Dataset Information

Direct RNA sequencing mediated identification of mRNA localized in protrusions of human MDA-MB-231 metastatic breast cancer cells.

ABSTRACT: BACKGROUND:Protrusions of cancer cells conferrers a vital function for cell migration and metastasis. Protein and RNA localization mechanisms have been extensively examined and shown to play pivotal roles for the functional presence of specific protein components in cancer cell protrusions. METHODS:To describe genome wide RNA localized in protrusions of the metastatic human breast cancer cell line MDA-MB-231 we used Boyden chamber based methodology followed by direct mRNA sequencing. RESULTS:In the hereby identified group of protrusion localized mRNA some previously were described to be localized exemplified by mRNA for Ras-Related protein 13 (RAB13) and p0071 (Plakophilin-4/PKP4). For other transcripts, exemplified by mRNA for SH3PXD2A/TKS5 and PPFIA1/Liprin-?1, only the corresponding proteins previously were described to have protrusion localization. Finally, a cohort of MDA-MB-231 protrusion localized transcripts represents novel candidates to mediate cancer cell subcellular region specific functions through mRNA direction to protrusions. We included a further characterization of p0071, an armadillo repeat protein of adherence junctions and desmosomes, in MDA-MB-231 and non-metastatic MCF7 cells including analysis of novel identified alternative spliced p0071 mRNA isoforms. The results showed isoform and cell type specific p0071 mRNA localization. CONCLUSIONS:Altogether, the presented data represents a genome wide and gene specific descriptive and functional analyses of RNA localization in protrusions of MDA-MB-231 metastatic cancer cells.

SUBMITTER: Jakobsen KR

PROVIDER: S-EPMC3844448 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Direct RNA sequencing mediated identification of mRNA localized in protrusions of human MDA-MB-231 metastatic breast cancer cells.

Jakobsen Kristine Raaby KR Sørensen Emilie E Brøndum Karin Kathrine KK Daugaard Tina Fuglsang TF Thomsen Rune R Nielsen Anders Lade AL

Journal of molecular signaling 20130901 1

<h4>Background</h4>Protrusions of cancer cells conferrers a vital function for cell migration and metastasis. Protein and RNA localization mechanisms have been extensively examined and shown to play pivotal roles for the functional presence of specific protein components in cancer cell protrusions.<h4>Methods</h4>To describe genome wide RNA localized in protrusions of the metastatic human breast cancer cell line MDA-MB-231 we used Boyden chamber based methodology followed by direct mRNA sequenci ...[more]

PMID: 24004954

Similar Datasets

Project description:Breast cancer is an important worldwide public health concern. The incidence rate of breast cancer increases every year. The primary cause of death is metastasis, a process by which cancer cells spread from a primary site to secondary organs. MicroRNAs (miRs/miRNAs) are small non-coding RNAs that control gene expression at the post-transcriptional level. Dysregulation of certain miRNAs is involved in carcinogenesis, cancer cell proliferation and metastasis. Therefore, the present study assessed miRNAs associated with breast cancer metastasis using two breast cancer cell lines, the low-metastatic MCF-7 and the highly metastatic MDA-MB-231. miRNA array analysis of both cell lines indicated that 46 miRNAs were differentially expressed when compared between the two cell lines. A total of 16 miRNAs were upregulated in MDA-MB-231 compared with MCF-7 cells, which suggested that their expression levels may be associated with the highly invasive phenotype of MDA-MB-231 cells. Among these miRNAs, miR-222-3p was selected for further study and its expression was confirmed by reverse transcription-quantitative PCR (RT-qPCR). Under both non-adherent and adherent culture conditions, the expression levels of miR-222-3p in the MDA-MB-231 cell line were higher than those noted in the MCF-7 cell line under the same conditions. Suppression of endogenous miR-222-3p expression in MDA-MB-231 cells using a miR-222-3p inhibitor resulted in a 20-40% reduction in proliferation, and a ~30% reduction in migration, which suggested that the aggressive phenotype of MDA-MB-231 cells was partly regulated by miR-222-3p. Bioinformatic analysis of miR-222-3p using TargetScan 8.0, miRDB and PicTar identified 25 common mRNA targets, such as cyclin-dependent kinase inhibitor 1B, ADP-ribosylation factor 4, iroquois homeobox 5 and Bcl2 modifying factor. The results of the present study indicated that miR-222-3p was potentially associated with the proliferation and migratory ability of the MDA-MB-231 cell line.

Project description:Since bone metastatic breast cancer is an incurable disease, causing significant morbidity and mortality, understanding of the underlying molecular mechanisms would be highly valuable. Here, we describe in vitro and in vivo evidence for the importance of serine biosynthesis in the metastasis of breast cancer to bone. We first characterized the bone metastatic propensity of the MDA-MB-231(SA) cell line variant as compared to the parental MDA-MB-231 cells by radiographic and histological observations in the inoculated mice. Genome-wide gene expression profiling of this isogenic cell line pair revealed that all the three genes involved in the L-serine biosynthesis pathway, phosphoglycerate dehydrogenase (PHGDH), phosphoserine aminotransferase 1 (PSAT1), and phosphoserine phosphatase (PSPH) were upregulated in the highly metastatic variant. This pathway is the primary endogenous source for L-serine in mammalian tissues. Consistently, we observed that the proliferation of MDA-MB-231(SA) cells in serine-free conditions was dependent on PSAT1 expression. In addition, we observed that L-serine is essential for the formation of bone resorbing human osteoclasts and may thus contribute to the vicious cycle of osteolytic bone metastasis. High expression of PHGDH and PSAT1 in primary breast cancer was significantly associated with decreased relapse-free and overall survival of patients and malignant phenotypic features of breast cancer. In conclusion, high expression of serine biosynthesis genes in metastatic breast cancer cells and the stimulating effect of L-serine on osteoclastogenesis and cancer cell proliferation indicate a functionally critical role for serine biosynthesis in bone metastatic breast cancer and thereby an opportunity for targeted therapeutic interventions. Parental MDA-MB-231 cells and MDA-MB-231(SA) cells were cultured in cell culture flasks. RNA was isolated in order to compare the gene expression profiles of these cell variants. Total of two samples. No replicates.

Project description:Breast cancer remains a leading cause of death in women worldwide. Although breast cancer therapies have greatly advanced in recent years, many patients still develop tumour recurrence and metastasis, and eventually succumb to the disease due to chemoresistance. Citral has been reported to show cytotoxic effect on various cancer cell lines. However, the potential of citral to specifically target the drug resistant breast cancer cells has not yet been tested, which was the focus of our current study.The cytotoxic activity of citral was first tested on MDA-MB-231 cells in vitro by MTT assay. Subsequently, spheroids of MDA-MB-231 breast cancer cells were developed and treated with citral at different concentrations. Doxorubicin, cisplatin and tamoxifen were used as positive controls to evaluate the drug resistance phenotype of MDA-MB-231 spheroids. In addition, apoptosis study was performed using AnnexinV/7AAD flowcytometry. Aldefluor assay was also carried out to examine whether citral could inhibit the ALDH-positive population, while the potential mechanism of the effect of citral was carried out by using quantitative real time- PCR followed by western blotting analysis.Citral was able to inhibit the growth of the MDA-MB-231 spheroids when compared to a monolayer culture of MDA-MB-231 cells at a lower IC50 value. To confirm the inhibition of spheroid self-renewal capacity, the primary spheroids were then cultured to additional passages in the absence of citral. A significant reduction in the number of secondary spheroids were formed, suggesting the reduction of self-renewal capacity of these aldehyde dehydrogenase positive (ALDH+) drug resistant spheroids. Moreover, the AnnexinV/7AAD results demonstrated that citral induced both early and late apoptotic changes in a dose-dependent manner compared to the vehicle control. Furthermore, citral treated spheroids showed lower cell renewal capacity compared to the vehicle control spheroids in the mammosphere formation assay. Gene expression studies using quantitative real time PCR and Western blotting assays showed that citral was able to suppress the self-renewal capacity of spheroids and downregulate the Wnt/?-catenin pathway.The results suggest that citral could be a potential new agent which can eliminate drug-resistant breast cancer cells in a spheroid model via inducing apoptosis.

Dataset Information

Direct RNA sequencing mediated identification of mRNA localized in protrusions of human MDA-MB-231 metastatic breast cancer cells.

Publications

Direct RNA sequencing mediated identification of mRNA localized in protrusions of human MDA-MB-231 metastatic breast cancer cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets