Short-term transcriptome and microRNAs responses to exposure to different air pollutants in two population studies.

Diesel vehicle emissions are the major source of genotoxic compounds in ambient air from urban areas. These pollutants are linked to risks of cardiovascular diseases, lung cancer, respiratory infections and adverse neurological effects. Biological events associated with exposure to some air pollutants are widely unknown but applying omics techniques may help to identify the molecular processes that link exposure to disease risk. Most data on health risks are related to long-term exposure, so the aim of this study is to investigate the impact of short-term exposure (two hours) to air pollutants on the blood transcriptome and microRNA expression levels. We analyzed transcriptomics and microRNA expression using microarray technology on blood samples from volunteers participating in studies in London, the Oxford Street cohort, and, in Barcelona, the TAPAS cohort. Personal exposure levels measurements of particulate matter (PM10, PM2.5), ultrafine particles (UFPC), nitrogen oxides (NO2, NO and NOx), black carbon (BC) and carbon oxides (CO and CO2) were registered for each volunteer. Associations between air pollutant levels and gene/microRNA expression were evaluated using multivariate normal models (MVN). MVN-models identified compound-specific expression of blood cell genes and microRNAs associated with air pollution despite the low exposure levels, the short exposure periods and the relatively small-sized cohorts. Hsa-miR-197-3p, hsa-miR-29a-3p, hsa-miR-15a-5p, hsa-miR-16-5p and hsa-miR-92a-3p are found significantly expressed in association with exposures. These microRNAs target also relevant transcripts, indicating their potential relevance in the research of omics-biomarkers responding to air pollution. Furthermore, these microRNAs are also known to be associated with diseases previously linked to air pollution exposure including several cancers such lung cancer and Alzheimer's disease. In conclusion, we identified in this study promising compound-specific mRNA and microRNA biomarkers after two hours of exposure to low levels of air pollutants during two hours that suggest increased cancer risks

Experimental Validation of Gene Expression of MYBL1, MYBL2, UBXN8, and ADRM1 Genes in Triple Negative Breast Cancer Cell Lines

A previous study conducted in our laboratory demonstrated V-Myb Avian Myeloblast Viral Oncogene Homolog Like 1 (MYBL1) gene over-expression in triple negative breast cancer (TNBC) compared to normal, some luminal, and a subpopulation of other TNBC. The MYBL1 gene belongs to the Avian myeloblastosis virus (MYB) family and is classified as proto-oncogene that functions as a strong transcription factor. The MYBL1 gene is related to cancer progression which involves dysregulation of cell cycle signaling, apoptosis and differentiation processes. A primary goal of our laboratory is to further characterize MYBL1 gene expression in TNBC samples. To achieve this goal, we performed a knockdown study to identify genes that co-operate with MYBL1 to affect the phenotype of TNBC. The MDA MB231 TNBC cells were ransduced with a short hairpin ribonucleic acid (shRNA) lentiviral knockdown of the MYBL1 gene. When MYBL1 was knocked down, MYBL2 and Adhesion Regulating Molecule 1 (ADRM1) genes were down regulated and UBX Domain Protein 8 (UBXN8) gene was unregulated. Since MYBL2, UBXN8 and ADRM1 were affected by MYBL1 knockdown, for the current study, we compared the gene expression patterns of MYBL2, UBXN8 and ADRM1 to that of MYBL1 using different methods. Two approaches are utilized to achieve our goal. For approach 1 we utilized polymerase chain reaction and immunohistochemistry to assess RNA and protein expression levels, respectively. For the second approach, we analyzed MYBL1, MYBL2, UBXN8 and ADRM1 transcript levels in TNBC patient samples etrieved from Gene Expression Omnibus Results from this project should assist in our understanding of MYBL1 in TNBC

V-Myb Avian Myel oblast Viral Oncogene Homolog Like 1 (MYBL1) Knockdown and its Role in a Triple Negative Breast Cancer

Triple Negative Breast Cancer (TNBC) is defined as negative for three genes, estrogen receptor (ESR), progesterone receptor (PR) and Human epidermal growth factor receptor (HER2-neu) genes. Previous data show the V-Myb Avian Myel oblast Viral Oncogene Homolog Like 1 (MYBL1) gene is over-expressed in Triple negative breast cancer cell line (MDA-MB231). MYBL1 belongs to the MYB family of genes which are transcription factors and proto-oncogenes which are associated with cell cycle regulation, apoptosis, and differentiation, all of which are key events associated with cancers. It could be that MYBL1 contributes to these same processes in TNBC. Instead of studying MYBL1’s contribution to several of these processes, we were mainly concerned with identifying genes that were either directly or indirectly affected by down-regulation of MYBL1 gene. Utilizing a gene silencing approach helps to identify genes that cooperate with MYBL1 in the signaling processes in cancer. Although the focus of our laboratory is TNBC, there are two parts to this current study, one that examines MYBL1 in luminal cancers cell line (MCF7) and one that examines MYBL1 in TNBC, designated Part 1 and Part 2, respectively. For Part 1, we performed analyses of MCF7 (Luminal breast cancer cell line) receptor positive cells where estrogen receptor gene was silenced; and another MCF7 preparation where cMYB gene was silenced. Both datasets were obtained from Gene Expression Omnibus (GEO). These datasets were chosen because even though they were neither TNBC or directly involved MYBL1 as the primary target, comparative analyses of both datasets showed MYBL1 knock-down (KD). We reasoned that even under these conditions, genes either directly or indirectly associated with MYBL1 might be identified. For Part 2 of this study, short hairpin RNA (shRNA) lentiviral transduction was used to down-regulate MYBL1 in MDA MB231 TNBC cells. A substantial number of reliable differentially expressed genes were identified here. Overall, genes recognized as associated with MYBL1 in the MCF7 luminal preparations (Part 1) were drastically different from genes identified as associated with MYBL1 in the TNBC KD study (Part 2). In both datasets, we identify novel genes that appear to be coordinately expressed with MYBL1 in breast cancers. This study led to identification of candidate genes that might be important towards the study of characterizing MYBL1 expression in TNBC. Two of these genes, transcription factor 19 (TCF19) and Kinesin-like protein (kinesin family member 18B) (KIF18B) have been experimentally validated. MYBL1 is a strong candidate gene to study for its contribution to the development of TNBC. Continued analyses of these genes and their relationship to MYBL1 should lead to a better understanding of signaling processes in breast cancers

A Supervised Network Analysis on Gene Expression Profiles of Breast Tumors Predicts a 41-Gene Prognostic Signature of the Transcription Factor MYB

Background. MYB is predicted to be a favorable prognostic predictor in a breast cancer population. We proposed to find the inferred mechanism(s) relevant to the prognostic features of MYB via a supervised network analysis. Methods. Both coefficient of intrinsic dependence (CID) and Galton Pierson’s correlation coefficient (GPCC) were combined and designated as CIDUGPCC. It is for the univariate network analysis. Multivariate CID is for the multivariate network analysis. Other analyses using bioinformatic tools and statistical methods are included. Results. ARNT2 is predicted to be the essential gene partner of MYB. We classified four prognostic relevant gene subpools in three breast cancer cohorts as feature types I–IV. Only the probes in feature type II are the potential prognostic feature of MYB. Moreover, we further validated 41 prognosis relevant probes to be the favorable prognostic signature. Surprisingly, two additional family members of MYB are elevated to promote poor prognosis when both levels of MYB and ARNT2 decline. Both MYBL1 and MYBL2 may partially decrease the tumor suppressive activities that are predicted to be up-regulated by MYB and ARNT2. Conclusions. The major prognostic feature of MYB is predicted to be determined by the MYB subnetwork (41 probes) that is relevant across subtypes

A Supervised Network Analysis on Gene Expression Profiles of Breast Tumors Predicts a 41-Gene Prognostic Signature of the Transcription Factor MYB across Molecular Subtypes

Background. MYB is predicted to be a favorable prognostic predictor in a breast cancer population. We proposed to find the inferred mechanism(s) relevant to the prognostic features of MYB via a supervised network analysis. Methods. Both coefficient of intrinsic dependence (CID) and Galton Pierson's correlation coefficient (GPCC) were combined and designated as CIDUGPCC. It is for the univariate network analysis. Multivariate CID is for the multivariate network analysis. Other analyses using bioinformatic tools and statistical methods are included. Results. ARNT2 is predicted to be the essential gene partner of MYB. We classified four prognostic relevant gene subpools in three breast cancer cohorts as feature types I-IV. Only the probes in feature type II are the potential prognostic feature of MYB. Moreover, we further validated 41 prognosis relevant probes to be the favorable prognostic signature. Surprisingly, two additional family members of MYB are elevated to promote poor prognosis when both levels of MYB and ARNT2 decline. Both MYBL1 and MYBL2 may partially decrease the tumor suppressive activities that are predicted to be up-regulated by MYB and ARNT2. Conclusions. The major prognostic feature of MYB is predicted to be determined by the MYB subnetwork (41 probes) that is relevant across subtypes

Microscopic analysis of aneuploidy induced by the mutation of the CCDC124 gene

The Coiled-coil domain containing protein 124 (Ccdc124) is a centrosomal protein that relocates to the midbody region at the cytokinesis stage of the cell cycle. Cytokinetic abscission is the cellular process that leads to physical separation of two postmitotic sister cells by severing the intercellular bridge. Mutation of the Ccdc124 gene by CRISPR/Cas9 genome editing in HEK293T cells leads to the failure of cytokinesis and formation of aneuploid (multinucleated-MN) aberrant cells. In this study, the MN cells were analyzed using flow cytometry and confocal imaging followed by quantitative image analysis. MN cells had mitotic and chromosome attachment aberrations, multiple centrosomes and micronuclei. These aberrations are known to occur in tumour cells, a finding that links Ccdc124 to cancer. MN cells also upregulated the p53 protein, which induced senescence. Furthermore, MN cells had increased numbers of 53BP1 foci which indicates that the mutation of Ccdc124 induces the DNA damage response and activates the p53 pathway. This study documents a relationship between Ccdc124 mutation-associated cytokinesis failure and p53-dependent senescence