# SCUBE3 downregulation modulates hepatocellular carcinoma by inhibiting CCNE1 via TGFβ/PI3K/AKT/GSK3β pathway **Authors:** Pan Xu, Aoran Luo, Chuan Xiong, Hong Ren, Liang Yan, Qiang Luo PMC · DOI: 10.1186/s12935-021-02402-z · Cancer Cell International · 2022-01-03 ## TL;DR This study shows that SCUBE3 promotes liver cancer growth by affecting cell cycle proteins and signaling pathways, suggesting it could be a new target for treatment. ## Contribution The novel finding is that SCUBE3 promotes hepatocellular carcinoma by regulating CCNE1 via the TGFβ/PI3K/AKT/GSK3β pathway. ## Key findings - SCUBE3 knockdown inhibits HCC cell proliferation and promotes apoptosis in vitro and in vivo. - SCUBE3 binds to TGFβRII and modulates the PI3K/AKT/GSK3β pathway to regulate CCNE1 expression. - SCUBE3 is upregulated in HCC tissues and may serve as a potential diagnostic and therapeutic target. ## Abstract We aimed to verify the role of signal peptide-CUB-EGF-like domain-containing protein3 (SCUBE3) in the hepatocellular carcinoma (HCC) progression. The role of SCUBE3 in HCC cell proliferation, apoptosis, and cell cycle in vitro were detected using MTT assay, colony formation assay, 5-ethynyl-2´-deoxyuridine assay (EDU), Celigo cell counting assay, Caspase3/7 activity assay, and flow cytometry. The effect of SCUBE3 on HCC cell proliferation in vivo was inspected by a xenograft tumour model in nude mice. The related mechanisms were further studied. The level of SCUBE3 was upregulated in HCC tissues and cell lines. Knockdown of SCUBE3 inhibited proliferation, promoted apoptosis, and induced cell cycle arrest in HCC cell lines in vitro and in vivo. Screening of cell cycle-related proteins revealed that CCNL2, CDK6, CCNE1, and CCND1 exhibited a significantly different expression profile. We found that SCUBE3 may promote the proliferation of HCC cells by regulating CCNE1 expression. The pathway enrichment analysis showed that the TGFβ signalling pathway and the PI3K/AKT signalling pathway were significantly altered. Co-immunoprecipitation results showed that SCUBE3 binds to the TGFβRII receptor. SCUBE3 knockdown inhibited the PI3K/AKT signalling pathway and the phosphorylation of GSK3β to inhibit its kinase activity. SCUBE3 promotes HCC development by regulating CCNE1 via TGFβ/PI3K/AKT/GSK3β pathway. In addition, SCUBE3 may be a new molecular target for the clinical diagnosis and treatment of HCC. The online version contains supplementary material available at 10.1186/s12935-021-02402-z. ## Linked entities - **Genes:** SCUBE3 (signal peptide, CUB domain and EGF like domain containing 3) [NCBI Gene 222663], CCNE1 (cyclin E1) [NCBI Gene 898], CCNL2 (cyclin L2) [NCBI Gene 81669], CDK6 (cyclin dependent kinase 6) [NCBI Gene 1021], CCND1 (cyclin D1) [NCBI Gene 595], TGFBR2 (transforming growth factor beta receptor 2) [NCBI Gene 100033860] - **Proteins:** SCUBE3 (signal peptide, CUB domain and EGF like domain containing 3), CCNE1 (cyclin E1), CCNL2 (cyclin L2), CDK6 (cyclin dependent kinase 6), CCND1 (cyclin D1), TGFBR2 (transforming growth factor beta receptor 2), PIK3CA (phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha), AKT1 (AKT serine/threonine kinase 1), GSK3B (glycogen synthase kinase 3 beta) - **Diseases:** hepatocellular carcinoma (MONDO:0007256), HCC (MONDO:0007256) ## Full-text entities - **Genes:** H3P16 (H3 histone pseudogene 16) [NCBI Gene 644914] {aka H3.6, H3F3AP6, p21}, NPAT (nuclear protein, coactivator of histone transcription) [NCBI Gene 4863] {aka E14, E14/NPAT, p220}, CDK6 (cyclin dependent kinase 6) [NCBI Gene 1021] {aka MCPH12, PLSTIRE}, Ccnd1 (cyclin D1) [NCBI Gene 12443] {aka CycD1, Cyl-1, PRAD1, bcl-1, cD1}, BMP1 (bone morphogenetic protein 1) [NCBI Gene 649] {aka OI13, PCOLC, PCP, TLD}, APC (APC regulator of Wnt signaling pathway) [NCBI Gene 324] {aka BTPS2, DESMD, DP2, DP2.5, DP3, GS}, C1S (complement C1s) [NCBI Gene 716] {aka EDSPD2}, E2F1 (E2F transcription factor 1) [NCBI Gene 1869] {aka E2F-1, RBAP1, RBBP3, RBP3}, AKT1 (AKT serine/threonine kinase 1) [NCBI Gene 207] {aka AKT, PKB, PKB-ALPHA, PRKBA, RAC, RAC-ALPHA}, Nfkb1 (nuclear factor of kappa light polypeptide gene enhancer in B cells 1, p105) [NCBI Gene 18033] {aka NF-KB1, NF-kappaB, NF-kappaB1, p105, p50, p50/p105}, Cdk6 (cyclin dependent kinase 6) [NCBI Gene 12571] {aka Crk2}, SCUBE3 (signal peptide, CUB domain and EGF like domain containing 3) [NCBI Gene 222663] {aka CEGF3, SSFSC2}, GAPDH (glyceraldehyde-3-phosphate dehydrogenase) [NCBI Gene 2597] {aka G3PD, GAPD, HEL-S-162eP}, BMP4 (bone morphogenetic protein 4) [NCBI Gene 652] {aka BMP2B, BMP2B1, MCOPS6, OFC11, ZYME}, CCNE1 (cyclin E1) [NCBI Gene 898] {aka CCNE, pCCNE1}, Tgfb1 (transforming growth factor, beta 1) [NCBI Gene 21803] {aka TGF-beta1, TGFbeta1, Tgfb, Tgfb-1}, Caspase3/7 [NCBI Gene 12367;12369], GRN (granulin precursor) [NCBI Gene 2896] {aka CLN11, FTD2, GEP, GP88, PCDGF, PEPI}, TGFB1 (transforming growth factor beta 1) [NCBI Gene 7040] {aka CAEND1, CED, DPD1, IBDIMDE, LAP, TGF-beta1}, SMAD3 (SMAD family member 3) [NCBI Gene 4088] {aka HSPC193, HsT17436, JV15-2, LDS1C, LDS3, MADH3}, TGFBR2 (transforming growth factor beta receptor 2) [NCBI Gene 7048] {aka AAT3, FAA3, LDS1B, LDS2, LDS2B, MFS2}, BRCA1 (BRCA1 DNA repair associated) [NCBI Gene 672] {aka BRCAI, BRCC1, BROVCA1, FANCS, IRIS, PNCA4}, C1R (complement C1r) [NCBI Gene 715] {aka EDS8, EDSPD1}, DCTN6 (dynactin subunit 6) [NCBI Gene 10671] {aka WS-3, WS3, p27}, CASP3 (caspase 3) [NCBI Gene 836] {aka CPP32, CPP32B, SCA-1}, GSK3B (glycogen synthase kinase 3 beta) [NCBI Gene 2932], CCND1 (cyclin D1) [NCBI Gene 595] {aka BCL1, D11S287E, PRAD1, U21B31}, Ccne1 (cyclin E1) [NCBI Gene 12447] {aka CycE1}, SMAD4 (SMAD family member 4) [NCBI Gene 4089] {aka DPC4, JIP, MADH4, MYHRS}, Scube3 (signal peptide, CUB domain, EGF-like 3) [NCBI Gene 268935] {aka CEGF3, D030038I21Rik}, Gsk3b (glycogen synthase kinase 3 beta) [NCBI Gene 56637] {aka 7330414F15Rik, 8430431H08Rik, GSK-3, GSK-3beta, GSK3}, TP53 (tumor protein p53) [NCBI Gene 7157] {aka BCC7, BMFS5, LFS1, P53, TRP53}, Ccnl2 (cyclin L2) [NCBI Gene 56036] {aka 1700010A01Rik, 1810019L15Rik, 2010319M22Rik, Ania-6b, HCLA-150, Pcee}, BMP2 (bone morphogenetic protein 2) [NCBI Gene 650] {aka BDA2, BMP2A, SSFSC, SSFSC1}, CCNL2 (cyclin L2) [NCBI Gene 81669] {aka ANIA-6B, CCNM, CCNS, HCLA-ISO, HLA-ISO, PCEE}, SMAD2 (SMAD family member 2) [NCBI Gene 4087] {aka CHTD8, JV18, JV18-1, LDS6, MADH2, MADR2}, EGF (epidermal growth factor) [NCBI Gene 1950] {aka HOMG4, URG}, Akt1 (Akt serine/threonine kinase 1) [NCBI Gene 11651] {aka Akt, LTR-akt, PKB, PKB/Akt, PKBalpha, Rac}, CDK2 (cyclin dependent kinase 2) [NCBI Gene 1017] {aka CDKN2, p33(CDK2)}, ANXA5 (annexin A5) [NCBI Gene 308] {aka ANX5, CPB-I, ENX2, HEL-S-7, PP4, RPRGL3} - **Diseases:** ovarian, breast, bladder and colorectal cancer (MESH:D010051), glioma (MESH:D005910), inflammation (MESH:D007249), metastasis (MESH:D009362), alcoholism (MESH:D000437), endocrine system disorder (MESH:D004700), KD (MESH:D009080), Infectious Diseases (MESH:D003141), liver injury (MESH:D017093), gastrointestinal disease (MESH:D005767), lung cancer (MESH:D008175), cirrhosis (MESH:D005355), Viral Hepatitis (MESH:D014777), non-small-cell lung cancer (MESH:D002289), osteosarcoma (MESH:D012516), liver disease (MESH:D008107), Cancer (MESH:D009369), liver fibrosis (MESH:D008103), reproductive system disease (MESH:D060737), cardiac hypertrophy (MESH:D006332), organism damage and abnormality (MESH:D000014), death (MESH:D003643), Hepatocellular carcinoma (MESH:D006528), breast cancer (MESH:D001943), liver tumour (MESH:D008113) - **Chemicals:** polybrene (MESH:D006583), FITC (MESH:D016650), Lipofectamine (MESH:C086724), ethanol (MESH:D000431), CO2 (MESH:D002245), streptomycin (MESH:D013307), EdU (MESH:C022811), agarose (MESH:D012685), MTT (MESH:C070243), 4',6-diamidino-2-phenylindole (MESH:C007293), Apollo (MESH:C062106), penicillin (MESH:D010406), 5-ethynyl-2 -deoxyuridine (MESH:C031086), PBS (MESH:D007854), crystal violet (MESH:D005840), TRIzol (MESH:C411644), isoflurane (MESH:D007530), Celigo (-), IP (MESH:C041508), PI (MESH:D010716), SDS (MESH:D012967), puromycin (MESH:D011691), PVDF (MESH:C024865), Alexa-Fluor 555 (MESH:C000608607), Paraformaldehyde (MESH:C003043) - **Species:** Mus musculus (house mouse, species) [taxon 10090], Escherichia coli (E. coli, species) [taxon 562], Lentivirus (genus) [taxon 11646], Homo sapiens (human, species) [taxon 9606], Culcita novaeguineae (species) [taxon 31170], Rattus norvegicus (brown rat, species) [taxon 10116] - **Cell lines:** BALB/c — Mus musculus (Mouse), Spontaneously immortalized cell line (CVCL_0184), ShSCUBE3 — Mus musculus (Mouse), Hybridoma (CVCL_C6V6), SMMC7721 — Homo sapiens (Human), Human papillomavirus-related endocervical adenocarcinoma, Cancer cell line (CVCL_0534), DH5alpha — Drosophila hydei (Fruit fly), Spontaneously immortalized cell line (CVCL_Z531), HepG2 — Homo sapiens (Human), Hepatoblastoma, Cancer cell line (CVCL_0027), Bel7402 — Homo sapiens (Human), Human papillomavirus-related endocervical adenocarcinoma, Cancer cell line (CVCL_5492), shCCNE1 — Mus musculus (Mouse), Hybridoma (CVCL_C7RB), S2 — Drosophila melanogaster (Fruit fly), Spontaneously immortalized cell line (CVCL_Z232), U251 — Homo sapiens (Human), Astrocytoma, Cancer cell line (CVCL_0021), shSCUBE3 — Homo sapiens (Human), Transformed cell line (CVCL_B3K0), CB101 — Mus musculus (Mouse), Hybridoma (CVCL_J815), Ubi- — Homo sapiens (Human), Induced pluripotent stem cell (CVCL_YM94), HR — Homo sapiens (Human), Ovarian serous cystadenocarcinoma, Cancer cell line (CVCL_ZE74), Bel7404 — Homo sapiens (Human), Human papillomavirus-related endocervical adenocarcinoma, Cancer cell line (CVCL_6568) ## Full text _Full body text omitted from this summary view._ Fetch the complete paper as Markdown: https://tomesphere.com/paper/PMC8725472/full.md ## Figures 9 figures with captions in the complete paper: https://tomesphere.com/paper/PMC8725472/full.md ## References 45 references — full list in the complete paper: https://tomesphere.com/paper/PMC8725472/full.md --- Source: https://tomesphere.com/paper/PMC8725472