Mcm5 Represses Endodermal Migration through Cxcr4a-itgb1b Cascade Instead of Cell Cycle Control.

Minichromosome maintenance protein 5 (MCM5) is a critical cell cycle regulator; its role in DNA replication is well known, but whether it is involved in the regulation of organogenesis in a cell cycle-independent way, is far from clear. In this study, we found that a loss of mcm5 function resulted in a mildly smaller liver, but that mcm5 overexpression led to liver bifida. Further, the data showed that mcm5 overexpression delayed endodermal migration in the ventral-dorsal axis and induced the liver bifida. Cell cycle analysis showed that a loss of mcm5 function, but not overexpression, resulted in cell cycle delay and increased cell apoptosis during gastrulation, implying that liver bifida was not the result of a cell cycle defect. In terms of its mechanism, our data proves that mcm5 represses the expression of cxcr4a, which sequentially causes a decrease in the expression of itgb1b during gastrulation. The downregulation of the cxcr4a-itgb1b cascade leads to an endodermal migration delay during gastrulation, as well as to the subsequent liver bifida during liver morphogenesis. In conclusion, our results suggest that in a cell cycle-independent way, mcm5 works as a gene expression regulator, either partially and directly, or indirectly repressing the expression of cxcr4a and the downstream gene itgb1b, to coordinate endodermal migration during gastrulation and liver location during liver organogenesis.

ALKBH5 promotes cadmium-induced transformation of human bronchial epithelial cells by regulating PTEN expression in an m6A-dependent manner.

Cadmium is a carcinogenic heavy metal that poses a severe threat to human beings. The underlying mechanism, however, remains elusive. N6-methyladenosine (m6A) is the most abundant post-transcriptional modification in mRNA that regulates RNA metabolism. Emerging evidence shows that m6A is involved in the pathogenesis of various cancers. In this study, human bronchial epithelial BEAS-2B cells were transformed by exposing to 2 µM of cadmium for 20 weeks to investigate the role of m6A in cadmium carcinogenesis. We found the level of m6A in mRNA was significantly decreased in cadmium-transformed BEAS-2B cells, and this change was regulated by m6A demethylase ALKBH5. ALKBH5 was significantly upregulated in the middle and late stages of cell transformation at week 8, 12, 16 and 20. Knockdown of ALKBH5 in cadmium-transformed cells alleviated cell proliferation, migration, invasion, and anchorage-independent growth, but co-transfection with ALKBH5 siRNA and PTEN siRNA restored the inhibitory effects of ALKBH5 knockdown on those transformation properties. ALKBH5 decreased the m6A level of PTEN mRNA, resulting in its instability and reduction of PTEN protein expression. These results indicate that ALKBH5-mediated demethylation m6A at PTEN mRNA is involved in cadmium-induced cell transformation. Our study provides a new perspective for the involvement of m6A modification in cadmium carcinogenesis.

Procyanidins mediates antineoplastic effects against non-small cell lung cancer via the JAK2/STAT3 pathway.

BACKGROUND: Lung cancer is a malignant tumor with one of the highest rates of cancer-related morbidity and mortality worldwide. Non-small cell lung cancer (NSCLC) account for 85% of all lung cancers and have a poor prognosis. Proanthocyanidins (PCs) are polyphenolic compounds that are found widely in natural plants. The present study aimed to determine the effects of PC on lung cancer and identify its possible mechanism. METHODS: A cell growth assay was used to detect the cell growth ability of A549 cancer cells, and a clonal formation assay was used to detect the cloning ability of A549 cancer cells. Flow cytometry was used to detect the effect of PCs on apoptosis and the cell cycle. The wound healing test, Transwell migration, and invasion test were used to detect the migration and invasion of human NSCLC A549 cells. Western blotting was utilized to detect the expression levels of N-cadherin, E-cadherin, vimentin, Janus kinase 2 (JAK2), p-signal transducer and activator of transcription 3 (p-STAT3), STAT3, matrix metalloproteinase 2 (MMP-2), MMP-9, and the apoptosis-related proteins, B-cell lymphoma-2 (Bcl-2) and BCL2-associated X (Bax). Cell immunofluorescence was used to detect the expression levels of the p-STAT3 primary antibody. RESULTS: PCs reduced the proliferation and cloning ability of A549 cells and significantly inhibited the migration and invasion of A549 cells in a dose-dependent manner. At the same time, PCs induced apoptosis in A549 cells and G2/M cell cycle arrest. PCs increased the pro-apoptotic protein expression, Bax, and down-regulated the anti-apoptotic protein expression, Bcl-2. PCs also inhibited the epithelial-mesothermal transition (EMT) process of A549 cells. We also found that the JAK2/STAT3 signaling pathway inhibitor, AG490, cooperated with PCs to inhibit A549 cell invasion and migration. Our results demonstrated that PCs could mediate the antitumor effect of NSCLC via the JAK2/STAT3 pathway. CONCLUSIONS: PCs can inhibit NSCLC A549 cell proliferation, invasion, metastasis, clone formation, EMT, and induced apoptosis and G2/M cell cycle arrest. They work by inhibiting the JAK2/STAT3 signaling pathway. As a novel antitumor drug, PCs have broad application prospects for the treatment of NSCLC.

The expression of zebrafish NAD(P)H:quinone oxidoreductase 1(nqo1) in adult organs and embryos.

NQO1, NAD(P)H: quinone oxidoreductase 1, was first identified in rat and its role has been extensively studied. Even the roles of NQO1 in the maintenance of physiological function and disease were largely addressed, whether the tissue specific functions of the NQO1 in organ development remains unknown. In the current study, we identified two NQO1 isoforms (isoform 1 and isoform 2) and examined the expression of nqo1 variants in adult zebrafish organs and embryos at different stages. In adult organs, RT-PCR result indicated that nqo1 variant 1 was mainly expressed in stomach and intestine, while nqo1 variant 2 was expressed in all organs investigated except for heart. Further, RT-PCR result showed that the nqo1 variant 1 and variant 2 were expressed at all the embryonic stages, but nqo1 variant 1 expression level was much lower than that of nqo1 variant 2. To specifically examine the expression pattern of these two different nqo1 variants, we did whole mount in situ hybridization and the results demonstrated that, both of them were maternally expressed at 8-cell stage, and they were all expressed ubiquitously at early stage. At 24 hpf, nqo1 variant 2 was mainly expressed in yolk cells, and slightly in head and eyes. At 48 hpf, nqo1 variant 2 was restricted in lateral line neuromasts. From 72 hpf to 144 hpf, nqo1 variant 2 was mainly restricted in branchial arch, liver, swimming bladder and lateral line neuromasts, while from 124 hpf to 192 hpf, nqo1 variant 2 only restricted in liver, and disappeared in lateral line neuromasts. On the contrary, at the late embryonic stage, nqo1 variant 1 was only expressed in liver and swimming bladder while not in branchial arch and lateral line neuromasts. In conclusion, we systematically analyzed the expression pattern of nqo1 variant 1 and variant 2 in zebrafish at different embryonic stages, and our data implied the possible role of nqo1 in regulating liver, branchial arch and lateral neuromasts development.