Abstract
Ambra1 is considered an autophagy and trafficking protein with roles in neurogenesis and cancer cell invasion. Here, we report that Ambra1 also localizes to the nucleus of cancer cells, where it has a novel nuclear scaffolding function that controls gene expression. Using biochemical fractionation and proteomics, we found that Ambra1 binds to multiple classes of proteins in the nucleus, including nuclear pore proteins, adaptor proteins such as FAK and Akap8, chromatin-modifying proteins, and transcriptional regulators like Brg1 and Atf2. We identified biologically important genes, such as Angpt1, Tgfb2, Tgfb3, Itga8, and Itgb7, whose transcription is regulated by Ambra1-scaffolded complexes, likely by altering histone modifications and Atf2 activity. Therefore, in addition to its recognized roles in autophagy and trafficking, Ambra1 scaffolds protein complexes at chromatin, regulating transcriptional signaling in the nucleus. This novel function for Ambra1, and the specific genes impacted, may help to explain the wider role of Ambra1 in cancer cell biology.
Highlights
Ambra1 is already known to be an important protein in physiology, e.g. in the development of the central nervous system, vertebrate embryogenesis, adult neurogenesis, and cancer cell invasion [1,2,3,4,5]
Like Ambra1, other proteins involved in autophagy have been reported in the nucleus, e.g. LC3B binds to Lamin B1, mediating the degradation of the nuclear lamina and Beclin 1, promoting autophagy-independent DNA damagerepair[46, 47]
No typical nuclear localization sequence is evident for Ambra1; the mechanism of nuclear translocation is unknown; as nuclear Ambra1 interacts with components of nuclear pore complexes and importins (Fig. 1C), it is likely that nuclear import of Ambra1 occurs via binding to these in some way
Summary
Ambra (activating molecule in Beclin1-regulated autophagy) is already known to be an important protein in physiology, e.g. in the development of the central nervous system, vertebrate embryogenesis, adult neurogenesis, and cancer cell invasion [1,2,3,4,5]. We investigate a nuclear function of Ambra and show that it binds to FAK in the nucleus, as well as to other nuclear adaptor proteins, nuclear pore components, histone-modifying enzymes, and regulators of transcription, in some cases regulating their recruitment to chromatin. Ambra forms complexes with Akap, Brg, and Atf and is responsible for the recruitment of Akap, Bgr, the Mediator complex component Cdk, and p-Atf T71 to chromatin. Both Ambra and its binding protein Akap regulate the binding of transcriptional proteins to chromatin, especially pAtf T71, and proteins that modulate histone modifications. Ambra acts as a scaffold protein in the nucleus, recruiting transcriptional regulators to chromatin. This creates an Ambra1-dependent nuclear microdomain that regulates gene expression
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