The PERK/ATF4/LAMP3-arm of the unfolded protein response affects radioresistance by interfering with the DNA damage response

Anika Nagelkerke; Johan Bussink; Albert J van der Kogel; Fred C G J Sweep; Paul N Span

doi:10.1016/j.radonc.2013.06.037

The PERK/ATF4/LAMP3-arm of the unfolded protein response affects radioresistance by interfering with the DNA damage response

Anika Nagelkerke, Johan Bussink, Albert J van der Kogel, Fred C G J Sweep, Paul N Span^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › Academic › peer-review

75 Citations (Scopus)

Abstract

BACKGROUND AND PURPOSE: Lysosome-associated membrane protein 3 (LAMP3) is induced by the PKR-like ER kinase (PERK)/activating transcription factor 4 (ATF4)-arm of the unfolded protein response (UPR) during hypoxia. LAMP3 has prognostic value in breast cancer patients treated with radiotherapy. Here, we specifically investigated the role of the PERK/ATF4/LAMP3-arm in the radiation response of breast cancer cells.

MATERIAL AND METHODS: Radiosensitivity of breast cancer cells was examined after siRNA-mediated knockdown of PERK, ATF4 and LAMP3. Activation of DNA damage repair proteins was evaluated by Western blotting and immunocytochemistry.

RESULTS: Knockdown of the PERK/ATF4/LAMP3-arm and chemical inhibition of PERK could radiosensitise MDA-MB-231 cells significantly. Western blot analysis of several DNA damage repair proteins showed that LAMP3 knockdowns had an attenuated DNA damage response after radiation compared to controls. γ-H2AX foci analysis revealed that LAMP3 knockdowns had a reduced number of positive cells after irradiation, indicating that their DNA damage repair signalling response is decreased. In addition, the effect of autophagy inhibition was examined and revealed a radiosensitising effect and the presence of residual γ-H2AX foci.

CONCLUSIONS: The PERK/ATF4/LAMP3-arm causes radioresistance of breast cancer cells by increasing DNA damage repair signalling. Inhibition of PERK and/or autophagy may sensitise tumours to radiotherapy.

Original language	English
Pages (from-to)	415-21
Number of pages	7
Journal	Radiotherapy and Oncology
Volume	108
Issue number	3
DOIs	https://doi.org/10.1016/j.radonc.2013.06.037
Publication status	Published - Sept-2013
Externally published	Yes

Keywords

Activating Transcription Factor 4/antagonists & inhibitors
Autophagy
Breast Neoplasms/radiotherapy
Cell Line, Tumor
DNA Damage
Female
Histones/analysis
Humans
Lysosome-Associated Membrane Glycoproteins/antagonists & inhibitors
Neoplasm Proteins/antagonists & inhibitors
Protein Phosphatase 1/physiology
Radiation Tolerance
Unfolded Protein Response
eIF-2 Kinase/antagonists & inhibitors

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@article{72cad51fbf4944518aebd2215eee584a,

title = "The PERK/ATF4/LAMP3-arm of the unfolded protein response affects radioresistance by interfering with the DNA damage response",

abstract = "BACKGROUND AND PURPOSE: Lysosome-associated membrane protein 3 (LAMP3) is induced by the PKR-like ER kinase (PERK)/activating transcription factor 4 (ATF4)-arm of the unfolded protein response (UPR) during hypoxia. LAMP3 has prognostic value in breast cancer patients treated with radiotherapy. Here, we specifically investigated the role of the PERK/ATF4/LAMP3-arm in the radiation response of breast cancer cells.MATERIAL AND METHODS: Radiosensitivity of breast cancer cells was examined after siRNA-mediated knockdown of PERK, ATF4 and LAMP3. Activation of DNA damage repair proteins was evaluated by Western blotting and immunocytochemistry.RESULTS: Knockdown of the PERK/ATF4/LAMP3-arm and chemical inhibition of PERK could radiosensitise MDA-MB-231 cells significantly. Western blot analysis of several DNA damage repair proteins showed that LAMP3 knockdowns had an attenuated DNA damage response after radiation compared to controls. γ-H2AX foci analysis revealed that LAMP3 knockdowns had a reduced number of positive cells after irradiation, indicating that their DNA damage repair signalling response is decreased. In addition, the effect of autophagy inhibition was examined and revealed a radiosensitising effect and the presence of residual γ-H2AX foci.CONCLUSIONS: The PERK/ATF4/LAMP3-arm causes radioresistance of breast cancer cells by increasing DNA damage repair signalling. Inhibition of PERK and/or autophagy may sensitise tumours to radiotherapy.",

keywords = "Activating Transcription Factor 4/antagonists & inhibitors, Autophagy, Breast Neoplasms/radiotherapy, Cell Line, Tumor, DNA Damage, Female, Histones/analysis, Humans, Lysosome-Associated Membrane Glycoproteins/antagonists & inhibitors, Neoplasm Proteins/antagonists & inhibitors, Protein Phosphatase 1/physiology, Radiation Tolerance, Unfolded Protein Response, eIF-2 Kinase/antagonists & inhibitors",

author = "Anika Nagelkerke and Johan Bussink and {van der Kogel}, {Albert J} and Sweep, {Fred C G J} and Span, {Paul N}",

year = "2013",

month = sep,

doi = "10.1016/j.radonc.2013.06.037",

language = "English",

volume = "108",

pages = "415--21",

journal = "Radiotherapy and Oncology",

issn = "0167-8140",

publisher = "ELSEVIER IRELAND LTD",

number = "3",

}

TY - JOUR

T1 - The PERK/ATF4/LAMP3-arm of the unfolded protein response affects radioresistance by interfering with the DNA damage response

AU - Nagelkerke, Anika

AU - Bussink, Johan

AU - van der Kogel, Albert J

AU - Sweep, Fred C G J

AU - Span, Paul N

PY - 2013/9

Y1 - 2013/9

N2 - BACKGROUND AND PURPOSE: Lysosome-associated membrane protein 3 (LAMP3) is induced by the PKR-like ER kinase (PERK)/activating transcription factor 4 (ATF4)-arm of the unfolded protein response (UPR) during hypoxia. LAMP3 has prognostic value in breast cancer patients treated with radiotherapy. Here, we specifically investigated the role of the PERK/ATF4/LAMP3-arm in the radiation response of breast cancer cells.MATERIAL AND METHODS: Radiosensitivity of breast cancer cells was examined after siRNA-mediated knockdown of PERK, ATF4 and LAMP3. Activation of DNA damage repair proteins was evaluated by Western blotting and immunocytochemistry.RESULTS: Knockdown of the PERK/ATF4/LAMP3-arm and chemical inhibition of PERK could radiosensitise MDA-MB-231 cells significantly. Western blot analysis of several DNA damage repair proteins showed that LAMP3 knockdowns had an attenuated DNA damage response after radiation compared to controls. γ-H2AX foci analysis revealed that LAMP3 knockdowns had a reduced number of positive cells after irradiation, indicating that their DNA damage repair signalling response is decreased. In addition, the effect of autophagy inhibition was examined and revealed a radiosensitising effect and the presence of residual γ-H2AX foci.CONCLUSIONS: The PERK/ATF4/LAMP3-arm causes radioresistance of breast cancer cells by increasing DNA damage repair signalling. Inhibition of PERK and/or autophagy may sensitise tumours to radiotherapy.

AB - BACKGROUND AND PURPOSE: Lysosome-associated membrane protein 3 (LAMP3) is induced by the PKR-like ER kinase (PERK)/activating transcription factor 4 (ATF4)-arm of the unfolded protein response (UPR) during hypoxia. LAMP3 has prognostic value in breast cancer patients treated with radiotherapy. Here, we specifically investigated the role of the PERK/ATF4/LAMP3-arm in the radiation response of breast cancer cells.MATERIAL AND METHODS: Radiosensitivity of breast cancer cells was examined after siRNA-mediated knockdown of PERK, ATF4 and LAMP3. Activation of DNA damage repair proteins was evaluated by Western blotting and immunocytochemistry.RESULTS: Knockdown of the PERK/ATF4/LAMP3-arm and chemical inhibition of PERK could radiosensitise MDA-MB-231 cells significantly. Western blot analysis of several DNA damage repair proteins showed that LAMP3 knockdowns had an attenuated DNA damage response after radiation compared to controls. γ-H2AX foci analysis revealed that LAMP3 knockdowns had a reduced number of positive cells after irradiation, indicating that their DNA damage repair signalling response is decreased. In addition, the effect of autophagy inhibition was examined and revealed a radiosensitising effect and the presence of residual γ-H2AX foci.CONCLUSIONS: The PERK/ATF4/LAMP3-arm causes radioresistance of breast cancer cells by increasing DNA damage repair signalling. Inhibition of PERK and/or autophagy may sensitise tumours to radiotherapy.

KW - Activating Transcription Factor 4/antagonists & inhibitors

KW - Autophagy

KW - Breast Neoplasms/radiotherapy

KW - Cell Line, Tumor

KW - DNA Damage

KW - Female

KW - Histones/analysis

KW - Humans

KW - Lysosome-Associated Membrane Glycoproteins/antagonists & inhibitors

KW - Neoplasm Proteins/antagonists & inhibitors

KW - Protein Phosphatase 1/physiology

KW - Radiation Tolerance

KW - Unfolded Protein Response

KW - eIF-2 Kinase/antagonists & inhibitors

U2 - 10.1016/j.radonc.2013.06.037

DO - 10.1016/j.radonc.2013.06.037

M3 - Article

C2 - 23891100

SN - 0167-8140

VL - 108

SP - 415

EP - 421

JO - Radiotherapy and Oncology

JF - Radiotherapy and Oncology

IS - 3

ER -