A del T poly T (8) mutation in the 3′ untranslated region (UTR) of the CDK2-AP1 gene is functionally significant causing decreased mRNA stability resulting in decreased CDK2-AP1 expression in human microsatellite unstable (MSI) colorectal cancer (CRC)

Joongho Shin, Ziqian Yuan, Kenneth Fordyce, Prashanth V. Sreeramoju, Tara S. Kent, John Kim, Victor Wang, David Schneyer, Thomas K. Weber

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Background: We have previously published results indicating that decreased expression of CDK2-AP1 in MSI human colorectal cancer is associated with deletion mutations in the poly (T) 8 repeat sequence within the 3′-UTR of the CDK2-AP1 gene. In this study, we test the hypothesis that the del T mutation results in decreased CDK2-AP1 expression by causing reduced mRNA stability. Methods: We introduced wild-type and mutant 3′-UTR sequences fused to a green fluorescent protein (GFP) gene separately into human CRC cell lines and quantified the expression of the GFP gene. Native CDK2-AP1 mRNA stability was measured in human CRC cell lines, using an actinomycin D assay and the mRNA structure folding software mfold 3.2. Results: Mutant GFP-3′-UTR samples demonstrated significantly reduced GFP expression compared with wild-type GFP-3′-UTR as measured by both FACS and real-time PCR. Both the actinomycin D assay and mfold software demonstrated significantly reduced mRNA stability for the del T poly (T) 8 transcript compared with the wild type. Conclusions: In summary, these novel results support our hypotheses that the del T poly (T) 8 observed in the 3′-UTR of the CDK2-AP1 gene in human MSI CRC is functionally significant and results in decreased CDK2-AP1 expression. The results also indicate the mechanism of this decreased expression is caused at least in part by decreased mRNA stability.

Original languageEnglish (US)
Pages (from-to)222-227
Number of pages6
JournalSurgery
Volume142
Issue number2
DOIs
StatePublished - Aug 2007

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Poly T
RNA Stability
3' Untranslated Regions
Green Fluorescent Proteins
Microsatellite Repeats
Colorectal Neoplasms
Mutation
Genes
Dactinomycin
Software
Cell Line
Sequence Deletion
Mutant Proteins
Real-Time Polymerase Chain Reaction
Messenger RNA

ASJC Scopus subject areas

  • Surgery

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A del T poly T (8) mutation in the 3′ untranslated region (UTR) of the CDK2-AP1 gene is functionally significant causing decreased mRNA stability resulting in decreased CDK2-AP1 expression in human microsatellite unstable (MSI) colorectal cancer (CRC). / Shin, Joongho; Yuan, Ziqian; Fordyce, Kenneth; Sreeramoju, Prashanth V.; Kent, Tara S.; Kim, John; Wang, Victor; Schneyer, David; Weber, Thomas K.

In: Surgery, Vol. 142, No. 2, 08.2007, p. 222-227.

Research output: Contribution to journalArticle

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title = "A del T poly T (8) mutation in the 3′ untranslated region (UTR) of the CDK2-AP1 gene is functionally significant causing decreased mRNA stability resulting in decreased CDK2-AP1 expression in human microsatellite unstable (MSI) colorectal cancer (CRC)",
abstract = "Background: We have previously published results indicating that decreased expression of CDK2-AP1 in MSI human colorectal cancer is associated with deletion mutations in the poly (T) 8 repeat sequence within the 3′-UTR of the CDK2-AP1 gene. In this study, we test the hypothesis that the del T mutation results in decreased CDK2-AP1 expression by causing reduced mRNA stability. Methods: We introduced wild-type and mutant 3′-UTR sequences fused to a green fluorescent protein (GFP) gene separately into human CRC cell lines and quantified the expression of the GFP gene. Native CDK2-AP1 mRNA stability was measured in human CRC cell lines, using an actinomycin D assay and the mRNA structure folding software mfold 3.2. Results: Mutant GFP-3′-UTR samples demonstrated significantly reduced GFP expression compared with wild-type GFP-3′-UTR as measured by both FACS and real-time PCR. Both the actinomycin D assay and mfold software demonstrated significantly reduced mRNA stability for the del T poly (T) 8 transcript compared with the wild type. Conclusions: In summary, these novel results support our hypotheses that the del T poly (T) 8 observed in the 3′-UTR of the CDK2-AP1 gene in human MSI CRC is functionally significant and results in decreased CDK2-AP1 expression. The results also indicate the mechanism of this decreased expression is caused at least in part by decreased mRNA stability.",
author = "Joongho Shin and Ziqian Yuan and Kenneth Fordyce and Sreeramoju, {Prashanth V.} and Kent, {Tara S.} and John Kim and Victor Wang and David Schneyer and Weber, {Thomas K.}",
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T1 - A del T poly T (8) mutation in the 3′ untranslated region (UTR) of the CDK2-AP1 gene is functionally significant causing decreased mRNA stability resulting in decreased CDK2-AP1 expression in human microsatellite unstable (MSI) colorectal cancer (CRC)

AU - Shin, Joongho

AU - Yuan, Ziqian

AU - Fordyce, Kenneth

AU - Sreeramoju, Prashanth V.

AU - Kent, Tara S.

AU - Kim, John

AU - Wang, Victor

AU - Schneyer, David

AU - Weber, Thomas K.

PY - 2007/8

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N2 - Background: We have previously published results indicating that decreased expression of CDK2-AP1 in MSI human colorectal cancer is associated with deletion mutations in the poly (T) 8 repeat sequence within the 3′-UTR of the CDK2-AP1 gene. In this study, we test the hypothesis that the del T mutation results in decreased CDK2-AP1 expression by causing reduced mRNA stability. Methods: We introduced wild-type and mutant 3′-UTR sequences fused to a green fluorescent protein (GFP) gene separately into human CRC cell lines and quantified the expression of the GFP gene. Native CDK2-AP1 mRNA stability was measured in human CRC cell lines, using an actinomycin D assay and the mRNA structure folding software mfold 3.2. Results: Mutant GFP-3′-UTR samples demonstrated significantly reduced GFP expression compared with wild-type GFP-3′-UTR as measured by both FACS and real-time PCR. Both the actinomycin D assay and mfold software demonstrated significantly reduced mRNA stability for the del T poly (T) 8 transcript compared with the wild type. Conclusions: In summary, these novel results support our hypotheses that the del T poly (T) 8 observed in the 3′-UTR of the CDK2-AP1 gene in human MSI CRC is functionally significant and results in decreased CDK2-AP1 expression. The results also indicate the mechanism of this decreased expression is caused at least in part by decreased mRNA stability.

AB - Background: We have previously published results indicating that decreased expression of CDK2-AP1 in MSI human colorectal cancer is associated with deletion mutations in the poly (T) 8 repeat sequence within the 3′-UTR of the CDK2-AP1 gene. In this study, we test the hypothesis that the del T mutation results in decreased CDK2-AP1 expression by causing reduced mRNA stability. Methods: We introduced wild-type and mutant 3′-UTR sequences fused to a green fluorescent protein (GFP) gene separately into human CRC cell lines and quantified the expression of the GFP gene. Native CDK2-AP1 mRNA stability was measured in human CRC cell lines, using an actinomycin D assay and the mRNA structure folding software mfold 3.2. Results: Mutant GFP-3′-UTR samples demonstrated significantly reduced GFP expression compared with wild-type GFP-3′-UTR as measured by both FACS and real-time PCR. Both the actinomycin D assay and mfold software demonstrated significantly reduced mRNA stability for the del T poly (T) 8 transcript compared with the wild type. Conclusions: In summary, these novel results support our hypotheses that the del T poly (T) 8 observed in the 3′-UTR of the CDK2-AP1 gene in human MSI CRC is functionally significant and results in decreased CDK2-AP1 expression. The results also indicate the mechanism of this decreased expression is caused at least in part by decreased mRNA stability.

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