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Cycloheximide (chx)

Manufactured by Selleck Chemicals
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Cycloheximide is a chemical compound commonly used as a laboratory reagent. It is a protein synthesis inhibitor, specifically targeting the 60S ribosomal subunit in eukaryotic cells. Cycloheximide is widely utilized in various research applications where the inhibition of protein synthesis is required.

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Lab products found in correlation

Mg132, by Selleck Chemicals (72 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (17 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (10 mentions) Lipofectamine 2000, by Thermo Fisher Scientific (10 mentions) Bortezomib, by Selleck Chemicals (8 mentions) Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (8 mentions) Penicillin, by Thermo Fisher Scientific (7 mentions) Bafilomycin a1, by Selleck Chemicals (6 mentions) Dimethyl sulfoxide (dmso), by Merck Group (6 mentions) Rpmi 1640, by Thermo Fisher Scientific (6 mentions) Erastin, by Selleck Chemicals (5 mentions) Z vad fmk, by Selleck Chemicals (5 mentions) Streptomycin, by Thermo Fisher Scientific (5 mentions) Mg132, by Merck Group (5 mentions) Mg132, by MedChemExpress (4 mentions) Palbociclib, by Selleck Chemicals (4 mentions) β actin, by Proteintech (4 mentions) Gefitinib, by Selleck Chemicals (4 mentions) Rapamycin, by MedChemExpress (3 mentions) Rpmi 1640 medium, by Thermo Fisher Scientific (3 mentions)

163 protocols using cycloheximide (chx)

1

Cycloheximide-Everolimus Protein Kinetics

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Jurkat cells were treated with either 10 μM cycloheximide (Sigma-Aldrich) alone, 10 μM cycloheximide + 15 μM everolimus or with 10 μM cycloheximide + 15 μM everolimus + 1 μM Vps34-IN1 (Selleck Chemicals). Cells were harvested in cell disruption buffer at 0, 1, 4, 8 and 12 h after indicated treatments. Lysates were subjected to SDS-PAGE and immunoblotting to detect G6PD and GAPDH as described above. The G6PD signals measured at the 4 timepoints were scaled against the value at 0 h; resulting values represented the fraction of protein remaining. Protein half-life was calculated through best fitting equations to the data.
Silic-Benussi M., Sharova E., Ciccarese F., Cavallari I., Raimondi V., Urso L., Corradin A., Kotler H., Scattolin G., Buldini B., Francescato S., Basso G., Minuzzo S.A., Indraccolo S., D'Agostino D.M, & Ciminale V. (2022). mTOR inhibition downregulates glucose-6-phosphate dehydrogenase and induces ROS-dependent death in T-cell acute lymphoblastic leukemia cells. Redox Biology, 51, 102268.
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2

Regulation of PTEN Protein Stability

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The human GCs line KGN was cultured in Dulbecco’s modified Eagle’s medium/nutrient mixture F-12 (DMEM/F12) (Gibco, Grand Island, NE, United States) containing 10% charcoal-stripped fetal bovine serum (Gibco) and 1% antibiotics (mixture of penicillin, streptomycin, and neomycin; Gibco) in a 37°C and 5% CO2 incubator (Thermo Fisher Scientific, Waltham, MA, United States). Cells were passaged every 2 days. For MG132 (100 μM, Selleck Chemicals, Houston, TX, United States) treatment, cells were harvested after 6 h of treatment. In addition, cells were treated with 100 μM cycloheximide (CHX, Selleck Chemicals, Houston, TX, United States) and collected every 12 h to assess PTEN protein stability. For inhibition of PI3K/AKT signaling, 30 μm of LY294002 (Selleck Chemicals), a specific inhibitor that blocks the PI3K/AKT pathway, was added to the cells for 12 h. Since MG132, CHX, and LY294002 were all dissolved in DMSO, an equal volume of DMSO was added to the control cells.
Gao Y., Chen J., Ji R., Ding J., Zhang Y, & Yang J. (2021). USP25 Regulates the Proliferation and Apoptosis of Ovarian Granulosa Cells in Polycystic Ovary Syndrome by Modulating the PI3K/AKT Pathway via Deubiquitinating PTEN. Frontiers in Cell and Developmental Biology, 9, 779718.
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3

Protein Stability Analysis by CHX-Chase

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CHX (14 μg/mL, Selleck Chemicals) was used in this study to perform CHX-chase test. CHX (14 μg/mL) was mixed with cells for 0, 2, 4, 8, respectively. Then, the protein was extracted, and protein expression levels were measured through western blotting.
Ye D., Wang S., Wang X., Lin Y., Huang Y, & Chi P. (2022). Overexpression of OTU domain-containing ubiquitin aldehyde-binding protein 1 exacerbates colorectal cancer malignancy by inhibiting protein degradation of β-Catenin via Ubiquitin-proteasome pathway. Bioengineered, 13(4), 9106-9116.
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4

Measuring CX3CL1 Protein Half-Life

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The half‐life of CX3CL1 protein in cells was determined using a CHX chase assay. 4T1 cells were treated with 10 μmol/L CHX (Selleck Chemicals) and lysed at the indicated times after the addition of CHX. CX3CL1 levels were measured using Western blotting.
Zhang W., Zhang Q., Yang N., Shi Q., Su H., Lin T., He Z., Wang W., Guo H, & Shen P. (2022). Crosstalk between IL‐15Rα+ tumor‐associated macrophages and breast cancer cells reduces CD8+ T cell recruitment. Cancer Communications, 42(6), 536-557.
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5

CHX-Chase Assay for P-STAT3 Protein

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CHX-chase assay was performed using CHX (Selleck Chemicals), an inhibitor of protein synthesis. The cells in each group were mixed with 12.5 μg/mL of CHX and the expression of P-STAT3 protein was determined by western blot analysis at 0, 3, 6, 12, and 24 h.
Zhang J, & Gao Y. (2019). Long non-coding RNA MEG3 inhibits cervical cancer cell growth by promoting degradation of P-STAT3 protein via ubiquitination. Cancer Cell International, 19, 175.
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6

Culturing and treating HCC cell lines

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The HCC cell lines were kept in the Institute of Liver and Gastrointestinal Diseases (Tongji Hospital, Huazhong University of Science and Technology). Chang liver, Huh7, MHCC97H, HCCLM3 and SK‐hep‐1 were cultured in DMEM medium, HL7702 and SNU398 were cultured in RPMI 1640 medium, and Hep3B and PLC/PRF/5 were cultured in MEM medium. These cell lines were incubated in at 37°C, 5% CO2. All of the medium were added 10% fetal bovine serum (Invitrogen Gibco, USA). SGC707 (MCE, cat# HY‐19715), 2‐deoxy‐D‐glucose (2‐DG) (MCE, cat# HY‐13966), oligomycin (MCE, cat# HY‐N6782) and XY‐1 (MCE, cat# HY‐19714) were purchased from MedChemExpress. Cycloheximide (Selleck, cat# HY‐12320) was purchased from Selleck.
Lei Y., Han P., Chen Y., Wang H., Wang S., Wang M., Liu J., Yan W., Tian D, & Liu M. (2022). Protein arginine methyltransferase 3 promotes glycolysis and hepatocellular carcinoma growth by enhancing arginine methylation of lactate dehydrogenase A. Clinical and Translational Medicine, 12(1), e686.
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7

Breast Cancer Cell Culture Protocol

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Cell lines were from the American Tissue Culture Collection (ATCC). MDA-MB-231 cells were cultured in DMEM with 10% fatal bovine serum (FBS) and 5% Penicillin/Streptomycin. BT549 and Hs578T cells were cultured per ATCC recommendations. Cycloheximide, bortezomib, carfilzomib, and MG132 were from Selleckchem (Houston, TX). Primers are listed in Tables 1 and 2, and antibodies are listed in Table 3.
Al-Marsoummi S., Vomhof-DeKrey E, & Basson M.D. (2019). Schlafen12 Reduces the Aggressiveness of Triple Negative Breast Cancer through Post-Transcriptional Regulation of ZEB1 That Drives Stem Cell Differentiation. Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology, 53(6), 999-1014.
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8

Cell line maintenance and treatments

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NCI-H1975, NCI-H1650, NCI-HCC827, NCI-H1299, A549 and PC9 cell lines were purchased from the Type Culture Collection of the Chinese Academy of Sciences (Shanghai, China). They were maintained in RPMI-1640 medium (Gibco-Life Technologies, NY, USA) supplemented with 10% fetal bovine serum (Gibco-Life Technologies), 100 μg/mL streptomycin and 100 U/mL penicillin (Gibco-Life Technologies). Cells were cultured at 37°C with a humid atmosphere of 5% CO2 and 95% air. Pemetrexed, decitabine, and cycloheximide were obtained from Selleck Chemicals (Houston, TX, USA).
Wang Y., Huang J., Wu Q., Zhang J., Ma Z., Zhu L., Xia B., Ma S, & Zhang S. (2021). Decitabine Sensitizes the Radioresistant Lung Adenocarcinoma to Pemetrexed Through Upregulation of Folate Receptor Alpha. Frontiers in Oncology, 11, 668798.
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9

Evaluation of Hu-17 Anticancer Potential

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Forskolin, exemestane, formestane, cisplatin, and PD98059 were purchased from Sigma Chemical Co. MG132, Z‐VAD‐FMK and cycloheximide were purchased from Selleckchem. Paclitaxel was kindly provided by the Shanghai Key Laboratory of Gynecologic Oncology, Ren Ji Hospital. Hu‐17 was synthesized by the laboratory of Professor Yanghua Yi, Second Military Medical University. Hu‐17 (molecular weight 1084 Da; structure in Figure 1) was stored at −20°C as a stock solution (20 mmol/L) in dimethyl sulfoxide.
Xi Y., Liu J., Wang H., Li S., Yi Y, & Du Y. (2020). New small‐molecule compound Hu‐17 inhibits estrogen biosynthesis by aromatase in human ovarian granulosa cancer cells. Cancer Medicine, 9(23), 9081-9095.
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10

Gastric Cancer Cell Line Culture

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Human GC cell lines, including AGS, BGC-823, MGC-803, MKN45, SGC-7901, and human normal gastric epithelial cell line GES-1 were purchased from iCell Bioscience Inc. (Shanghai, China). AGS, BGC-823, and MGC-803 were cultured in DMEM medium, and MKN45, SGC-7901, and GES-1 were cultured in RPMI-1640 medium. All the mediums were purchased from Gibco (Carlsbad, CA, USA) and 10% fetal bovine serum (Gibco) and 1% penicillin-streptomycin sulfate (Gibco) were added. 10 μM of MG132 (Selleckchem, Radnor, PA, USA), a proteasome inhibitor, or 100 μg/ml of cycloheximide (Selleckchem), a protein synthesis inhibitor were used to treat cells.
Xie H., He Y., Wu Y, & Lu Q. (2021). Silencing of UBE2D1 inhibited cell migration in gastric cancer, decreasing ubiquitination of SMAD4. Infectious Agents and Cancer, 16, 63.
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