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Kinase-independent role of cyclin D1 in chromosomal instability and mammary tumorigenesis.

Casimiro MC, Di Sante G, Crosariol M, Loro E, Dampier W, Ertel A, Yu Z, Saria EA, Papanikolaou A, Li Z, Wang C, Addya S, Lisanti MP, Fortina P, Cardiff RD, Tozeren A, Knudsen ES, Arnold A, Pestell RG - Oncotarget (2015)

Bottom Line: Sustained expression of cyclin D1(KE) induced mammary adenocarcinoma with similar kinetics to that of the wild-type cyclin D1.ChIP-Seq studies demonstrated recruitment of cyclin D1(WT) and cyclin D1(KE) to the genes governing CIN.We conclude that the CDK-activating function of cyclin D1 is not necessary to induce either chromosomal instability or mammary tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Departments of Cancer Biology, Thomas Jefferson University & Hospital, Philadelphia, PA 19107, USA.

ABSTRACT
Cyclin D1 is an important molecular driver of human breast cancer but better understanding of its oncogenic mechanisms is needed, especially to enhance efforts in targeted therapeutics. Currently, pharmaceutical initiatives to inhibit cyclin D1 are focused on the catalytic component since the transforming capacity is thought to reside in the cyclin D1/CDK activity. We initiated the following study to directly test the oncogenic potential of catalytically inactive cyclin D1 in an in vivo mouse model that is relevant to breast cancer. Herein, transduction of cyclin D1(-/-) mouse embryonic fibroblasts (MEFs) with the kinase dead KE mutant of cyclin D1 led to aneuploidy, abnormalities in mitotic spindle formation, autosome amplification, and chromosomal instability (CIN) by gene expression profiling. Acute transgenic expression of either cyclin D1(WT) or cyclin D1(KE) in the mammary gland was sufficient to induce a high CIN score within 7 days. Sustained expression of cyclin D1(KE) induced mammary adenocarcinoma with similar kinetics to that of the wild-type cyclin D1. ChIP-Seq studies demonstrated recruitment of cyclin D1(WT) and cyclin D1(KE) to the genes governing CIN. We conclude that the CDK-activating function of cyclin D1 is not necessary to induce either chromosomal instability or mammary tumorigenesis.

No MeSH data available.


Related in: MedlinePlus

Identification of transcription factor motifs found in cyclin D1WT and cyclin D1KE interval sequences(A) Selection of transcription factor motif hits common between Cyclin D1WT and cyclin D1KE interval sequences (B) Representative TF motifs found in the interval regions associated with cyclin D1WT and cyclin D1KE(C) Luciferase reporter gene assays were conducted using the Peroxisome Proliferator-Activated Receptor γ (AOX-LUC) (left panel) and Hypoxia Responsive Element (HRE-LUC) (right panel) luciferase reporter constructs. The number of responsive elements for each construct is depicted in the reporter schematic. HEK293T cells were co-transfected with cyclin D1 (50 ng). Data are of n = 2 separate experiments, mean ± SEM.
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Figure 6: Identification of transcription factor motifs found in cyclin D1WT and cyclin D1KE interval sequences(A) Selection of transcription factor motif hits common between Cyclin D1WT and cyclin D1KE interval sequences (B) Representative TF motifs found in the interval regions associated with cyclin D1WT and cyclin D1KE(C) Luciferase reporter gene assays were conducted using the Peroxisome Proliferator-Activated Receptor γ (AOX-LUC) (left panel) and Hypoxia Responsive Element (HRE-LUC) (right panel) luciferase reporter constructs. The number of responsive elements for each construct is depicted in the reporter schematic. HEK293T cells were co-transfected with cyclin D1 (50 ng). Data are of n = 2 separate experiments, mean ± SEM.

Mentions: Select CIN associated genes showed similar ChIP-Seq tag density profiles for cyclin D1WT and cyclin D1KE (Figure 4D). ChIP analysis of selected target genes governing CIN demonstrated similar relative occupancy for cyclin D1WT and cyclin D1KE (Figure 5A). We then analyzed a broader array of genes governing CIN by QT-PCR, demonstrating similar upregulation of the transcript level by cyclin D1WT and cyclin D1KE (Figure 5B). The enrichment for transcription factor (TF) binding sites identified TF motifs and their statistical significance for the cyclin D1WT and cyclin D1KE (Supplementary Figure 6A and Supplementary Table S3). For the examples shown the prevalence of the TF binding site was similar and significant for both cyclin D1WT and cyclin D1KE. Representative TF motifs most significantly enriched in the cyclin D1WT intervals are shown for the cyclin D1KE intervals (Figure 6B). In addition to associating with the TF motifs, we verified that cyclin D1KE regulated the reporter activity of selected TF responsive elements in a similar manner to cyclin D1WT (Figure 6C).


Kinase-independent role of cyclin D1 in chromosomal instability and mammary tumorigenesis.

Casimiro MC, Di Sante G, Crosariol M, Loro E, Dampier W, Ertel A, Yu Z, Saria EA, Papanikolaou A, Li Z, Wang C, Addya S, Lisanti MP, Fortina P, Cardiff RD, Tozeren A, Knudsen ES, Arnold A, Pestell RG - Oncotarget (2015)

Identification of transcription factor motifs found in cyclin D1WT and cyclin D1KE interval sequences(A) Selection of transcription factor motif hits common between Cyclin D1WT and cyclin D1KE interval sequences (B) Representative TF motifs found in the interval regions associated with cyclin D1WT and cyclin D1KE(C) Luciferase reporter gene assays were conducted using the Peroxisome Proliferator-Activated Receptor γ (AOX-LUC) (left panel) and Hypoxia Responsive Element (HRE-LUC) (right panel) luciferase reporter constructs. The number of responsive elements for each construct is depicted in the reporter schematic. HEK293T cells were co-transfected with cyclin D1 (50 ng). Data are of n = 2 separate experiments, mean ± SEM.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4496164&req=5

Figure 6: Identification of transcription factor motifs found in cyclin D1WT and cyclin D1KE interval sequences(A) Selection of transcription factor motif hits common between Cyclin D1WT and cyclin D1KE interval sequences (B) Representative TF motifs found in the interval regions associated with cyclin D1WT and cyclin D1KE(C) Luciferase reporter gene assays were conducted using the Peroxisome Proliferator-Activated Receptor γ (AOX-LUC) (left panel) and Hypoxia Responsive Element (HRE-LUC) (right panel) luciferase reporter constructs. The number of responsive elements for each construct is depicted in the reporter schematic. HEK293T cells were co-transfected with cyclin D1 (50 ng). Data are of n = 2 separate experiments, mean ± SEM.
Mentions: Select CIN associated genes showed similar ChIP-Seq tag density profiles for cyclin D1WT and cyclin D1KE (Figure 4D). ChIP analysis of selected target genes governing CIN demonstrated similar relative occupancy for cyclin D1WT and cyclin D1KE (Figure 5A). We then analyzed a broader array of genes governing CIN by QT-PCR, demonstrating similar upregulation of the transcript level by cyclin D1WT and cyclin D1KE (Figure 5B). The enrichment for transcription factor (TF) binding sites identified TF motifs and their statistical significance for the cyclin D1WT and cyclin D1KE (Supplementary Figure 6A and Supplementary Table S3). For the examples shown the prevalence of the TF binding site was similar and significant for both cyclin D1WT and cyclin D1KE. Representative TF motifs most significantly enriched in the cyclin D1WT intervals are shown for the cyclin D1KE intervals (Figure 6B). In addition to associating with the TF motifs, we verified that cyclin D1KE regulated the reporter activity of selected TF responsive elements in a similar manner to cyclin D1WT (Figure 6C).

Bottom Line: Sustained expression of cyclin D1(KE) induced mammary adenocarcinoma with similar kinetics to that of the wild-type cyclin D1.ChIP-Seq studies demonstrated recruitment of cyclin D1(WT) and cyclin D1(KE) to the genes governing CIN.We conclude that the CDK-activating function of cyclin D1 is not necessary to induce either chromosomal instability or mammary tumorigenesis.

View Article: PubMed Central - PubMed

Affiliation: Departments of Cancer Biology, Thomas Jefferson University & Hospital, Philadelphia, PA 19107, USA.

ABSTRACT
Cyclin D1 is an important molecular driver of human breast cancer but better understanding of its oncogenic mechanisms is needed, especially to enhance efforts in targeted therapeutics. Currently, pharmaceutical initiatives to inhibit cyclin D1 are focused on the catalytic component since the transforming capacity is thought to reside in the cyclin D1/CDK activity. We initiated the following study to directly test the oncogenic potential of catalytically inactive cyclin D1 in an in vivo mouse model that is relevant to breast cancer. Herein, transduction of cyclin D1(-/-) mouse embryonic fibroblasts (MEFs) with the kinase dead KE mutant of cyclin D1 led to aneuploidy, abnormalities in mitotic spindle formation, autosome amplification, and chromosomal instability (CIN) by gene expression profiling. Acute transgenic expression of either cyclin D1(WT) or cyclin D1(KE) in the mammary gland was sufficient to induce a high CIN score within 7 days. Sustained expression of cyclin D1(KE) induced mammary adenocarcinoma with similar kinetics to that of the wild-type cyclin D1. ChIP-Seq studies demonstrated recruitment of cyclin D1(WT) and cyclin D1(KE) to the genes governing CIN. We conclude that the CDK-activating function of cyclin D1 is not necessary to induce either chromosomal instability or mammary tumorigenesis.

No MeSH data available.


Related in: MedlinePlus