Structure-Selective Differentiation of Deletion Mutation in Circulating Tumor DNA by Dual Probe-Based Isothermal Amplification

A rapid dual probe-based fluorimetric assay was developed to detect deletion mutation in circulating tumor DNA, through structure-selective isothermal amplification and pattern recognition. This method can detect both homozygous and...

Transcriptional co-activator regulates melanocyte differentiation and oncogenesis by integrating cAMP and MAPK/ERK pathways

SummaryThe cyclic AMP pathway promotes melanocyte differentiation in part by triggering gene expression changes mediated by CREB and its coactivators (CRTC1-3). Differentiation is dysregulated in melanomas, although the contributions of different cAMP effectors in this setting is unclear. We report a selective differentiation impairment in CRTC3 KO melanocytes and melanoma cells, due to downregulation of OCA2 and block of melanosome maturation. CRTC3 stimulated OCA2 expression via binding to CREB on a conserved enhancer, a regulatory site for pigmentation and melanoma risk in humans. Response to cellular signaling differed between CRTC3 and its family members; CRTC3 was uniquely activated by ERK1/2-mediated phosphorylation at Ser391 and by low levels of cAMP. Phosphorylation at Ser391 was constitutively elevated in human melanoma cells with hyperactivated ERK1/2 signaling; knockout of CRTC3 in this setting impaired anchorage-independent growth, migration and invasiveness while CRTC3 overexpression supported cell survival in response to MAPK inhibition by vemurafenib. Human melanomas expressing gain of function mutations in CRTC3 were associated with poorer clinical outcome. Our results suggest that CRTC3 inhibition may provide benefit in the treatment of hyperpigmentation and melanoma, and potentially other disorders with deregulated cAMP/MAPK crosstalk.

Transcriptome Analysis of Induced Pluripotent Stem Cell (iPSC)-derived Pancreatic β-like Cell Differentiation

Diabetes affects millions of people worldwide, and β-cell replacement is one of the promising new strategies for treatment. Induced pluripotent stem cells (iPSCs) can differentiate into any cell type, including pancreatic β cells, providing a potential treatment for diabetes. However, the molecular mechanisms underlying the differentiation of iPSC-derived β cells have not yet been fully elucidated. Here, we generated pancreatic β-like cells from mouse iPSCs using a 3-step protocol and performed deep RNA sequencing to get a transcriptional landscape of iPSC-derived pancreatic β-like cells during the selective differentiation period. We then focused on the differentially expressed genes (DEGs) during the time course of the differentiation period, and these genes underwent Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathway analysis. In addition, gene-act networks were constructed for these DEGs, and the expression of pivotal genes detected by quantitative real-time polymerase chain reaction was well correlated with RNA sequence (RNA-seq). Overall, our study provides valuable information regarding the transcriptome changes in β cells derived from iPSCs during differentiation, elucidates the biological process and pathways underlying β-cell differentiation, and promotes the identification and functional analysis of potential genes that could be used for improving functional β-cell generation from iPSCs.