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Trp-tRNA synthetase bridges DNA-PKcs to PARP-1 to link IFN-? and p53 signaling.

ABSTRACT: Interferon-? (IFN-?) engenders strong antiproliferative responses, in part through activation of p53. However, the long-known IFN-?-dependent upregulation of human Trp-tRNA synthetase (TrpRS), a cytoplasmic enzyme that activates tryptophan to form Trp-AMP in the first step of protein synthesis, is unexplained. Here we report a nuclear complex of TrpRS with the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) and with poly(ADP-ribose) polymerase 1 (PARP-1), the major PARP in human cells. The IFN-?-dependent poly(ADP-ribosyl)ation of DNA-PKcs (which activates its kinase function) and concomitant activation of the tumor suppressor p53 were specifically prevented by Trp-SA, an analog of Trp-AMP that disrupted the TrpRS-DNA-PKcs-PARP-1 complex. The connection of TrpRS to p53 signaling in vivo was confirmed in a vertebrate system. These and further results suggest an unexpected evolutionary expansion of the protein synthesis apparatus to a nuclear role that links major signaling pathways.

SUBMITTER: Sajish M

PROVIDER: S-EPMC3780985 | biostudies-literature | 2012 Apr

REPOSITORIES: biostudies-literature

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Trp-tRNA synthetase bridges DNA-PKcs to PARP-1 to link IFN-γ and p53 signaling.

Sajish Mathew M Zhou Quansheng Q Kishi Shuji S Valdez Delgado M DM Kapoor Mili M Guo Min M Lee Sunhee S Kim Sunghoon S Yang Xiang-Lei XL Schimmel Paul P

Nature chemical biology 20120415 6

Interferon-γ (IFN-γ) engenders strong antiproliferative responses, in part through activation of p53. However, the long-known IFN-γ-dependent upregulation of human Trp-tRNA synthetase (TrpRS), a cytoplasmic enzyme that activates tryptophan to form Trp-AMP in the first step of protein synthesis, is unexplained. Here we report a nuclear complex of TrpRS with the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) and with poly(ADP-ribose) polymerase 1 (PARP-1), the major PARP in human cel ...[more]

PMID: 22504299

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Project description:BACKGROUND:Immune checkpoint inhibition and in particular anti-PD-1 immunotherapy have revolutionized the treatment of advanced melanoma. In this regard, higher tumoral PD-L1 protein (gene name: CD274) expression is associated with better clinical response and increased survival to anti-PD-1 therapy. Moreover, there is increasing evidence that tumor suppressor proteins are involved in immune regulation and are capable of modulating the expression of immune checkpoint proteins. Here, we determined the role of p53 protein (gene name: TP53) in the regulation of PD-L1 expression in melanoma. METHODS:We analyzed publicly available mRNA and protein expression data from the cancer genome/proteome atlas and performed immunohistochemistry on tumors with known TP53 status. Constitutive and IFN-?-induced PD-L1 expression upon p53 knockdown in wildtype, TP53-mutated or JAK2-overexpressing melanoma cells or in cells, in which p53 was rendered transcriptionally inactive by CRISPR/Cas9, was determined by immunoblot or flow cytometry. Similarly, PD-L1 expression was investigated after overexpression of a transcriptionally-impaired p53 (L22Q, W23S) in TP53-wt or a TP53-knockout melanoma cell line. Immunoblot was applied to analyze the IFN-? signaling pathway. RESULTS:For TP53-mutated tumors, an increased CD274 mRNA expression and a higher frequency of PD-L1 positivity was observed. Interestingly, positive correlations of IFNG mRNA and PD-L1 protein in both TP53-wt and -mutated samples and of p53 and PD-L1 protein suggest a non-transcriptional mode of action of p53. Indeed, cell line experiments revealed a diminished IFN-?-induced PD-L1 expression upon p53 knockdown in both wildtype and TP53-mutated melanoma cells, which was not the case when p53 wildtype protein was rendered transcriptionally inactive or by ectopic expression of p53L22Q,W23S, a transcriptionally-impaired variant, in TP53-wt cells. Accordingly, expression of p53L22Q,W23S in a TP53-knockout melanoma cell line boosted IFN-?-induced PD-L1 expression. The impaired PD-L1-inducibility after p53 knockdown was associated with a reduced JAK2 expression in the cells and was almost abrogated by JAK2 overexpression. CONCLUSIONS:While having only a small impact on basal PD-L1 expression, both wildtype and mutated p53 play an important positive role for IFN-?-induced PD-L1 expression in melanoma cells by supporting JAK2 expression. Future studies should address, whether p53 expression levels might influence response to anti-PD-1 immunotherapy.

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Trp-tRNA synthetase bridges DNA-PKcs to PARP-1 to link IFN-? and p53 signaling.

Publications

Trp-tRNA synthetase bridges DNA-PKcs to PARP-1 to link IFN-γ and p53 signaling.

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