a Department of Biochemistry, and Department of Cardiology of the Second Affiliated Hospital , Zhejiang University School of Medicine , Hangzhou , China.
b Collaborative Innovation Center for Diagnosis and Treatment of Infectious Disease , First Affiliated Hospital, Zhejiang University School of Medicine , Hangzhou , China.
Autophagy. 2019 Aug;15(8):1309-1321. doi: 10.1080/15548627.2019.1580510. Epub 2019 Feb 20.
TP53INP2/DOR (tumor protein p53-inducible nuclear protein 2) contributes to mammalian macroautophagy/autophagy by carrying nuclear deacetylated MAP1LC3/LC3 to the cytoplasm. Here, we report that in the cytoplasm, TP53INP2 further functions in autophagosome biogenesis by promoting LC3B-ATG7 interaction. Cytoplasmic expression of the N-terminal region of TP53INP2, which includes the LC3-interacting region, effectively triggered LC3B-PE production and autophagosome formation. In the cytoplasm, TP53INP2 colocalized to early autophagic membrane structures containing ATG14, ZFYVE1/DFCP1 or WIPI2. While knockdown of did not affect the formation of these autophagic structures, deletion of or , or mutations preventing TP53INP2 from LC3 interaction, disrupted the membrane binding of TP53INP2. TP53INP2 interacted directly with ATG7 to form a LC3B-TP53INP2-ATG7 complex in the cytoplasm. Loss of TP53INP2-LC3 or TP53INP2-ATG7 interaction significantly reduced LC3B-ATG7 binding. Together, these results suggest that after shifting from the nucleus, cytoplasmic TP53INP2 is targeted to early autophagic membranes accompanied by LC3, where it contributes to autophagosome biogenesis by mediating LC3-ATG7 interaction. : 3-MA, 3-methyladenine; 3NES, 3 repeated nuclear export signal; 3NLS, 3 repeated nuclear localization signal; ACTB, actin beta; ATG, autophagy related; BECN1, Beclin 1; mCherry, monomeric red fluorescent protein mCherry; GFP, green fluorescent protein; GST, glutathione S-transferase; KO, knockout; LC3B/MAP1LC3B, microtubule-associated protein 1 light chain 3 beta; LC3B[G120], LC3B mutant lacking amino acids after glycine 120; LDH, lactate dehydrogenase; LMNB1, lamin B1; LIR, LC3-interacting region; MTORC1, mechanistic target of rapamycin complex 1; PE, phosphatidylethanolamine; PtdIns3K, phosphatidylinositol 3-kinase; PtdIns3P, phosphatidylinositol 3-phosphate; rDNA, ribosomal DNA; RFP, red fluorescent protein; RNAi, RNA interference; SQSTM1, sequestosome 1; TP53INP2, tumor protein p53-inducible nuclear protein 2; TP53INP2[1-28], TP53INP2 mutant containing amino acids 1 to 28; TP53INP2[28-45], TP53INP2 mutant containing amino acids 28 to 45; TP53INP2[LIRΔ], TP53INP2 mutant lacking amino acids 1 to 144; TP53INP2[NLSΔ], TP53INP2 mutant lacking amino acids 145 to 221; TP53INP2, TP53INP2 mutant in which tryptophan 35 and isoleucine 38 are replaced with alanine; TP53INP2[NLSΔ], TP53INP2 mutant lacking amino acids 145 to 221, and tryptophan 35 and isoleucine 38 are replaced with alanine; TP53INP2[Δ1-28],[NLSΔ], TP53INP2 mutant lacking amino acids 1 to 28 and amino acids 145 to 221, and tryptophan 35 and isoleucine 38 are replaced with alanine; TP53INP2[Δ1-28],[NLSΔ], TP53INP2 mutant lacking amino acids 1 to 28 and amino acids 145 to 221; TP53INP2[Δ67-111],[NLSΔ], TP53INP2 mutant lacking amino acids 67 to 111 and amino acids 145 to 221; TP53INP2[Δ112-144],[NLSΔ], TP53INP2 mutant lacking amino acids 112 to 144 and amino acids 145 to 221; TUBB, tubulin beta class I; ULK1, unc-51 like autophagy activating kinase 1; VMP1, vacuole membrane protein 1; WIPI2, WD repeat domain phosphoinositide-interacting 2; WT, wild-type; ZFYVE1/DFCP1, zinc finger FYVE-type containing 1.
TP53INP2/DOR（肿瘤抑制基因 p53 诱导核蛋白 2）通过将核去乙酰化 MAP1LC3/LC3 携带到细胞质中，促进哺乳动物巨自噬/自噬体的形成。在这里，我们报告说在细胞质中，TP53INP2 通过促进 LC3B-ATG7 相互作用进一步参与自噬体生物发生。TP53INP2 N 端区域的细胞质表达，包括 LC3 相互作用区域，有效地触发了 LC3B-PE 的产生和自噬体的形成。在细胞质中，TP53INP2 与包含 ATG14、ZFYVE1/DFCP1 或 WIPI2 的早期自噬膜结构共定位。虽然 的敲低并不影响这些自噬结构的形成，但 或 或突变阻止 TP53INP2 与 LC3 相互作用，破坏了 TP53INP2 的膜结合。TP53INP2 与 ATG7 直接相互作用，在细胞质中形成一个 LC3B-TP53INP2-ATG7 复合物。失去 TP53INP2-LC3 或 TP53INP2-ATG7 相互作用显著降低了 LC3B-ATG7 的结合。综上所述，TP53INP2 从核转移后，靶向到伴有 LC3 的早期自噬膜，通过介导 LC3-ATG7 相互作用，促进自噬体生物发生。
