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CSE1L interaction with MSH6 promotes osteosarcoma progression and predicts poor patient survival

View Article: PubMed Central - PubMed

ABSTRACT

To discover tumor-associated proteins in osteosarcoma, a quantitative proteomic analysis was performed to identify proteins that were differentially expressed between osteosarcoma and human osteoblastic cells. Through clinical screening and a functional evaluation, chromosome segregation 1-like (CSE1L) protein was found to be related to the growth of osteosarcoma cells. To date, little is known about the function and underlying mechanism of CSE1L in osteosarcoma. In the present study, we show that knockdown of CSE1L inhibits osteosarcoma growth in vitro and in vivo. By co-immunoprecipitation and RNA-seq analysis, CSE1L was found to interact with mutS homolog 6 (MSH6) and function as a positive regulator of MSH6 protein in osteosarcoma cells. A rescue study showed that decreased growth of osteosarcoma cells by CSE1L knockdown was reversed by MSH6 overexpression, indicating that the activity of CSE1L was an MSH6-dependent function. In addition, depletion of MSH6 hindered cellular proliferation in vitro and in vivo. Notably, CSE1L expression was correlated with MSH6 expression in tumor samples and was associated with poor prognosis in patients with osteosarcoma. Taken together, our results demonstrate that the CSE1L-MSH6 axis has an important role in osteosarcoma progression.

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CSE1L knockdown inhibits osteosarcoma cell growth in vivo.(A) Representative blots displaying CSE1L protein expression in MNNG/HOS cells stably expressing sh-control or sh-CSE1L. β-actin was used as an internal control. (B) The upper diagram shows a photograph of tumor-bearing mice; the lower panel shows a photograph of the tumors when the mice were euthanized. (C) Growth curve drawn by measuring tumor volumes on the indicated days. Error bars represent the SEM. (D) This diagram shows the tumor weight in the sh-control group and the sh-CSE1L group. (E,F) These diagrams show representative green fluorescent protein (GFP) imaging and average photon number of the sh-control group and the sh-CSE1L group. (G,H) Representative images of Ki-67 staining in the sh-control group and the sh-CSE1L group. Magnification: 50×, 200×. For western blotting, full-length gels are presented in Supplementary Figure S9.
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f3: CSE1L knockdown inhibits osteosarcoma cell growth in vivo.(A) Representative blots displaying CSE1L protein expression in MNNG/HOS cells stably expressing sh-control or sh-CSE1L. β-actin was used as an internal control. (B) The upper diagram shows a photograph of tumor-bearing mice; the lower panel shows a photograph of the tumors when the mice were euthanized. (C) Growth curve drawn by measuring tumor volumes on the indicated days. Error bars represent the SEM. (D) This diagram shows the tumor weight in the sh-control group and the sh-CSE1L group. (E,F) These diagrams show representative green fluorescent protein (GFP) imaging and average photon number of the sh-control group and the sh-CSE1L group. (G,H) Representative images of Ki-67 staining in the sh-control group and the sh-CSE1L group. Magnification: 50×, 200×. For western blotting, full-length gels are presented in Supplementary Figure S9.

Mentions: To further determine the effect of CSE1L on osteosarcoma growth in vivo, MNNG/HOS cells stably expressing sh-control or sh-CSE1L were constructed. Protein expression was validated by western blotting (Fig. 3A). Transfected cells were subcutaneously injected into the left scapulae of nude mice, and the animals were closely monitored for tumor growth for 4 weeks. The tumor growth curve demonstrated that CSE1L knockdown significantly inhibited tumor growth in vivo. Both tumor volume and weight were reduced in the CSE1L knockdown group compared to that in the scramble control group (Fig. 3B,C and D). Following euthanization of the mice, green fluorescent protein (GFP) imaging was performed using the small animal imaging system, and the sum photon number was calculated. Typical GFP imaging of the sh-control group and the sh-CSE1L group is shown in Fig. 3E. The average photon number in the sh-CSE1L group was smaller than that in the sh-control group (Fig. 3F). Our findings revealed that the fluorescence signal in sh-CSE1L mice was lower than that in sh-control mice. As shown in Fig. 3G and H, IHC analysis identified that staining of the tissue proliferation marker Ki-67 was decreased in sh-CSE1L tumors than in sh-control tumors. Taken together, these results indicate that CSE1L knockdown hinders tumorigenesis of osteosarcoma cells in vivo.


CSE1L interaction with MSH6 promotes osteosarcoma progression and predicts poor patient survival
CSE1L knockdown inhibits osteosarcoma cell growth in vivo.(A) Representative blots displaying CSE1L protein expression in MNNG/HOS cells stably expressing sh-control or sh-CSE1L. β-actin was used as an internal control. (B) The upper diagram shows a photograph of tumor-bearing mice; the lower panel shows a photograph of the tumors when the mice were euthanized. (C) Growth curve drawn by measuring tumor volumes on the indicated days. Error bars represent the SEM. (D) This diagram shows the tumor weight in the sh-control group and the sh-CSE1L group. (E,F) These diagrams show representative green fluorescent protein (GFP) imaging and average photon number of the sh-control group and the sh-CSE1L group. (G,H) Representative images of Ki-67 staining in the sh-control group and the sh-CSE1L group. Magnification: 50×, 200×. For western blotting, full-length gels are presented in Supplementary Figure S9.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5384328&req=5

f3: CSE1L knockdown inhibits osteosarcoma cell growth in vivo.(A) Representative blots displaying CSE1L protein expression in MNNG/HOS cells stably expressing sh-control or sh-CSE1L. β-actin was used as an internal control. (B) The upper diagram shows a photograph of tumor-bearing mice; the lower panel shows a photograph of the tumors when the mice were euthanized. (C) Growth curve drawn by measuring tumor volumes on the indicated days. Error bars represent the SEM. (D) This diagram shows the tumor weight in the sh-control group and the sh-CSE1L group. (E,F) These diagrams show representative green fluorescent protein (GFP) imaging and average photon number of the sh-control group and the sh-CSE1L group. (G,H) Representative images of Ki-67 staining in the sh-control group and the sh-CSE1L group. Magnification: 50×, 200×. For western blotting, full-length gels are presented in Supplementary Figure S9.
Mentions: To further determine the effect of CSE1L on osteosarcoma growth in vivo, MNNG/HOS cells stably expressing sh-control or sh-CSE1L were constructed. Protein expression was validated by western blotting (Fig. 3A). Transfected cells were subcutaneously injected into the left scapulae of nude mice, and the animals were closely monitored for tumor growth for 4 weeks. The tumor growth curve demonstrated that CSE1L knockdown significantly inhibited tumor growth in vivo. Both tumor volume and weight were reduced in the CSE1L knockdown group compared to that in the scramble control group (Fig. 3B,C and D). Following euthanization of the mice, green fluorescent protein (GFP) imaging was performed using the small animal imaging system, and the sum photon number was calculated. Typical GFP imaging of the sh-control group and the sh-CSE1L group is shown in Fig. 3E. The average photon number in the sh-CSE1L group was smaller than that in the sh-control group (Fig. 3F). Our findings revealed that the fluorescence signal in sh-CSE1L mice was lower than that in sh-control mice. As shown in Fig. 3G and H, IHC analysis identified that staining of the tissue proliferation marker Ki-67 was decreased in sh-CSE1L tumors than in sh-control tumors. Taken together, these results indicate that CSE1L knockdown hinders tumorigenesis of osteosarcoma cells in vivo.

View Article: PubMed Central - PubMed

ABSTRACT

To discover tumor-associated proteins in osteosarcoma, a quantitative proteomic analysis was performed to identify proteins that were differentially expressed between osteosarcoma and human osteoblastic cells. Through clinical screening and a functional evaluation, chromosome segregation 1-like (CSE1L) protein was found to be related to the growth of osteosarcoma cells. To date, little is known about the function and underlying mechanism of CSE1L in osteosarcoma. In the present study, we show that knockdown of CSE1L inhibits osteosarcoma growth in vitro and in vivo. By co-immunoprecipitation and RNA-seq analysis, CSE1L was found to interact with mutS homolog 6 (MSH6) and function as a positive regulator of MSH6 protein in osteosarcoma cells. A rescue study showed that decreased growth of osteosarcoma cells by CSE1L knockdown was reversed by MSH6 overexpression, indicating that the activity of CSE1L was an MSH6-dependent function. In addition, depletion of MSH6 hindered cellular proliferation in vitro and in vivo. Notably, CSE1L expression was correlated with MSH6 expression in tumor samples and was associated with poor prognosis in patients with osteosarcoma. Taken together, our results demonstrate that the CSE1L-MSH6 axis has an important role in osteosarcoma progression.

No MeSH data available.


Related in: MedlinePlus