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J Biol Chem. 2017 Nov 3;292(44):18227-18239. doi: 10.1074/jbc.M117.781492. Epub 2017 Sep 18.

Nuclear insulin-like growth factor 1 receptor phosphorylates proliferating cell nuclear antigen and rescues stalled replication forks after DNA damage.

Author information

1
From the Department of Oncology-Pathology, Cancer Center Karolinska, Karolinska Institutet, Stockholm SE-171 76, Sweden.
2
From the Department of Oncology-Pathology, Cancer Center Karolinska, Karolinska Institutet, Stockholm SE-171 76, Sweden olle.larsson@ki.se.

Abstract

We have previously shown that the insulin-like growth factor 1 receptor (IGF-1R) translocates to the cell nucleus, where it binds to enhancer-like regions and increases gene transcription. Further studies have demonstrated that nuclear IGF-1R (nIGF-1R) physically and functionally interacts with some nuclear proteins, i.e. the lymphoid enhancer-binding factor 1 (Lef1), histone H3, and Brahma-related gene-1 proteins. In this study, we identified the proliferating cell nuclear antigen (PCNA) as a nIGF-1R-binding partner. PCNA is a pivotal component of the replication fork machinery and a main regulator of the DNA damage tolerance (DDT) pathway. We found that IGF-1R interacts with and phosphorylates PCNA in human embryonic stem cells and other cell lines. In vitro MS analysis of PCNA co-incubated with the IGF-1R kinase indicated tyrosine residues 60, 133, and 250 in PCNA as IGF-1R targets, and PCNA phosphorylation was followed by mono- and polyubiquitination. Co-immunoprecipitation experiments suggested that these ubiquitination events may be mediated by DDT-dependent E2/E3 ligases (e.g. RAD18 and SHPRH/HLTF). Absence of IGF-1R or mutation of Tyr-60, Tyr-133, or Tyr-250 in PCNA abrogated its ubiquitination. Unlike in cells expressing IGF-1R, externally induced DNA damage in IGF-1R-negative cells caused G1 cell cycle arrest and S phase fork stalling. Taken together, our results suggest a role of IGF-1R in DDT.

KEYWORDS:

insulin-like growth factor (IGF); phosphorylation; post-transcriptional regulation; proliferating cell nuclear antigen (PCNA); ubiquitin

PMID:
28924044
PMCID:
PMC5672045
DOI:
10.1074/jbc.M117.781492
[Indexed for MEDLINE]
Free PMC Article

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