TP53INP1, a tumor suppressor, interacts with LC3 and ATG8-family proteins through the LC3-interacting region (LIR) and promotes autophagy-dependent cell death

Abstract

TP53INP1 (tumor protein 53-induced nuclear protein 1) is a tumor suppressor, whose expression is downregulated in cancers from different organs. It was described as a p53 target gene involved in cell death, cell-cycle arrest and cellular migration. In this work, we show that TP53INP1 is also able to interact with ATG8-family proteins and to induce autophagy-dependent cell death. In agreement with this finding, we observe that TP53INP1, which is mainly nuclear, relocalizes in autophagosomes during autophagy where it is eventually degraded. TP53INP1-LC3 interaction occurs via a functional LC3-interacting region (LIR). Inactivating mutations of this sequence abolish TP53INP1-LC3 interaction, relocalize TP53INP1 in autophagosomes and decrease TP53INP1 ability to trigger cell death. Interestingly, TP53INP1 binds to ATG8-family proteins with higher affinity than p62, suggesting that it could partially displace p62 from autophagosomes, modifying thereby their composition. Moreover, silencing the expression of autophagy related genes (ATG5 or Beclin-1) or inhibiting caspase activity significantly decreases cell death induced by TP53INP1. These data indicate that cell death observed after TP53INP1-LC3 interaction depends on both autophagy and caspase activity. We conclude that TP53INP1 could act as a tumor suppressor by inducing cell death by caspasedependent autophagy. Increased resistance to cell death participates in pancreatic cancer progression. Cells unable to undergo self elimination accumulate mutations and epigenetic modifications that in turn induce uncontrolled replication. 1 Various concomitant mechanisms exist in normal cells to induce death and, as consequence, a number of known cell-death regulators are missing in cancer cells. One of the most studied is the tumor suppressor TP53, which is inactivated in 450% of pancreatic tumors. 2 p53 induces cell death by both direct permeabilization of the outer mitochondrial membrane or translocation to the nucleus where it activates the transcription of several target genes. One of the p53 target genes is TP53INP1 (tumor protein 53-induced nuclear protein 1). 3-6 p53-dependent expression of TP53INP1 is triggered in response to several stress agents such as mutagens, ethanol, heat shock or conditions promoting reactive oxygen species formation (i.e., exposure to UV light or g-irradiation). 4,6 TP53INP1 interacts with kinases, HIPK2 and PKCd, which in turn phosphorylate p53 creating a positive feedback loop between p53 and TP53INP1. 7,8 Our laboratory demonstrated that TP53INP1 is a tumor suppressor on the basis of the following observations: (i) TP53INP1 deficient mice present with an increased susceptibility to tumor development; (ii) TP53INP1 is lost at very early stages of pancreatic carcinogenesis through a mechanism involving the oncogenic miR-155 microRNA and (iii) when TP53INP1 expression is restored in pancreatic cells, it suppresses xenograft growth by increasing apoptotic cell death through a caspase-dependent mechanism. 3,9,10 More recently, in an attempt t...

Oxidative stress-induced p53 activity is enhanced by a redox-sensitive TP53INP1 SUMOylation

“…27 When TP53INP1 expression is strongly induced by high oxidative stress, it interacts with LC3 in the autophagosomes, displacing p62, and inducing autophagic-dependent cell death. However, under low oxidative stress conditions, the intracellular concentration of TP53INP1 is very low (negligible in comparison with p62), and TP53INP1 does not displace p62 from the autophagosomes and therefore autophagy works as a cell survival mechanism.…”

“…We and others have characterized the importance of TP53INP1 in macroautophagy. 27,28 We previously showed that TP53INP1 colocalizes with LC3 (microtubuleassociated protein light chain 3) in autophagosomes and also increases the number of autophagosomes in the cytoplasm Where indicated, the WT or K113R mutant TP53INP1a was co-transfected with the previous constructs. Sixteen hours later, cells were exposed to H 2 O 2 1 mM for 1 h, and incubated for an additional 24-h period, luciferase activity was measured in cell lysates.…”

Molecular dynamics of the full-length p53 monomer

“…Just as an example, p53, with different degrees depending on its isoforms or its polymorphism at codon 72, 53 plays a crucial role in single-strand breaks during muscle function, 54 interacts with PRAP1, 55 as well as with DNA damage response. 56,57 The regulation of cell death by the p53 protein is quite complex, acting both at the level of autophagy, 58 lysosomes, 59 or at the core machinery of programmed cell death. [60][61][62][63][64][65] In addition to DNA damage response and cell death, p53 plays a crucial role in regulating cellular senescence [66][67][68] by interacting, for example, with MageA2, 69 PATZ1, 70 4E-BP1, 71 mTOR, 72,73 highlighting the vast complexity of this crucial regulation.…”

Sestrins Link Tumor Suppressors with the AMPK-TOR Signaling Network

“…p53 activates autophagy via transcriptional activation of genes involved in the regulation of the AMPK-mTORC1 pathway [70,73], although other p53 targets are directly involved in the autophagic process or operate through other mechanisms. The list of pro-autophagic genes regulated by p53 includes lysosomal protein damage-regulated autophagy modulator (DRAM), ULK1 [74], p53INP1 (which interacts with ATG8 and promote autophagic cell death) [75], Bax, Puma [18] and a new gene with unknown function ISG20L1 [76]. Interestingly that autophagy also contributes to p53-induced cell death, potentially providing the ground for elimination of damaged cells, if they are irreparable.…”