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Oncogenic Role of miR-15a-3p in 13q Amplicon-Driven Colorectal Adenoma-to-Carcinoma Progression.

de Groen FL, Timmer LM, Menezes RX, Diosdado B, Hooijberg E, Meijer GA, Steenbergen RD, Carvalho B - PLoS ONE (2015)

Bottom Line: A subset of these genes, including the mir-17~92 cluster, are functionally involved in CRC development.These results could be confirmed by qRT-PCR in a series of 100 colon adenomas and carcinomas.Functional analysis of both mature miRNAs encoded by mir-15a, i.e. miR-15a-5p and miR-15a-3p, showed that silencing of miR-15a-3p significantly inhibited viability of CRC cells.Upon silencing of miR-15a-3p, mRNA expression of both genes increased in CRC cells, supporting miR-15a-3p mediated regulation of UPC2 and COPS2 expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, VU University Medical Center, Amsterdam, the Netherlands.

ABSTRACT
Progression from colorectal adenoma to carcinoma is strongly associated with an accumulation of genomic alterations, including gain of chromosome 13. This gain affects the whole q arm and is present in 40%-60% of all colorectal cancers (CRCs). Several genes located at this amplicon are known to be overexpressed in carcinomas due to copy number dosage. A subset of these genes, including the mir-17~92 cluster, are functionally involved in CRC development. The present study set out to explore whether apart from mir-17~92, other miRNAs located at the 13q amplicon show a copy number dependent dosage effect that may contribute to 13q-driven colorectal adenoma-to-carcinoma progression. Integration of publically available miRNA expression, target mRNA expression and DNA copy number data from 125 CRCs yielded three miRNAs, miR-15a, -17, and -20a, of which high expression levels were significantly correlated with a 13q gain and which influenced target mRNA expression. These results could be confirmed by qRT-PCR in a series of 100 colon adenomas and carcinomas.Functional analysis of both mature miRNAs encoded by mir-15a, i.e. miR-15a-5p and miR-15a-3p, showed that silencing of miR-15a-3p significantly inhibited viability of CRC cells. Integration of miR-15a expression levels with mRNA expression data of predicted target genes identified mitochondrial uncoupling protein 2 (UCP2) and COP9 signalosome subunit 2 (COPS2) as candidates with significantly decreased expression in CRCs with 13q gain. Upon silencing of miR-15a-3p, mRNA expression of both genes increased in CRC cells, supporting miR-15a-3p mediated regulation of UPC2 and COPS2 expression. In conclusion, significant overexpression of miR-15a-3p due to gain of 13q is functionally relevant in CRC, with UCP2 and COPS2 as candidate target genes. Taken together our findings suggest that miR-15a-3p may contribute to adenoma-to-carcinoma progression.

No MeSH data available.


Related in: MedlinePlus

Schematic overview of in silico analysis pipeline used in this study.First level (association 1) shows a graphical representation of integration analysis of miRNA expression with copy number (CN) data. CN values for miRNAs on 13q were determined by combining values within a 2 Mb window surrounding the start of the miRNA. These values defined the covariate set used to determine association with miRNA expression. Second level (association 2) gives a graphical representation of the association of miRNA expression with predicted target mRNA expression. For each miRNA on 13q target mRNAs were determined by combining three or more target prediction tools. Expression levels of these target mRNAs defined the covariate set used to determine association with miRNA expression.
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pone.0132495.g001: Schematic overview of in silico analysis pipeline used in this study.First level (association 1) shows a graphical representation of integration analysis of miRNA expression with copy number (CN) data. CN values for miRNAs on 13q were determined by combining values within a 2 Mb window surrounding the start of the miRNA. These values defined the covariate set used to determine association with miRNA expression. Second level (association 2) gives a graphical representation of the association of miRNA expression with predicted target mRNA expression. For each miRNA on 13q target mRNAs were determined by combining three or more target prediction tools. Expression levels of these target mRNAs defined the covariate set used to determine association with miRNA expression.

Mentions: To investigate 13q miRNAs of which expression was regulated by DNA copy number changes, associations between DNA copy number and miRNA expression levels were studied. To this end we used an integrated approach, based on the use of covariate sets rather than single covariates, to determine subtle consistent associations with copy number alterations of a chromosomal region. The covariate set was defined by considering each miRNA probe individually and all copy number measurements within a 2Mb-window around the genomic start position of the miRNA. The analysis pipeline and definition of the covariate set is graphically presented in Fig 1 (association 1). The method used is implemented in the BioConductor package SIM [18]. This procedure finds miRNAs of which expression is likely to be regulated by DNA copy number changes within this 2MB-window, by assigning one p-value to each miRNA. Multiple testing correction was done using the false-discovery rate (FDR) step-down procedure of Benjamini & Hoghberg [19]. This analysis yielded a list of FDR values for miRNAs, of which expression is likely to be regulated by copy number changes in these samples. MiRNAs with FDR<0.05 were considered significant.


Oncogenic Role of miR-15a-3p in 13q Amplicon-Driven Colorectal Adenoma-to-Carcinoma Progression.

de Groen FL, Timmer LM, Menezes RX, Diosdado B, Hooijberg E, Meijer GA, Steenbergen RD, Carvalho B - PLoS ONE (2015)

Schematic overview of in silico analysis pipeline used in this study.First level (association 1) shows a graphical representation of integration analysis of miRNA expression with copy number (CN) data. CN values for miRNAs on 13q were determined by combining values within a 2 Mb window surrounding the start of the miRNA. These values defined the covariate set used to determine association with miRNA expression. Second level (association 2) gives a graphical representation of the association of miRNA expression with predicted target mRNA expression. For each miRNA on 13q target mRNAs were determined by combining three or more target prediction tools. Expression levels of these target mRNAs defined the covariate set used to determine association with miRNA expression.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4492490&req=5

pone.0132495.g001: Schematic overview of in silico analysis pipeline used in this study.First level (association 1) shows a graphical representation of integration analysis of miRNA expression with copy number (CN) data. CN values for miRNAs on 13q were determined by combining values within a 2 Mb window surrounding the start of the miRNA. These values defined the covariate set used to determine association with miRNA expression. Second level (association 2) gives a graphical representation of the association of miRNA expression with predicted target mRNA expression. For each miRNA on 13q target mRNAs were determined by combining three or more target prediction tools. Expression levels of these target mRNAs defined the covariate set used to determine association with miRNA expression.
Mentions: To investigate 13q miRNAs of which expression was regulated by DNA copy number changes, associations between DNA copy number and miRNA expression levels were studied. To this end we used an integrated approach, based on the use of covariate sets rather than single covariates, to determine subtle consistent associations with copy number alterations of a chromosomal region. The covariate set was defined by considering each miRNA probe individually and all copy number measurements within a 2Mb-window around the genomic start position of the miRNA. The analysis pipeline and definition of the covariate set is graphically presented in Fig 1 (association 1). The method used is implemented in the BioConductor package SIM [18]. This procedure finds miRNAs of which expression is likely to be regulated by DNA copy number changes within this 2MB-window, by assigning one p-value to each miRNA. Multiple testing correction was done using the false-discovery rate (FDR) step-down procedure of Benjamini & Hoghberg [19]. This analysis yielded a list of FDR values for miRNAs, of which expression is likely to be regulated by copy number changes in these samples. MiRNAs with FDR<0.05 were considered significant.

Bottom Line: A subset of these genes, including the mir-17~92 cluster, are functionally involved in CRC development.These results could be confirmed by qRT-PCR in a series of 100 colon adenomas and carcinomas.Functional analysis of both mature miRNAs encoded by mir-15a, i.e. miR-15a-5p and miR-15a-3p, showed that silencing of miR-15a-3p significantly inhibited viability of CRC cells.Upon silencing of miR-15a-3p, mRNA expression of both genes increased in CRC cells, supporting miR-15a-3p mediated regulation of UPC2 and COPS2 expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, VU University Medical Center, Amsterdam, the Netherlands.

ABSTRACT
Progression from colorectal adenoma to carcinoma is strongly associated with an accumulation of genomic alterations, including gain of chromosome 13. This gain affects the whole q arm and is present in 40%-60% of all colorectal cancers (CRCs). Several genes located at this amplicon are known to be overexpressed in carcinomas due to copy number dosage. A subset of these genes, including the mir-17~92 cluster, are functionally involved in CRC development. The present study set out to explore whether apart from mir-17~92, other miRNAs located at the 13q amplicon show a copy number dependent dosage effect that may contribute to 13q-driven colorectal adenoma-to-carcinoma progression. Integration of publically available miRNA expression, target mRNA expression and DNA copy number data from 125 CRCs yielded three miRNAs, miR-15a, -17, and -20a, of which high expression levels were significantly correlated with a 13q gain and which influenced target mRNA expression. These results could be confirmed by qRT-PCR in a series of 100 colon adenomas and carcinomas.Functional analysis of both mature miRNAs encoded by mir-15a, i.e. miR-15a-5p and miR-15a-3p, showed that silencing of miR-15a-3p significantly inhibited viability of CRC cells. Integration of miR-15a expression levels with mRNA expression data of predicted target genes identified mitochondrial uncoupling protein 2 (UCP2) and COP9 signalosome subunit 2 (COPS2) as candidates with significantly decreased expression in CRCs with 13q gain. Upon silencing of miR-15a-3p, mRNA expression of both genes increased in CRC cells, supporting miR-15a-3p mediated regulation of UPC2 and COPS2 expression. In conclusion, significant overexpression of miR-15a-3p due to gain of 13q is functionally relevant in CRC, with UCP2 and COPS2 as candidate target genes. Taken together our findings suggest that miR-15a-3p may contribute to adenoma-to-carcinoma progression.

No MeSH data available.


Related in: MedlinePlus