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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

Analysis of miRNA expression in colorectal adenomas and carcinomas.A) Box plots comparing relative expression determined by qRT-PCR in adenomas (n = 48) versus carcinomas (n = 52) of four top candidate miRNAs, miR-15a-3p, miR-15a-5p, miR-17 and miR-20a. B) Box plots comparing miRNA expression between tumours (adenomas and carcinomas) with (n = 24) or without (n = 73) 13q gain. P-values, determined by Mann-Whitney U-test, ≤ 0.05 are considered significant (* p ≤ 0.05; ** p < 0.01).
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pone.0132495.g002: Analysis of miRNA expression in colorectal adenomas and carcinomas.A) Box plots comparing relative expression determined by qRT-PCR in adenomas (n = 48) versus carcinomas (n = 52) of four top candidate miRNAs, miR-15a-3p, miR-15a-5p, miR-17 and miR-20a. B) Box plots comparing miRNA expression between tumours (adenomas and carcinomas) with (n = 24) or without (n = 73) 13q gain. P-values, determined by Mann-Whitney U-test, ≤ 0.05 are considered significant (* p ≤ 0.05; ** p < 0.01).

Mentions: Next we analysed by qRT-PCR the expression levels of the top candidate miRNAs found in the in silico analyses in a series of 100 colon adenoma and carcinoma samples. Given the fact that mir-15a encodes for two mature miRNA strands, miR-15a-5p and miR-15a-3p, both were studied using strand-specific qRT-PCR. In line with the in silico data, expression levels of miR-15a-3p, miR-17 and miR-20a, were significantly higher in carcinomas compared to adenomas (Fig 2A). However, expression of miR-15a-5p was not significantly increased. Comparison of tumours with 13q gain with tumours without 13q gain showed that expression levels of all four miRNAs were significantly higher in tumours with gain (Fig 2B), corroborating the results from the in silico analysis.


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)

Analysis of miRNA expression in colorectal adenomas and carcinomas.A) Box plots comparing relative expression determined by qRT-PCR in adenomas (n = 48) versus carcinomas (n = 52) of four top candidate miRNAs, miR-15a-3p, miR-15a-5p, miR-17 and miR-20a. B) Box plots comparing miRNA expression between tumours (adenomas and carcinomas) with (n = 24) or without (n = 73) 13q gain. P-values, determined by Mann-Whitney U-test, ≤ 0.05 are considered significant (* p ≤ 0.05; ** p < 0.01).
© Copyright Policy
Related In: Results  -  Collection

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

pone.0132495.g002: Analysis of miRNA expression in colorectal adenomas and carcinomas.A) Box plots comparing relative expression determined by qRT-PCR in adenomas (n = 48) versus carcinomas (n = 52) of four top candidate miRNAs, miR-15a-3p, miR-15a-5p, miR-17 and miR-20a. B) Box plots comparing miRNA expression between tumours (adenomas and carcinomas) with (n = 24) or without (n = 73) 13q gain. P-values, determined by Mann-Whitney U-test, ≤ 0.05 are considered significant (* p ≤ 0.05; ** p < 0.01).
Mentions: Next we analysed by qRT-PCR the expression levels of the top candidate miRNAs found in the in silico analyses in a series of 100 colon adenoma and carcinoma samples. Given the fact that mir-15a encodes for two mature miRNA strands, miR-15a-5p and miR-15a-3p, both were studied using strand-specific qRT-PCR. In line with the in silico data, expression levels of miR-15a-3p, miR-17 and miR-20a, were significantly higher in carcinomas compared to adenomas (Fig 2A). However, expression of miR-15a-5p was not significantly increased. Comparison of tumours with 13q gain with tumours without 13q gain showed that expression levels of all four miRNAs were significantly higher in tumours with gain (Fig 2B), corroborating the results from the in silico analysis.

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