GADD45A inhibits autophagy by regulating the interaction between BECN1 and PIK3C3

GADD45A is a TP53-regulated and DNA damage-inducible tumor suppressor protein, which regulates cell cycle arrest, apoptosis, and DNA repair, and inhibits tumor growth and angiogenesis. However, the function of GADD45A in autophagy remains unknown. In this report, we demonstrate that GADD45A plays an important role in regulating the process of autophagy. GADD45A is able to decrease LC3-II expression and numbers of autophagosomes in mouse tissues and different cancer cell lines. Using bafilomycin A1 treatment, we have observed that GADD45A regulates autophagosome initiation. Likely, GADD45A inhibition of autophagy is through its influence on the interaction between BECN1 and PIK3C3. Immunoprecipitation and GST affinity isolation assays exhibit that GADD45A directly interacts with BECN1, and in turn dissociates the BECN1-PIK3C3 complex. Furthermore, we have mapped the 71 to 81 amino acids of the GADD45A protein that are necessary for the GADD45A interaction with BECN1. Knockdown of BECN1 can abolish autophagy alterations induced by GADD45A. Taken together, these findings provide the novel evidence that GADD45A inhibits autophagy via impairing the BECN1-PIK3C3 complex formation.

GADD45A是受TP53调控且可被DNA损伤诱导的抑癌蛋白，其可调控细胞周期阻滞、细胞凋亡与DNA修复过程，并抑制肿瘤生长及血管生成。然而，GADD45A在细胞自噬（autophagy）中的功能仍有待阐明。本研究证实，GADD45A在调控细胞自噬进程中发挥关键作用。GADD45A能够降低小鼠组织及不同癌细胞系中的LC3-II表达水平与自噬体（autophagosome）数量。借助巴弗洛霉素A1（bafilomycin A1）处理实验，我们发现GADD45A可调控自噬体的起始过程。GADD45A对自噬的抑制作用，大概率是通过影响BECN1与PIK3C3之间的相互作用实现的。免疫沉淀（immunoprecipitation）与GST亲和分离实验结果表明，GADD45A可直接与BECN1结合，并进而解离BECN1-PIK3C3复合物。此外，我们已明确GADD45A蛋白中71至81位氨基酸是其与BECN1相互作用所必需的结构区域。敲低BECN1可消除GADD45A诱导的自噬异常。综上，本研究发现GADD45A通过损害BECN1-PIK3C3复合物的形成来抑制细胞自噬，该发现为相关机制提供了全新的实验证据。