Journal: The Journal of Biological Chemistry

Article Title: Lin-28 Homologue A (LIN28A) Promotes Cell Cycle Progression via Regulation of Cyclin-dependent Kinase 2 (CDK2), Cyclin D1 (CCND1), and Cell Division Cycle 25 Homolog A (CDC25A) Expression in Cancer *

doi: 10.1074/jbc.M111.321158

Figure Lengend Snippet: Reactivation of strong LIN28A expression in human epithelial tumor specimens. LIN28A protein expression in six types of common human epithelial tumors (n = 369) was examined using tumor tissue arrays. Strong LIN28A expression was detected in ∼10% of the tumor specimens. Top panels, examples of LIN28A-negative tumors; bottom panels, examples of LIN28A-positive tumors.

Article Snippet: The pSin-EF2-LIN28A lentiviral expression vector was purchased from Addgene, and the pSin-EF2 empty vector was used as control.

Techniques: Expressing

Journal: The Journal of Biological Chemistry

Article Title: Lin-28 Homologue A (LIN28A) Promotes Cell Cycle Progression via Regulation of Cyclin-dependent Kinase 2 (CDK2), Cyclin D1 (CCND1), and Cell Division Cycle 25 Homolog A (CDC25A) Expression in Cancer *

doi: 10.1074/jbc.M111.321158

Figure Lengend Snippet: Expression of LIN28A in human breast and ovarian tumors. A, expression of LIN28A in tumor cells was confirmed by real-time RT-PCR (top panels) and Western blots (bottom panels) in breast and ovarian cancer cell lines. Two cultured mammary gland epithelial cell lines and four cultured ovarian surface epithelial cell lines were used as controls. B, expression of LIN28A in human benign and DCIS tumor specimens was confirmed by real-time RT-PCR (top panels) and Western blots (bottom panels). T47D cells were used as positive controls in the Western blots.

Article Snippet: The pSin-EF2-LIN28A lentiviral expression vector was purchased from Addgene, and the pSin-EF2 empty vector was used as control.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Cell Culture

Journal: The Journal of Biological Chemistry

Article Title: Lin-28 Homologue A (LIN28A) Promotes Cell Cycle Progression via Regulation of Cyclin-dependent Kinase 2 (CDK2), Cyclin D1 (CCND1), and Cell Division Cycle 25 Homolog A (CDC25A) Expression in Cancer *

doi: 10.1074/jbc.M111.321158

Figure Lengend Snippet: LIN28A promotes cancer cell growth in vitro. A, two independent shRNAs were used to stably knock down LIN28A expression in A2780, IGROV1, and T47D cell lines. Inhibition of LIN28A by shRNAs was confirmed by both real-time RT-PCR and Western blots. B, the in vitro growth of LIN28A knockdown and control cells was monitored by cell counts. C and D, anchorage-independent growth of LIN28A knockdown and control cells was examined using a soft agar colony formation assay. Error bars, S.D.

Article Snippet: The pSin-EF2-LIN28A lentiviral expression vector was purchased from Addgene, and the pSin-EF2 empty vector was used as control.

Techniques: In Vitro, Stable Transfection, Expressing, Inhibition, Quantitative RT-PCR, Western Blot, Soft Agar Assay

Journal: The Journal of Biological Chemistry

Article Title: Lin-28 Homologue A (LIN28A) Promotes Cell Cycle Progression via Regulation of Cyclin-dependent Kinase 2 (CDK2), Cyclin D1 (CCND1), and Cell Division Cycle 25 Homolog A (CDC25A) Expression in Cancer *

doi: 10.1074/jbc.M111.321158

Figure Lengend Snippet: LIN28A promotes cancer cell growth in vivo. A and B, stable knockdown of LIN28A expression in vivo was confirmed by Western blots (A) and real-time RT-PCR (B). *, p < 0.05. C, LIN28A knockdown and control cells were transplanted into nude mice, and tumor growth was monitored every 3 days. Error bars, S.D.

Article Snippet: The pSin-EF2-LIN28A lentiviral expression vector was purchased from Addgene, and the pSin-EF2 empty vector was used as control.

Techniques: In Vivo, Expressing, Western Blot, Quantitative RT-PCR

Journal: The Journal of Biological Chemistry

Article Title: Lin-28 Homologue A (LIN28A) Promotes Cell Cycle Progression via Regulation of Cyclin-dependent Kinase 2 (CDK2), Cyclin D1 (CCND1), and Cell Division Cycle 25 Homolog A (CDC25A) Expression in Cancer *

doi: 10.1074/jbc.M111.321158

Figure Lengend Snippet: LIN28A promotes cell cycle progression in cancer cells. A and B, LIN28A was stably knocked down in A2780 and IGROV1 cells using shRNA. The cell cycle was analyzed using BrdU labeling and flow cytometry; stable inhibition of endogenous LIN28A expression significantly blocked cell cycle in both cell lines. C, two independent siRNAs were used to confirm the above findings. siRNA and control oligonucleotides were transiently transfected into A2780 cells, and the cell cycle was analyzed using BrdU labeling and flow cytometry. Transient inhibition of endogenous LIN28A expression significantly blocked cell cycle in these cells. D, immunochemical staining for the proliferation marker Ki-67 in LIN28A knockdown and control A2780 xenograft tumors. *, p < 0.05. Error bars, S.D.

Article Snippet: The pSin-EF2-LIN28A lentiviral expression vector was purchased from Addgene, and the pSin-EF2 empty vector was used as control.

Techniques: Stable Transfection, shRNA, Labeling, Flow Cytometry, Inhibition, Expressing, Transfection, Staining, Marker

Journal: The Journal of Biological Chemistry

Article Title: Lin-28 Homologue A (LIN28A) Promotes Cell Cycle Progression via Regulation of Cyclin-dependent Kinase 2 (CDK2), Cyclin D1 (CCND1), and Cell Division Cycle 25 Homolog A (CDC25A) Expression in Cancer *

doi: 10.1074/jbc.M111.321158

Figure Lengend Snippet: LIN28A binds to thousands of mRNAs, including a large group of cell cycle regulatory mRNAs. A, RNA-IP-chip technology was used to search for the whole-genome-wide LIN28A-binding RNAs in tumor and ES cells. B, microarray experiments found that 1,707 and 2,806 mRNA transcripts bound to LIN28A in A2780 cells and ES cells, respectively. Notably, there were 801 mRNA transcripts shared by both microarray experiments. C, the molecular category of the transcripts that bound to LIN28A in A2780 cells was explored by GO analysis. Pathways associated with cell cycle regulation were the most frequently identified pathways. The molecular category of the transcripts that bound to LIN28A in both A2780 and ES cells was also explored by GO analysis. Again, pathways associated with cell cycle regulation were also the most frequently identified pathways. Notably, the ranking of the cell cycle-associated pathways was higher when using transcripts that bound to both A2780 and ES cells, compared with the analysis using A2780 cells alone. D, a list of cyclins, cyclin-dependent kinases, and proteins involved in the control of the cell cycle that were identified by RNA-IP-chip in A2780 cells. The genes in green were also identified in ES cells using RNA-IP-chip.

Article Snippet: The pSin-EF2-LIN28A lentiviral expression vector was purchased from Addgene, and the pSin-EF2 empty vector was used as control.

Techniques: Genome Wide, Binding Assay, Microarray

Journal: The Journal of Biological Chemistry

Article Title: Lin-28 Homologue A (LIN28A) Promotes Cell Cycle Progression via Regulation of Cyclin-dependent Kinase 2 (CDK2), Cyclin D1 (CCND1), and Cell Division Cycle 25 Homolog A (CDC25A) Expression in Cancer *

doi: 10.1074/jbc.M111.321158

Figure Lengend Snippet: LIN28A regulates CDK2 protein expression in cancer cells. A, the RNA-IP-chip results were further validated in two tumor cell lines (A2780 and IGROV1) by real-time RT-PCR. The LIN28A-RNA-IP significantly enriched CDK2, CDC2, CDC20, CCNA2, and CCNB2 proteins compared with the IgG control. B, LIN28A expression was stably knocked down by shRNA in A2780 and IGROV1 cells in vitro. The expression of LIN28A-binding targets was detected by Western blots. Blocking endogenous LIN28A remarkably reduced CDK2, CDC2, and CDC20 protein expression in vitro. C, LIN28A expression was stably knocked down by shRNA in A2780 xenograft tumors in vivo. The expression of LIN28A-binding targets was detected by immunohistochemistry. Blocking endogenous LIN28A remarkably reduced CDK2, CDC2, and CDC20 protein expression in vivo. *, p < 0.05. Error bars, S.D.

Article Snippet: The pSin-EF2-LIN28A lentiviral expression vector was purchased from Addgene, and the pSin-EF2 empty vector was used as control.

Techniques: Expressing, Quantitative RT-PCR, Stable Transfection, shRNA, In Vitro, Binding Assay, Western Blot, Blocking Assay, In Vivo, Immunohistochemistry

Journal: The Journal of Biological Chemistry

Article Title: Lin-28 Homologue A (LIN28A) Promotes Cell Cycle Progression via Regulation of Cyclin-dependent Kinase 2 (CDK2), Cyclin D1 (CCND1), and Cell Division Cycle 25 Homolog A (CDC25A) Expression in Cancer *

doi: 10.1074/jbc.M111.321158

Figure Lengend Snippet: LIN28A regulates CCND1 and CDC25A expression via a let-7-dependent mechanism. A, using real-time RT-PCR, we showed that knocking down LIN28A expression reduced the expression of CCND1 and CDC25A mRNA in A2780 and IGROV1 cells. B, transfection with a let-7 mimic reduced CCND1 and CDC25A mRNA expression in A2780 and IGROV1 cells. C, knocking down LIN28A or enforcing the expression of let-7 reduced CCND1 and CDC25A protein expression in A2780 and IGROV1 cells. D, knocking down LIN28A reduced CCND1 and CDC25A protein expression in A2780 xenograft tumors. *, p < 0.05. Error bars, S.D.

Article Snippet: The pSin-EF2-LIN28A lentiviral expression vector was purchased from Addgene, and the pSin-EF2 empty vector was used as control.

Techniques: Expressing, Quantitative RT-PCR, Transfection