Limits...
Targeted Knockdown of the Kinetochore Protein D40/Knl-1 Inhibits Human Cancer in a p53 Status-Independent Manner.

Urata YN, Takeshita F, Tanaka H, Ochiya T, Takimoto M - Sci Rep (2015)

Bottom Line: The D40 gene encodes a kinetochore protein that plays an essential role in kinetochore formation during mitosis.These results indicated that D40 siRNA induced apoptotic cell death in human cancer cell lines, and inhibited their growth in vitro and in vivo regardless of p53 status.Therefore, D40 siRNA is a potential candidate anti-cancer reagent.

View Article: PubMed Central - PubMed

Affiliation: Division of Cancer Gene Regulation, Institute for Genetic Medicine, Hokkaido University, Sapporo, Hokkaido, Japan.

ABSTRACT
The D40 gene encodes a kinetochore protein that plays an essential role in kinetochore formation during mitosis. Short inhibitory RNA against D40, D40 siRNA, has been shown to deplete the D40 protein in the human cancer cell line HeLa, which harbors wild-type p53, and this activity was followed by the significant inhibition of cell growth and induction of apoptotic cell death. The p53- cancer cell line, PC-3M-luc, is also sensitive to the significant growth inhibition and cell death induced by D40 siRNA. The growth of PC-3M-luc tumors transplanted into nude mice was inhibited by the systemic administration of D40 siRNA and the atelocollagen complex. Furthermore, D40 siRNA significantly inhibited growth and induced apoptotic cell death in a cell line with a gain-of-function (GOF) mutation in p53, MDA-MB231-luc, and also inhibited the growth of tumors transplanted into mice when administered as a D40 siRNA/atelocollagen complex. These results indicated that D40 siRNA induced apoptotic cell death in human cancer cell lines, and inhibited their growth in vitro and in vivo regardless of p53 status. Therefore, D40 siRNA is a potential candidate anti-cancer reagent.

No MeSH data available.


Related in: MedlinePlus

Gain-of-Function (GOF) mutant p53 cell line was susceptible to Growth Inhibition and Apoptotic Cell death by D40 siRNA.(a) The GOF mutant p53 cell line MDA-MB231-luc was susceptible to depletion of D40 protein by D40 siRNA. MDA-MB231-luc cells were transfected with D40 or control siRNA, and WB analyses of D40 protein expression were performed 48 hrs after transfection. The gels for D40 and β-actin were run under the same experimental conditions, and then the proteins were transferred to a filter membrane. The membrane was cut and divided into two parts. One was probed with the anti-D40 antibody and the other was probed with the anti-β-actin antibody. (b) D40 siRNA induced Growth Inhibition and Caspase 3/7 activation in the MDA-MB231-luc cell line. Left: Growth curves of the MDA-MB231-Luc cell line. The transfection of siRNAs and the determination of the viable cell number were performed as described in Fig. 2B. Right: Ninety-six hours after transfection, the cells were harvested and caspase 3/7 activities were determined.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4562263&req=5

f4: Gain-of-Function (GOF) mutant p53 cell line was susceptible to Growth Inhibition and Apoptotic Cell death by D40 siRNA.(a) The GOF mutant p53 cell line MDA-MB231-luc was susceptible to depletion of D40 protein by D40 siRNA. MDA-MB231-luc cells were transfected with D40 or control siRNA, and WB analyses of D40 protein expression were performed 48 hrs after transfection. The gels for D40 and β-actin were run under the same experimental conditions, and then the proteins were transferred to a filter membrane. The membrane was cut and divided into two parts. One was probed with the anti-D40 antibody and the other was probed with the anti-β-actin antibody. (b) D40 siRNA induced Growth Inhibition and Caspase 3/7 activation in the MDA-MB231-luc cell line. Left: Growth curves of the MDA-MB231-Luc cell line. The transfection of siRNAs and the determination of the viable cell number were performed as described in Fig. 2B. Right: Ninety-six hours after transfection, the cells were harvested and caspase 3/7 activities were determined.

Mentions: We next determined whether D40 siRNA inhibited the growth of the GOF p53 mutant cell line MDA-MB231-luc, which exhibits malignant phenotypes such as a high potentials for invasiveness and metastasis as well as resistance to therapeutics. As such, it was important to assess the sensitivity of this cell line to D40 siRNA. The results revealed that D40 siRNA could significantly reduce D40 protein expression (Fig. 4a). The growth of cells transfected with D40 siRNA was also inhibited compared with that of cells transfected with control siRNA (Fig. 4b Left). Furthermore, caspase 3/7 activity was higher in cells transfected with D40 siRNA than in those transfected with control siRNA (Fig. 4b Right). These results indicated that D40 siRNA effectively induced apoptosis in tumors with a GOF mutation in p53.


Targeted Knockdown of the Kinetochore Protein D40/Knl-1 Inhibits Human Cancer in a p53 Status-Independent Manner.

Urata YN, Takeshita F, Tanaka H, Ochiya T, Takimoto M - Sci Rep (2015)

Gain-of-Function (GOF) mutant p53 cell line was susceptible to Growth Inhibition and Apoptotic Cell death by D40 siRNA.(a) The GOF mutant p53 cell line MDA-MB231-luc was susceptible to depletion of D40 protein by D40 siRNA. MDA-MB231-luc cells were transfected with D40 or control siRNA, and WB analyses of D40 protein expression were performed 48 hrs after transfection. The gels for D40 and β-actin were run under the same experimental conditions, and then the proteins were transferred to a filter membrane. The membrane was cut and divided into two parts. One was probed with the anti-D40 antibody and the other was probed with the anti-β-actin antibody. (b) D40 siRNA induced Growth Inhibition and Caspase 3/7 activation in the MDA-MB231-luc cell line. Left: Growth curves of the MDA-MB231-Luc cell line. The transfection of siRNAs and the determination of the viable cell number were performed as described in Fig. 2B. Right: Ninety-six hours after transfection, the cells were harvested and caspase 3/7 activities were determined.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4562263&req=5

f4: Gain-of-Function (GOF) mutant p53 cell line was susceptible to Growth Inhibition and Apoptotic Cell death by D40 siRNA.(a) The GOF mutant p53 cell line MDA-MB231-luc was susceptible to depletion of D40 protein by D40 siRNA. MDA-MB231-luc cells were transfected with D40 or control siRNA, and WB analyses of D40 protein expression were performed 48 hrs after transfection. The gels for D40 and β-actin were run under the same experimental conditions, and then the proteins were transferred to a filter membrane. The membrane was cut and divided into two parts. One was probed with the anti-D40 antibody and the other was probed with the anti-β-actin antibody. (b) D40 siRNA induced Growth Inhibition and Caspase 3/7 activation in the MDA-MB231-luc cell line. Left: Growth curves of the MDA-MB231-Luc cell line. The transfection of siRNAs and the determination of the viable cell number were performed as described in Fig. 2B. Right: Ninety-six hours after transfection, the cells were harvested and caspase 3/7 activities were determined.
Mentions: We next determined whether D40 siRNA inhibited the growth of the GOF p53 mutant cell line MDA-MB231-luc, which exhibits malignant phenotypes such as a high potentials for invasiveness and metastasis as well as resistance to therapeutics. As such, it was important to assess the sensitivity of this cell line to D40 siRNA. The results revealed that D40 siRNA could significantly reduce D40 protein expression (Fig. 4a). The growth of cells transfected with D40 siRNA was also inhibited compared with that of cells transfected with control siRNA (Fig. 4b Left). Furthermore, caspase 3/7 activity was higher in cells transfected with D40 siRNA than in those transfected with control siRNA (Fig. 4b Right). These results indicated that D40 siRNA effectively induced apoptosis in tumors with a GOF mutation in p53.

Bottom Line: The D40 gene encodes a kinetochore protein that plays an essential role in kinetochore formation during mitosis.These results indicated that D40 siRNA induced apoptotic cell death in human cancer cell lines, and inhibited their growth in vitro and in vivo regardless of p53 status.Therefore, D40 siRNA is a potential candidate anti-cancer reagent.

View Article: PubMed Central - PubMed

Affiliation: Division of Cancer Gene Regulation, Institute for Genetic Medicine, Hokkaido University, Sapporo, Hokkaido, Japan.

ABSTRACT
The D40 gene encodes a kinetochore protein that plays an essential role in kinetochore formation during mitosis. Short inhibitory RNA against D40, D40 siRNA, has been shown to deplete the D40 protein in the human cancer cell line HeLa, which harbors wild-type p53, and this activity was followed by the significant inhibition of cell growth and induction of apoptotic cell death. The p53- cancer cell line, PC-3M-luc, is also sensitive to the significant growth inhibition and cell death induced by D40 siRNA. The growth of PC-3M-luc tumors transplanted into nude mice was inhibited by the systemic administration of D40 siRNA and the atelocollagen complex. Furthermore, D40 siRNA significantly inhibited growth and induced apoptotic cell death in a cell line with a gain-of-function (GOF) mutation in p53, MDA-MB231-luc, and also inhibited the growth of tumors transplanted into mice when administered as a D40 siRNA/atelocollagen complex. These results indicated that D40 siRNA induced apoptotic cell death in human cancer cell lines, and inhibited their growth in vitro and in vivo regardless of p53 status. Therefore, D40 siRNA is a potential candidate anti-cancer reagent.

No MeSH data available.


Related in: MedlinePlus