Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

The homeobox transcription factor Nkx2-1 regulates microRNAs controlling downstream gene silencing in lung epithelial cells (mRNA)

ABSTRACT: Cell-specific gene expression is achieved by a combination of mechanisms including transcriptional and post-transcriptional regulation. The transcription factor Nkx2-1, essential for lung cell differentiation, mainly acts in transcriptional activation but can directly or indirectly repress gene expression. microRNAs are a class of small non-coding RNA that control one of the major mechanisms of gene repression. To identify miRNAs regulated by Nkx2-1 that may mediate its repressing effects, we knocked-down Nkx2-1 in mouse lung epithelial cell lines and systematically identified targets by genome-wide miR and mRNA expression analyses. Nkx2-1 controls expression of miRs known to contribute to lung cell differentiation in development and disease and others not previously described. Amongst the significantly altered miRs, the mir-106a-363 cluster, miR-1195, miR-378, and miR-346 are directly correlated with the levels of Nkx2-1, whereas miR-200c/b, miR-221, and miR- 222 are inversely correlated. These miRNAs are expressed in embryonic lung at day E11.5, and/or E19.5 determined by in-situ hybridization. Expression of predicted targets of mir-1195, mir-346 and miR-200c and mir-221/222 were evaluated by mRNA expression microarrays in Nkx2-1 knockdown cells identifying those anti-correlated to the corresponding miRNA expression. Genes regulated by mir-1195, Cyp2s1 and Map3k2, by mir-346, Klf6, and miR-200c, Myb, Nfib, and Six1, were validated by qRT-PCR. Inhibition of mir-1195 confirms the inverse correlation of this miRNA with its putative targets Cyp2s1 and Map3k2. This miRNA-mRNA expression analysis identifies potential paths of Nkx2-1 mediated gene repression, and contributes to the understanding of gene regulation in lung epithelial differentiation and development. Nkx2-1 mRNA was knocked down in lung epithelial cells using a lentivirus expressing a shRNA targeting Nkx2-1 (n=3) and compared to empty vector controls (n=3).

ORGANISM(S): Mus musculus

SUBMITTER: Joshua Campbell

PROVIDER: E-GEOD-47054 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Transcription factor and microRNA interactions in lung cells: an inhibitory link between NK2 homeobox 1, miR-200c and the developmental and oncogenic factors Nfib and Myb.

Tagne Jean-Bosco JB Mohtar Omar R OR Campbell Joshua D JD Lakshminarayanan Meenakshi M Huang Jingshu J Hinds Anne C AC Lu Jining J Ramirez Maria I MI

Respiratory research 20150213

<h4>Background</h4>The transcription factor NK2 homeobox 1 (Nkx2-1) plays essential roles in epithelial cell proliferation and differentiation in mouse and human lung development and tumorigenesis. A better understanding of genes and pathways downstream of Nkx2-1 will clarify the multiple roles of this critical lung factor. Nkx2-1 regulates directly or indirectly numerous protein-coding genes; however, there is a paucity of information about Nkx2-1-regulated microRNAs (miRNAs).<h4>Methods and re ...[more]

PMID: 25763778

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Project description:Despite the high prevalence and poor outcome of patients with metastatic lung cancer, the mechanisms of tumour progression and metastasis remain largely uncharacterized. We modelled human lung adenocarcinoma, which frequently harbours activating point mutations in KRAS1 and inactivation of the p53-pathway2, using conditional alleles in mice3-5. Lentiviral-mediated somatic activation of oncogenic Kras and deletion of p53 in the lung epithelial cells of KrasLSL-G12D/+;p53flox/flox mice initiates lung adenocarcinoma development4. Although tumours are initiated synchronously by defined genetic alterations, only a subset become malignant, suggesting that disease progression requires additional alterations. Identification of the lentiviral integration sites allowed us to distinguish metastatic from non-metastatic tumours and determine the gene expression alterations that distinguish these tumour types. Cross-species analysis identified the NK-2 related homeobox transcription factor Nkx2-1 (Ttf-1/Titf1) as a candidate suppressor of malignant progression. In this mouse model, Nkx2-1-negativity is pathognomonic of high-grade poorly differentiated tumours. Gain- and loss-of-function experiments in cells derived from metastatic and non-metastatic tumours demonstrated that Nkx2-1 controls tumour differentiation and limits metastatic potential in vivo. Interrogation of Nkx2-1 regulated genes, analysis of tumours at defined developmental stages, and functional complementation experiments indicate that Nkx2-1 constrains tumours in part by repressing the embryonically-restricted chromatin regulator Hmga2. While focal amplification of NKX2-1 in a fraction of human lung adenocarcinomas has focused attention on its oncogenic function6-9, our data specifically link Nkx2-1 downregulation to loss of differentiation, enhanced tumour seeding ability, and increased metastatic proclivity. Thus, the oncogenic and suppressive functions of Nkx2-1 in the same tumour type substantiate its role as a dual function lineage factor. 23 cell lines derived from primary tumor or metastasis. 6 samples analyzed to determine the effect of Nkx2-1 knockdown on gene expression

Dataset Information

The homeobox transcription factor Nkx2-1 regulates microRNAs controlling downstream gene silencing in lung epithelial cells (mRNA)

Publications

Transcription factor and microRNA interactions in lung cells: an inhibitory link between NK2 homeobox 1, miR-200c and the developmental and oncogenic factors Nfib and Myb.

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