Induction of secreted acid phosphatase (APase) is a universal response of higher plants to phosphate (Pi) limitation. These enzymes are thought to scavenge Pi from organophosphate compounds in the rhizosphere and thus to increase Pi availability to plants when Pi is deficient. The tight association of secreted APase with the root surface may make plants more efficient in the utilization of soil Pi around root tissues, which is present in organophosphate forms. To date, however, no systematic molecular, biochemical, and functional studies have been reported for any of the Pi starvation-induced APases that are associated with the root surface after secretion. In this work, using genetic and molecular approaches, we identified Arabidopsis (Arabidopsis thaliana) Purple Acid Phosphatase10 (AtPAP10) as a Pi starvation-induced APase that is predominantly associated with the root surface. The AtPAP10 protein has phosphatase activity against a variety of substrates. Expression of AtPAP10 is specifically induced by Pi limitation at both transcriptional and posttranscriptional levels. Functional analyses of multiple atpap10 mutant alleles and overexpressing lines indicated that AtPAP10 plays an important role in plant tolerance to Pi limitation. Genetic manipulation of AtPAP10 expression may provide an effective means for engineering new crops with increased tolerance to Pi deprivation.
IntroductionThe miR-183/-96/-182 cluster is a conserved polycistronic microRNA (miRNA) cluster which is highly expressed in most breast cancers. Although there are some sporadic reports which demonstrate the importance of each miRNA in this cluster in breast cancer, the biological roles of this cluster as a whole and its regulation mechanisms in breast cancer are still unclear. We compared the expression of this cluster in different cancer types, analyzed the regulation mechanism of this cluster, identified new target genes, and examined the impact of this cluster on breast cancer cells.MethodsThe miRNA level was detected by LNA-based northern blot and Real-time PCR, and was also analyzed from TCGA dataset. Bioinformatics research and luciferase assay were applied to find the promoter regions and transcription factors. To investigate the biological effects of the miR-183/-96 /-182 cluster in breast cancer, we generated miR-96, miR-182 and miR-183 overexpression stable cell lines to check the overdose effects; we also used miR-Down™ antagomir for each miRNA as well as miR-183/-96 /-182 cluster sponge lentivirus to check the knockdown effects. Growth, migration, cell cycle profile and survival of these cells was then monitored by colony formation assay, MTT assay, cell wound healing assay, flow cytometry and microscopy. The target gene was validated by Real-time PCR, luciferase assay, Western blot and Phalloidin/DAPI counterstaining.ResultsThe miR-183/-96/-182 cluster was highly expressed in most breast cancers, and its transcription is disordered in breast cancer. The miR-183/-96/-182 cluster was transcribed in the same pri-miRNA and its transcription was regulated by ZEB1 and HSF2. It increased breast cell growth by promoting more rapid completion of mitosis, promoted cell migration and was essential for cell survival. MiR-183 targeted the RAB21 mRNA directly in breast cancer.ConclusionThe miR-183/-96/-182 cluster is up-regulated in most breast cancer. It functions as an oncogene in breast cancer as it increases cell proliferation and migration.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-014-0473-z) contains supplementary material, which is available to authorized users.
Purpose Lung squamous cell carcinoma (LSCC) currently lacks effective targeted therapies. Previous studies reported overexpression of HEDGEHOG (HH)-GLI signaling components in LSCC. However, they addressed neither the tumor heterogeneity nor the requirement for HH-GLI signaling. Here, we investigated the role of HH-GLI signaling in LSCC, and studied the therapeutic potential of HH-GLI suppression. Experimental Design Gene expression datasets of two independent LSCC patient cohorts were analyzed to study the activation of HH-GLI signaling. Four human LSCC cell lines were examined for HH-GLI signaling components. Cell proliferation and apoptosis were assayed in these cells after blocking the HH-GLI pathway by lentiviral-shRNA knockdown or small molecule inhibitors. Xenografts in immunodeficient mice were used to determine the in vivo efficacy of GLI inhibitor GANT61. Results In both cohorts, activation of HH-GLI signaling was significantly associated with the classical subtype of LSCC. In cell lines, genetic knockdown of SMO produced minor effects on cell survival, while GLI2 knockdown significantly reduced proliferation and induced extensive apoptosis. Consistently, the SMO inhibitor GDC-0449 resulted in limited cytotoxicity in LSCC cells, whereas the GLI inhibitor GANT61 was very effective. Importantly, GANT61 demonstrated specific in vivo anti-tumor activity in xenograft models of GLI-positive cell lines. Conclusion Our studies demonstrate an important role for GLI2 in LSCC, and suggest GLI inhibition as a novel and potent strategy to treat a subset of LSCC patients.
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