter activity and intracellular glutathione levels, leading to ROS modulation for cell survival. Tm4sf5-null mice exhibited attenuated bleomycin-induced pulmonary fibrosis with lower CD44v8-10 and ESRPs levels than wild-type mice. Primary mouse alveolar epithelial cells (AECs) revealed type II AECs (AECII), but not type I, to adapt the TM4SF5-mediated characteristics, suggesting TM4SF5-mediated AECII survival following AECI injury during idiopathic pulmonary fibrosis (IPF). Thus, the TM4SF5-mediated CD44v8-10 splice variant could be targeted against IPF.

Transmembrane 4 L Six Family Member 5 Senses Arginine for mTORC1 Signaling

The mechanistic target of rapamycin complex (mTORC1) is a signaling hub on the lysosome surface, responding to lysosomal amino acids. Although arginine is metabolically important, the physiological arginine sensor that activates mTOR remains unclear. Here, we show that transmembrane 4L six family member 5 (TM4SF5) translocates from plasma membrane to lysosome upon arginine sufficiency and senses arginine, culminating in mTORC1/S6K1 activation. TM4SF5 bound active mTOR upon arginine sufficiency and constitutively bound amino acid transporter SLC38A9. TM4SF5 binding to the cytosolic arginine sensor Castor1 decreased upon arginine sufficiency, thus allowing TM4SF5-mediated sensing of metabolic amino acids. TM4SF5 directly bound free L-arginine via its extracellular loop possibly for the efflux, being supported by mutant study and homology and molecular docking modeling. Therefore, we propose that lysosomal TM4SF5 senses and enables arginine efflux for mTORC1/S6K1 activation, and arginine-auxotroph in hepatocellular carcinoma may be targeted by blocking the arginine sensing using anti-TM4SF5 reagents.

Clinicopathological significance of TM4SF5 expression in human hepatocellular carcinoma tissues.

Hepatocellular carcinoma (HCC) is a major cause of cancer-related mortality. Recent studies have indicated that transmembrane 4 L six family member 5 (TM4SF5) serves a vital role in tumor progression and metastasis in human cell lines and mouse models. However, little is known about the association between TM4SF5 expression and clinicopathological factors. The aim of the current study was to investigate this association and evaluate overall survival (OS) of patients. Immunohistochemistry and clinical record analysis revealed that TM4SF5 expression was significantly downregulated in HCC tissues. In addition, TM4SF5 expression was significantly associated with tumor size, vascular invasion, tumor differentiation, and tumor-node-metastasis stage. The survival analysis also demonstrated that low TM4SF5 expression resulted in shorter OS. In conclusion, the association between TM4SF5 expression and clinicopathologic factors was established, and prognostic significance of TM4SF5 as a potential biomarker was evaluated using human HCC formalin-fixed paraffin-embedded tissue samples. The results of the present study demonstrated that low TM4SF5 expression was associated with tumor malignant progression and may be a good prognostic biomarker for OS in HCC.

CD133-induced TM4SF5 expression promotes sphere growth via recruitment and blocking of protein tyrosine phosphatase receptor type F (PTPRF)

CD133 is a surface marker of liver cancer stem cells. Transmembrane 4 L six family member 5 (TM4SF5) promotes sphere growth and circulation. However, it is unknown how CD133 and TM4SF5 cross-talk with each other for cancer stem cell properties. Here, we investigated the significance of inter-relationships between CD133, TM4SF5, CD44, and protein tyrosine phosphatase receptor type F (PTPRF) in a three-dimensional (3D) sphere growth system. We found that CD133 upregulated TM4SF5 and CD44, whereas TM4SF5 and CD44 did not affect CD133 expression. Signaling activity following CD133 phosphorylation caused TM4SF5 expression and sphere growth. TM4SF5 bound to CD133 and promoted c-Src activity for CD133 phosphorylation as a positive feedback loop, leading to CD133-mediated sphere growth that was inhibited by TM4SF5 inhibition or suppression. TM4SF5 also bound PTPRF and promoted paxillin phosphorylation. Decreased sphere growth upon CD133 suppression was recovered by TM4SF5 expression and partially by PTPRF suppression. TM4SF5 inhibition enhanced PTPRF levels and abolished PTPRF suppression-mediated sphere growth. Altogether, CD133-induced TM4SF5 expression and function were important for liver cancer sphere growth and may be a promising target to block metastasis.

TM4SF5 promotes metastatic behavior of cells in 3D extracellular matrix gels by reducing dependency on environmental cues.

Transmembrane 4 L six family member 5 (TM4SF5) is highly expressed in hepatocellular carcinoma tissues and enhances migration in two-dimensional environments. Here, we investigated how TM4SF5 is involved in diverse pro-metastatic phenotypes in in vivo-like three-dimensional (3D) extracellular matrix gels. TM4SF5-positive cells aggressively formed invasive foci in 3D Matrigel, depending on TM4SF5-mediated signaling activity, cytoskeletal organization, and matrix metallopeptidase (MMP) 2-mediated extracellular remodeling, whereas TM4SF5-null cells did not. The TM4SF5-null cells did, however, form invasive foci in 3D Matrigel following inhibition of Rho-associated protein kinase or addition of collagen I, suggesting that collagen I compensated for TM4SF5 expression. Similarly, TM4SF5-positive cells expressing vascular endothelial-cadherin formed network-like vasculogenic mimicry in 3D Matrigel and collagen I mixture gels, whereas TM4SF5-negative cells in the mixture gels displayed the network structures only upon further treatment with epidermal growth factor. The foci formation also required MMP2-mediated remodeling of the extracellular matrix. Co-cultures exhibited TM4SF5-positive or cancer-associated fibroblasts at the outward edges of TM4SF5-null cell clusters. Compared with TM4SF5-null cells, TM4SF5-positive cells in 3D collagen gels showed a more invasive outgrowth with dramatic invadopodia. These observations suggest that TM4SF5 plays roles in the promotion of diverse metastatic properties with fewer environmental requirements than TM4SF5-negative cells.

TM4SF5-Mediated Roles in the Development of Fibrotic Phenotypes.

Transmembrane 4 L six family member 5 (TM4SF5) can form tetraspanin-enriched microdomains (TERMs) on the cell's surface. TERMs contain protein-protein complexes comprised of tetraspanins, growth factor receptors, and integrins. These complexes regulate communication between extracellular and intracellular spaces to control diverse cellular functions. TM4SF5 influences the epithelial-mesenchymal transition (EMT), aberrant multilayer cellular growth, drug resistance, enhanced migration and invasion, circulation through the bloodstream, tumor-initiation property, metastasis, and muscle development in zebrafish. Here, current data on TM4SF5's roles in the development of fibrotic phenotypes are reviewed. TM4SF5 is induced by transforming growth factor β1 (TGFβ1) signaling via a collaboration with epidermal growth factor receptor (EGFR) activation. TM4SF5, by itself or in concert with other receptors, transduces signals intracellularly. In hepatocytes, TM4SF5 expression regulates cell cycle progression, migration, and expression of extracellular matrix components. In CCl4-treated mice, TM4SF5, α-smooth muscle actin (α-SMA), and collagen I expression are observed together along the fibrotic septa regions of the liver. These fibrotic phenotypes are diminished by anti-TM4SF5 reagents, such as a specific small compound [TSAHC, 4′-(p-toluenesulfonylamido)-4-hydroxychalcone] or a chimeric antibody. This review discusses the antifibrotic strategies that target TM4SF5 and its associated protein networks that regulate the intracellular signaling necessary for fibrotic functions of hepatocytes.

Interaction of tetraspan(in) TM4SF5 with CD44 promotes self‐renewal and circulating capacities of hepatocarcinoma cells

Tumor metastasis involves circulating and tumor-initiating capacities of metastatic cancer cells. Epithelial-mesenchymal transition (EMT) is related to self-renewal capacity and circulating tumor cell (CTC) characteristics for tumor metastasis. Although tumor metastasis is a life-threatening, complicated process that occurs through circulation of tumor cells, mechanistic aspects of self-renewal and circulating capacities have been largely unknown. Hepatic transmembrane 4 L six family member 5 (TM4SF5) promotes EMT for malignant growth and migration, so it was rationalized that TM4SF5, as a hepatocellular carcinoma (HCC) biomarker, might be important for metastatic potential. Here, self-renewal capacity by TM4SF5 was mechanistically explored using hepatocarcinoma cells with or without TM4SF5 expression, and we explored whether they became CTCs using mouse liver-orthotopic model systems. We found that TM4SF5-dependent sphere growth correlated with CD24(-) , aldehyde dehydrogenase (ALDH) activity, as well as a physical association between CD44 and TM4SF5. Interaction between TM4SF5 and CD44 was through their extracellular domains with N-glycosylation modifications. TM4SF5/CD44 interaction activated proto-oncogene tyrosine-protein kinase Src (c-Src)/signal transducer and activator of transcription 3 (STAT3)/Twist-related protein 1 (Twist1)/B-cell-specific Moloney murine leukemia virus integration site 1 (Bmi1) signaling for spheroid formation, whereas disturbing the interaction, expression, or activity of any component in this signaling pathway inhibited spheroid formation. In serial xenografts using 200∼5,000 cells per injection, TM4SF5-positive tumors exhibited subpopulations with locally increased CD44 expressions, supporting for tumor cell differentiation. TM4SF5-positive, but not TM4SF5- or CD44-knocked-down, cells were identified circulating in blood 4-6 weeks after orthotopic liver injection using in vivo laser scanning endomicroscopy. Anti-TM4SF5 reagent blocked their metastasis to distal intestinal organs. TM4SF5 promotes self-renewal and CTC properties supported by TM4SF5(+) /CD44(+(TM4SF5-bound)) /ALDH(+) /CD24(-) markers during HCC metastasis.

Transmembrane 4 L Six Family Member 5 (TM4SF5)-Mediated Epithelial-Mesenchymal Transition in Liver Diseases.

The membrane protein TM4SF5, a member of the transmembrane 4L six family, forms a tetraspanin-enriched microdomain (TEM) on the cell surface, where many different membrane proteins and receptors form a massive protein-protein complex to regulate cellular functions including transdifferentiation, migration, and invasion. We recently reported that TM4SF5 causes epithelial-mesenchymal transition (EMT), eventually contributing to aberrant multilayer cellular growth, drug resistance, enhanced migration, invasion, its circulation in the blood, tumor initiation for successful metastasis, and muscle development in zebrafish. In this review, I summarize the information on the role of TM4SF5 in EMT-related functions at TM4SF5-enriched microdomain (T5EM) on cell surface, where proteins such as TM4SF5, CD151, CD44, integrins, and epidermal growth factor receptor (EGFR) can form numerous protein complexes. TM4SF5-mediated EMT contributes to diverse cellular functions, leading to fibrotic phenotypes and initiating and maintaining tumors in primary and/or metastatic regions, in addition to its role in muscle development in zebrafish. Anti-TM4SF5 strategies for addressing the protein networks can lead to regulation of the fibrotic, tumorigenic, and tumor-maintaining functions of TM4SF5-positive hepatic cells. This review is for us to (re)consider the antifibrotic or antitumorigenic (i.e., anti-EMT-related diseases) strategies of dealing with protein networks that would be involved in cross-talks to regulate various cellular functions during TM4SF5-dependent progression from fibrotic to cancerous hepatic cells.

TM4SF5-mediated protein-protein networks and tumorigenic roles.

Transmembrane 4 L six family member 5 (TM4SF5), as a membrane glycoprotein with 4 transmembrane domains, is similar to the tetraspanins in terms of membrane topology and plays important roles in tumorigenesis and tumor metastasis. Especially, TM4SF5 appears to form a massive protein-protein complex consisting of diverse membrane proteins and/or receptors in addition to cytosolic signaling molecules to regulate their signaling activities during the pathological processes. TM4SF5 is shown to interact with integrins α2, α5, and β1, EGFR, IL6R, CD151, focal adhesion kinase (FAK), and c-Src. This review focuses on the significance of the interactions with regards to TM4SF5-positive tumorigenesis and metastasis. [BMB Reports 2014; 47(9): 483-487]

TM4SF5 suppression disturbs integrin α5-related signalling and muscle development in zebrafish.

TM4SF5 (transmembrane 4L six family member 5) is involved in EMT (epithelial-mesenchymal transition) for liver fibrosis and cancer metastasis; however, the function(s) of TM4SF5 during embryogenesis remains unknown. In the present study the effects of TM4SF5 on embryogenesis of zebrafish were investigated. tm4sf5 mRNA was expressed in the posterior somites during somitogenesis and in whole myotome 1 dpf (day post-fertilization). tm4sf5 suppression impaired development of the trunk with aberrant morphology of muscle fibres and altered expression of integrin α5. The arrangement and adhesion of muscle cells were abnormally disorganized in tm4sf5 morphants with reduced muscle fibre masses, where integrin α5-related signalling molecules, including fibronectin, FAK (focal adhesion kinase), vinculin and actin were aberrantly localized, compared with those in control fish. Aberrant muscle developments in tm4sf5 morphants were recovered by additional tm4sf5 or integrin α5 mRNA injection. Such a role for TM4SF5 was observed in the differentiation of C2C12 mouse myoblast cells to multinuclear muscle cells. Taken together, the results show that TM4SF5 controls muscle differentiation via co-operation with integrin α5-related signalling.