Limits...
Dynamic changes in subcellular localization of cattle XLF during cell cycle, and focus formation of cattle XLF at DNA damage sites immediately after irradiation.

Koike M, Yutoku Y, Koike A - J. Vet. Med. Sci. (2015)

Bottom Line: Moreover, nuclear localization and accumulation of cattle XLF at DSB sites are dependent on 12 amino acids (288-299) of the C-terminal region of XLF (XLF CTR).Furthermore, basic amino acids on the XLF CTR are highly conserved among domestic animals including cattle, goat and horses, suggesting that the CTR is essential for the function of XLF in domestic animals.These findings might be useful to develop the molecular-targeting therapeutic drug taking XLF as a target molecule for human and domestic animals.

View Article: PubMed Central - PubMed

Affiliation: Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555, Japan.

ABSTRACT
Clinically, many chemotherapeutics and ionizing radiation (IR) have been applied for the treatment of various types of human and animal malignancies. These treatments kill tumor cells by causing DNA double-strand breaks (DSBs). Core factors of classical nonhomologous DNA-end joining (C-NHEJ) play a vital role in DSB repair. Thus, it is indispensable to clarify the mechanisms of C-NHEJ in order to develop next-generation chemotherapeutics for cancer. The XRCC4-like factor (XLF; also called Cernunnos or NHEJ1) is the lastly identified core NHEJ factor. The localization of core NHEJ factors might play a critical role in regulating NHEJ activity. The localization and function of XLF have not been elucidated in animal species other than mice and humans. Domestic cattle (Bos taurus) are the most common and vital domestic animals in many countries. Here, we show that the localization of cattle XLF changes dynamically during the cell cycle. Furthermore, EYFP-cattle XLF accumulates quickly at microirradiated sites and colocalizes with the DSB marker γH2AX. Moreover, nuclear localization and accumulation of cattle XLF at DSB sites are dependent on 12 amino acids (288-299) of the C-terminal region of XLF (XLF CTR). Furthermore, basic amino acids on the XLF CTR are highly conserved among domestic animals including cattle, goat and horses, suggesting that the CTR is essential for the function of XLF in domestic animals. These findings might be useful to develop the molecular-targeting therapeutic drug taking XLF as a target molecule for human and domestic animals.

No MeSH data available.


Related in: MedlinePlus

Expression and localization of XLF in cattle cells. (A) Total cell lysates from eachcell line (MDBK, 50 µg; HeLa, 10 µg) were analyzedby Western blotting using an anti-XLF antibody, an anti-Ku70 antibody or ananti-β-actin antibody. (B, C) Subcellular localization of XLF in cattle (MDBK) cellsduring the cell cycle. The cells were fixed and stained with an anti-XLF antibody.Nuclear DNA was counterstained with DAPI. The stained cells were analyzed by confocallaser microscopy. Arrowheads indicate the mitotic phase cells (B). The images shownare a representative example for interphase cells or mitotic phase cells (C).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4591152&req=5

fig_001: Expression and localization of XLF in cattle cells. (A) Total cell lysates from eachcell line (MDBK, 50 µg; HeLa, 10 µg) were analyzedby Western blotting using an anti-XLF antibody, an anti-Ku70 antibody or ananti-β-actin antibody. (B, C) Subcellular localization of XLF in cattle (MDBK) cellsduring the cell cycle. The cells were fixed and stained with an anti-XLF antibody.Nuclear DNA was counterstained with DAPI. The stained cells were analyzed by confocallaser microscopy. Arrowheads indicate the mitotic phase cells (B). The images shownare a representative example for interphase cells or mitotic phase cells (C).

Mentions: Expression and localization of cattle XLF in cattle cells: We examined theexpression and subcellular localization of XLF in cattle cells. First, we examined theexpression of XLF and Ku70 in the cattle cell line MDBK and the human cell line HeLa byWestern blot analysis using the anti-XLF antibody and anti-Ku70 antibody. As shown in Fig. 1AFig. 1.


Dynamic changes in subcellular localization of cattle XLF during cell cycle, and focus formation of cattle XLF at DNA damage sites immediately after irradiation.

Koike M, Yutoku Y, Koike A - J. Vet. Med. Sci. (2015)

Expression and localization of XLF in cattle cells. (A) Total cell lysates from eachcell line (MDBK, 50 µg; HeLa, 10 µg) were analyzedby Western blotting using an anti-XLF antibody, an anti-Ku70 antibody or ananti-β-actin antibody. (B, C) Subcellular localization of XLF in cattle (MDBK) cellsduring the cell cycle. The cells were fixed and stained with an anti-XLF antibody.Nuclear DNA was counterstained with DAPI. The stained cells were analyzed by confocallaser microscopy. Arrowheads indicate the mitotic phase cells (B). The images shownare a representative example for interphase cells or mitotic phase cells (C).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591152&req=5

fig_001: Expression and localization of XLF in cattle cells. (A) Total cell lysates from eachcell line (MDBK, 50 µg; HeLa, 10 µg) were analyzedby Western blotting using an anti-XLF antibody, an anti-Ku70 antibody or ananti-β-actin antibody. (B, C) Subcellular localization of XLF in cattle (MDBK) cellsduring the cell cycle. The cells were fixed and stained with an anti-XLF antibody.Nuclear DNA was counterstained with DAPI. The stained cells were analyzed by confocallaser microscopy. Arrowheads indicate the mitotic phase cells (B). The images shownare a representative example for interphase cells or mitotic phase cells (C).
Mentions: Expression and localization of cattle XLF in cattle cells: We examined theexpression and subcellular localization of XLF in cattle cells. First, we examined theexpression of XLF and Ku70 in the cattle cell line MDBK and the human cell line HeLa byWestern blot analysis using the anti-XLF antibody and anti-Ku70 antibody. As shown in Fig. 1AFig. 1.

Bottom Line: Moreover, nuclear localization and accumulation of cattle XLF at DSB sites are dependent on 12 amino acids (288-299) of the C-terminal region of XLF (XLF CTR).Furthermore, basic amino acids on the XLF CTR are highly conserved among domestic animals including cattle, goat and horses, suggesting that the CTR is essential for the function of XLF in domestic animals.These findings might be useful to develop the molecular-targeting therapeutic drug taking XLF as a target molecule for human and domestic animals.

View Article: PubMed Central - PubMed

Affiliation: Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555, Japan.

ABSTRACT
Clinically, many chemotherapeutics and ionizing radiation (IR) have been applied for the treatment of various types of human and animal malignancies. These treatments kill tumor cells by causing DNA double-strand breaks (DSBs). Core factors of classical nonhomologous DNA-end joining (C-NHEJ) play a vital role in DSB repair. Thus, it is indispensable to clarify the mechanisms of C-NHEJ in order to develop next-generation chemotherapeutics for cancer. The XRCC4-like factor (XLF; also called Cernunnos or NHEJ1) is the lastly identified core NHEJ factor. The localization of core NHEJ factors might play a critical role in regulating NHEJ activity. The localization and function of XLF have not been elucidated in animal species other than mice and humans. Domestic cattle (Bos taurus) are the most common and vital domestic animals in many countries. Here, we show that the localization of cattle XLF changes dynamically during the cell cycle. Furthermore, EYFP-cattle XLF accumulates quickly at microirradiated sites and colocalizes with the DSB marker γH2AX. Moreover, nuclear localization and accumulation of cattle XLF at DSB sites are dependent on 12 amino acids (288-299) of the C-terminal region of XLF (XLF CTR). Furthermore, basic amino acids on the XLF CTR are highly conserved among domestic animals including cattle, goat and horses, suggesting that the CTR is essential for the function of XLF in domestic animals. These findings might be useful to develop the molecular-targeting therapeutic drug taking XLF as a target molecule for human and domestic animals.

No MeSH data available.


Related in: MedlinePlus