Crucial roles of UCH-L1 on insulin-producing cells and carbohydrate metabolism in Drosophila melanogaster model.

Ubiquitin carboxyl-terminal hydrolase L1 (UCH-L1) is a highly expressed protein in ß cells and has been implicated in ß cells' viability and function, however, the role of UCH-L1 in ß cells remains unclear. Herein, we examined the functions of UCH-L1 in ß cells by utilizing the Drosophila melanogaster model. Our results showed that specific knockdown of dUCH (D.melanogaster homolog of UCH-L1) in Drosophila Insulin-producing cells (D.melanogaster homolog of ß cells) induced mitochondria fusion, IPCs death/degeneration, interfered with DILP2 secretion, and triggered the rise of glycogen storage and body weight. Strikingly, the impairment in IPCs cellular activities can be rescued by vitamin C- a strong antioxidant compound, which suggested the relationship between knockdown dUCH and oxidative stress in IPCs; and the potential of this model in screening compounds for ß cells function moderation. Since carbohydrate metabolism is an important function of beta cells, we continued to examine the ability to regulate carbohydrate metabolism of knockdown dUCH flies. Our results showed that knockdown dUCH caused the decline of IPCs number under a high-sucrose diet, which finally led to metabolic and physiological disturbances, including total lipid rise, glycogen storage reduction, circulating carbohydrate increase, and weight loss. These symptoms could be early indications of metabolic disorders, particularly ß cell dysfunction-related diseases. Taken together, our results indicate that dUCH is essential in the viability and functions of IPCs through the regulation of carbohydrate metabolism in the Drosophila model.

Elephantopus mollis Kunth extracts induce antiproliferation and apoptosis in human lung cancer and myeloid leukemia cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Elephantopus mollis Kunth (EM), which belongs to Asteraceae family, has been used as a folk medicine with diverse therapeutic properties. Previous studies reported that crude extracts of this plant could inhibit several cancer cell lines, including breast carcinoma MCF-7, liver carcinoma HepG2, colorectal carcinoma DLD-1, lung carcinoma NCI-H23, etc. AIM: In this study, the anticancer activity and associated molecular mechanism of EM which is distributed in Vietnam were investigated. MATERIALS AND METHODS: The cytotoxicity of various EM extracts was evaluated on different cell lines by MTT assay. In addition, the effects of EM extracts on cell growth, cell morphology, nuclear morphology, caspase-3 activation, and mRNA expression levels of apoptosis-related genes were also examined. RESULTS: Our results demonstrated that ethyl acetate extract (EM-EA) caused proliferative inhibition and apoptotic induction towards A549 lung cancer cells (IC50 = 18.66 µg/ml, SI = 5.8) and HL60 leukemia cells (IC50 = 7.45 µg/ml, SI = 14.5) while petroleum ether extract (EM-PE) showed high toxicity to HL60 cell line (IC50 = 11.14 µg/ml, SI = 6.7). Notably, Raji lymphoma cells were also affected by these extracts (IC50 < 20 µg/ml, SI > 4), which has not been reported yet. Furthermore, mechanisms of EM extracts were elucidated. The significant downregulation of PCNA mRNA level induced by EM-EA/PE extracts contributed to the cell-growth restraint. EM-EA extract might activate apoptosis in A549 cells through both extrinsic and intrinsic signaling pathways by causing a 1.55-fold increase in BID, 3.65-fold increase in BAK and 3.11-fold decrease in BCL-2 expression level. Meanwhile, with EM-EA-extract treatment, HL60 cells might encounter P53-dependent apoptotic deaths. CONCLUSIONS: The combination of antiproliferation and apoptosis activation contributed to the high efficacy of EM extracts. These findings not only proved the anticancer potential of EM but also provided further insights into the mechanisms of EM extracts.

Identification of the Drosophila skpA gene as a novel target of the transcription factor DREF.

SKPa is component of a Drosophila SCF complex that functions in combination with the ubiquitin-conjugating enzyme UbcD1. skpA null mutation results in centrosome overduplication, unusual chromatin condensation, defective endoreduplication and cell-cycle progression. While the molecular mechanisms that regulate expression of the skpA gene are poorly understood, the DNA replication-related element (DRE) and the DRE-binding factor (DREF) play important roles in regulating proliferation-related genes in Drosophila and DRE (5'-TATCGATA) and DRE-like (5'-CATCGATT) sequences were here found to be involved in skpA promoter activity. Thus both luciferase transient expression assays in cultured Drosophila S2 cells using skpA promoter-luciferase fusion plasmids and anti-lacZ immunostaining of various tissues from transgenic third instar larvae carrying the skpA promoter-lacZ fusion genes provided supportive evidence. Furthermore, anti-SKPa immunostaining of eye imaginal discs from flies overexpressing DREF showed ectopic expression of protein in the region posterior to the morphogenetic furrow where DREF is overexpressed. Knockdown of DREF in some tissues where SKPa distribution is well known almost completely abrogated the skpA gene expression. These findings, taken together, indicate that the Drosophila skpA gene is a novel target of the transcription factor DREF.