Reversible lysine acetylation regulates nuclear translocation of TyrRS to counteract genotoxic oxidative stress

ABSTRACT

Aminoacyl-tRNA synthetases, catalyzing the first step of protein synthesis, have been shown to involve with multiple additional physiologic responses. Here, we summarize our findings that p300/CBP-Associated Factor and Sirtuin 1 play the reversible acetylation role in regulating the nuclear translocation of Tyrosyl-tRNA synthetase and activating transcription factor E2F1, thus facilitating the repair of damaged DNA.

Our recent finding shows that the reversible lysine acetylation regulates the nuclear translocation of TyrRS in response to oxidative stress.¹ Aminoacyl-tRNA synthetases (AARSs) catalyze the first step of protein synthesis by esterifying specific amino acids on the 3′ ends of their cognate tRNAs. AARSs family contain 20 enzymes and are extremely conserved during evolution.² However, recent studies have uncovered a role of multiple AARSs in pathology, and implicated their potential applications as pharmacological targets and therapeutic reagents. Recently, the increasing discovery of genetic mutations in human AARSs has been regarded as a significant determinant of disease etiology. Strikingly, mutations in cytoplasmic AARSs are always associated with Charcot–Marie–Tooth and related neuropathies. However, mutations in mitochondrial AARSs are related to a wider variety of syndromes and diseases.³ Furthermore, several chemical inhibitors targeting bacterial, fungal, and human AARSs have been developed as antibiotics or disease-targeting medicines. Remarkably, several studies have revealed that in addition to the well-known function in translation, many AARSs have been explored the noncanonical roles during angiogenesis, inflammation, DNA damage response, tumorigenesis, and other important physiopathological processes.³ For example, glutamyl-prolyl-tRNA synthetase (GluProRS) is involved in gene-specific silencing of translation,⁴ glycyl-tRNA synthetase (GlyRS) plays a critical role in neddylation through direct interacting with various components of the neddylation pathway.⁵ It was shown that TyrRS can translocate to the nucleus in response to oxidative damage or serum starvation stress, where it protects against DNA damage by activating the transcription factor E2F1 and subsequent downstream DNA repair genes.⁶ However, the mechanism of how the nuclear translocation of TyrRS is regulated is unknown.

Protein lysine acetylation has recently emerged as a broadly used modification in the regulation of diverse cellular processes, including gene silencing, oxidative stress, DNA repair, cell survival and migration, and metabolism.⁷ Most identified acetylated proteins are transcription factors in the nucleus and metabolic enzymes outside the nucleus.⁷ Interestingly, numbers of AARSs, including TyrRS, are also acetylated in several proteomic studies. However, the connection between acetylation modification and these AARSs needs to be established.

In our study, we used an antibody recognizing pan-acetylated lysine to confirm that TyrRS is highly acetylated in vivo under oxidative stress, and that acetylation promotes its nuclear translocation. K244 was identified as the major acetylation site of TyrRS, and K244 acetylation significantly decreased TyrRS activity while enhancing its nuclear translocation, thereby protecting against DNA damage in vivo.¹ Additional biochemical studies demonstrated that PCAF acts as the acetyltransferase, and SIRT1 is a bona fide deacetylase of TyrRS, whose deacetylation activity prevents TyrRS nuclear localization.¹ From these results, we propose a model in which reversible acetylation in the nuclear localization signal (NLS) of TyrRS regulates its nuclear transport under oxidative stress (see Fig. 1). Furthermore, DNA repair genes downstream of E2F1 are activated in response to the oxidative stress. Our findings imply that it is able to generate a therapeutic strategy for physiologic diseases characterized by DNA damage through targeting the K244 residue of TyrRS.

Figure 1.

Mechanisms through which acetylation mediates TyrRS nuclear translocation in response to oxidative stress: PCAF acetylates TyrRS under oxidative stress and promotes its nuclear transport to activate DNA repair genes downstream of E2F1, and SIRT1 deacetylates TyrRS, thus regulating its nuclear export. Ac: acetylation.

Notably, the histone deacetylase (HDAC) inhibitor trichostatin A can be used to chemically mimic the protective effect of nuclear TyrRS.⁶ Our results are supportive with this finding and show that the Sirtuin inhibitor nicotinamide (NAM) promotes TyrRS nuclear translocation by favoring K244 acetylation, thereby preventing DNA damage. The K244 site is the most evolutionarily conserved lysine in TyrRS and is located in its NLS. Despite the controversial effect of HDAC inhibitors on DNA damage, three HDAC inhibitors have been licensed by the Food and Drug Administration (FDA) for the treatment of cutaneous/peripheral T-cell lymphoma,⁸ and one HDAC inhibitor has been approved by the Chinese government for the treatment of pancreatic cancer. Moreover, more than a dozen HDAC inhibitors are in various stages of clinical trials for the treatment of hematological malignancies and solid tumors.⁸ Taken together with previous data, our findings imply that acetylated K244 TyrRS may strengthen the value of HDAC inhibitors as tumor therapies.

Intracellular oxidative stress refers to increased levels of reactive oxygen species that can cause damage to lipids, proteins, and DNA, which have been associated with a variety of pathologies. SIRT1 is the most studied NAD⁺-dependent deacetylase and has been involved in a wide variety of cellular processes from cancer to aging.⁹ SIRT1 was reported to form an intricate complex with autopoly-ADP ribosylation of poly(ADP ribose) polymerase 1 (PARP1), another major signaling molecule in DNA damage response, and protect cells against DNA damage. More interestingly, serum starvation or resveratrol supplements promote the nuclear import of TyrRS and increase the PARP1 activity.¹⁰ Consistent with this finding, we demonstrated that resveratrol increased the acetylation level of TyrRS at K244 site in a dose-dependent manner through upregulating the expression of PCAF.¹ Together, these data suggest that under oxidative stress, PARP1 activation is due to negative feedback among SIRT1, TyrRS, and PARP1.

AARSs are among the higher abundant proteins in cells, which was assumed to meet a large demand for protein synthesis. Recently, many studies have shown that AARSs take on multiple functions in addition to aminoacylation activity, which may explain that the levels of cellular AARSs are larger, excessive for what is demanded in protein synthesis. Our study provides another evidence that PCAF and SIRT1 acting the reversible acetylation in TyrRS regulate its nuclear transport to involve with DNA damage repair, as proved in our in vivo models.¹ The present study not only reveals an unknown regulatory mechanism linking posttranslational modification to tRNA synthetases TyrRS but also enriches our knowledge of the regulatory pathway between PCAF/SIRT1 and DNA damage response. More importantly, we found that K244 acetylation of TyrRS is protective against DNA damage, suggesting a potential therapeutic opportunity to treat DNA-damage-associated diseases by targeting the K244 residue of TyrRS.

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.