CRISPR Screens Identify Essential Cell Growth Mediators in BRAF Inhibitor-resistant Melanoma / 基因组蛋白质组与生物信息学报·英文版

BRAF is a serine/threonine kinase that harbors activating mutations in ∼7% of human malignancies and ∼60% of melanomas. Despite initial clinical responses to BRAF inhibitors, patients frequently develop drug resistance. To identify candidate therapeutic targets for BRAF inhibitor resistant melanoma, we conduct CRISPR screens in melanoma cells harboring an activating BRAF mutation that had also acquired resistance to BRAF inhibitors. To investigate the mechanisms and pathways enabling resistance to BRAF inhibitors in melanomas, we integrate expression, ATAC-seq, and CRISPR screen data. We identify the JUN family transcription factors and the ETS family transcription factor ETV5 as key regulators of CDK6, which together enable resistance to BRAF inhibitors in melanoma cells. Our findings reveal genes contributing to resistance to a selective BRAF inhibitor PLX4720, providing new insights into gene regulation in BRAF inhibitor resistant melanoma cells.

TDP-43 regulates cancer-associated microRNAs

Aberrant regulation of miRNA genes contributes to pathogenesis of a wide range of human diseases, including cancer. The TAR DNA binding protein 43 (TDP-43), a RNA/DNA binding protein associated with neurodegeneration, is involved in miRNA biogenesis. Here, we systematically examined miRNAs regulated by TDP-43 using RNA-Seq coupled with an siRNA-mediated knockdown approach. TDP-43 knockdown affected the expression of a number of miRNAs. In addition, TDP-43 down-regulation led to alterations in the patterns of different isoforms of miRNAs (isomiRs) and miRNA arm selection, suggesting a previously unknown role of TDP-43 in miRNA processing. A number of TDP-43 associated miRNAs, and their candidate target genes, are associated with human cancers. Our data reveal highly complex roles of TDP-43 in regulating different miRNAs and their target genes. Our results suggest that TDP-43 may promote migration of lung cancer cells by regulating miR-423-3p. In contrast, TDP-43 increases miR-500a-3p expression and binds to the mature miR-500a-3p sequence. Reduced expression of miR-500a-3p is associated with poor survival of lung cancer patients, suggesting that TDP-43 may have a suppressive role in cancer by regulating miR-500a-3p. Cancer-associated genes LIF and PAPPA are possible targets of miR-500a-3p. Our work suggests that TDP-43-regulated miRNAs may play multifaceted roles in the pathogenesis of cancer.

Biocompatibility of macrophages with quantum dots / 中国组织工程研究

BACKGROUND: Compared with the traditional organic fluorescent dyes, quantum dots present good biomarker characteristics. Especially, quantum dots for cell labeling and targeted bioimaging present unique optical properties.OBJECTIVE: To investigate the biocompatibility of CdSe/ZnS quantum dots with mononuclear macrophages.METHODS: The macrophages RAW264.7 were inoculated into 96-well plates containing 0, 50, 100 mg/L CdSe/ZnS quantum dots for 1 or 2 hours. Then, the fluorescent signal was detected by flow cytometry. After 0-24 hours of culture,the fluorescence signal intensity of the macrophages cultured with 50 mg/L CdSe/ZnS quantum dots was detected by flow cytometry. After 18 hours of culture, quantitative PCR was used to detect the levels of tumor necrosis factor α and interleukin 1β in macrophages, and macrophage proliferation cell apoptosis were detected by MTT and flow cytometry,respectively.RESULTS AND CONCLUSION: The fluorescence signal intensity was positively correlated with the mass concentration of CdSe/ZnS quantum dots, and the intensity of the fluorescent signal was increased with the labeling time. After labeling using 50 mg/L CdSe/ZnS quantum dots, the fluorescence signal of macrophages increased continuously with time, and reached the peak at 18 hours. Compared with 0 mg/L quantum dot group, 50, 100mg/L quantum dot groups could significantly promote the expression of tumor necrosis factor α and interleukin 1β in macrophages (P < 0.01 or P < 0.05).The level of tumor necrosis factor α in the 100 mg/L quantum dot group was higher than that in the 50 mg/L quantum dot group (P < 0.01). The expression of interleukin-1β showed no difference between 50 and 100 mg/L quantum dot groups.The cell proliferation in the 50 and 100 mg/L CdSe/ZnS quantum dot groups was significantly higher than that in the 0 mg/L quantum dot group (P < 0.05), but there was no difference between the former two groups. In addition, 50 and 100 mg/L CdSe/ZnS quantum dots had no significant effect on apoptosis of macrophages. To conclude, CdSe/ZnS quantum dots could activate macrophages and promote their proliferation and secretion of inflammatory factors, but did not affect their apoptosis.