Structure of the Novel C-terminal Domain of Vacuolar Protein Sorting 30/Autophagy-related Protein 6 and Its Specific Role in Autophagy

Institute of Microbial Chemistry, Tokyo, Tokyo 141-0021, Japan.
Journal of Biological Chemistry (Impact Factor: 4.57). 03/2012; 287(20):16256-66. DOI: 10.1074/jbc.M112.348250
Source: PubMed


Vacuolar protein sorting 30 (Vps30)/autophagy-related protein 6 (Atg6) is a common component of two distinct phosphatidylinositol 3-kinase complexes. In complex I, Atg14 links Vps30 to Vps34 lipid kinase and exerts its specific role in autophagy, whereas in complex II, Vps38 links Vps30 to Vps34 and plays a crucial role in vacuolar protein sorting. However, the molecular role of Vps30 in each pathway remains unclear. Here, we report the crystal structure of the carboxyl-terminal domain of Vps30. The structure is a novel globular fold comprised of three β-sheet-α-helix repeats. Truncation analyses showed that the domain is dispensable for the construction of both complexes, but is specifically required for autophagy through the targeting of complex I to the pre-autophagosomal structure. Thus, the domain is named the β-α repeated, autophagy-specific (BARA) domain. On the other hand, the N-terminal region of Vps30 was shown to be specifically required for vacuolar protein sorting. These structural and functional investigations of Vps30 domains, which are also conserved in the mammalian ortholog, Beclin 1, will form the basis for studying the molecular functions of this protein family in various biological processes.

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    • "tion of Vps34 kinase activity ( Furuya et al . , 2005 ) . This domain is essential to both autophagy and Beclin1 ' s tumor suppressive function ( Liang et al . , 1999 ) . The recent crystal structures of human Beclin1 ECD ( residues 248 – 450 ) and yeast Atg6 ECD ( 319 – 540 ) have revealed additional roles for this domain ( Huang et al . , 2012 ; Noda et al . , 2012 ) . Human Beclin1 ECD consists of four α - helices and three anti - parallel β - sheets , arranged into three α / β repeats showing three fold symmetry . The α - helices form an internal three - helix bundle surrounded by the three β - sheets , with additional loops and the last α - helix are found around the perimeter of the α / β repe"
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    • "Among them, 18 ATG genes encode core components required for autophagosome formation (Atg1–10, 12–14, 16–18, 29, and 31), the majority of which localize at least partly to the preautophagosomal structure (PAS), the nucleation site from which the autophagosome originates (Suzuki et al., 2001, 2007). These 18 proteins are classified into six functional groups: Atg1 kinase and its regulators, the PI3-kinase complex, the Atg9 vesicle, the Atg2–Atg18 complex, and two ubiquitin-like conjugation systems (Nakatogawa et al., 2009). "
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    • "The preparation of yeast protein extracts was carried out as previously reported [23] [24]. Immunoblotting was performed using anti-Ape1 antiserum (API-2, Hamasaki and Ohsumi, unpublished), anti-Atg2 antiserum [17], anti-HA antiserum (Covance, 16B12), anti-Atg8 (IN-13) or anti-Pgk1 antiserum (Invitrogen/Molecular Probes). "
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