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Title: Structural transitions in conserved, ordered Beclin 1 domains essential to regulating autophagy

Abstract

Beclin 1 (BECN1) is a key regulator of autophagy, a critical catabolic homeostasis pathway that involves sequestration of selected cytoplasmic components by multilayered vesicles called autophagosomes, followed by lysosomal fusion and degradation. BECN1 is a core component of class III phosphatidylinositol-3-kinase complexes responsible for autophagosome nucleation. Without heterologous binding partners, BECN1 forms an antiparallel homodimer via its coiled-coil domain (CCD). However, the last 16 CCD residues, composing an “overlap helix” (OH), have been crystallized in two mutually exclusive states: either as part of the CCD or packed against the C-terminal β-α repeated, autophagy-specific domain (BARAD). Here, using CD spectroscopy, isothermal titration calorimetry, and small-angle X-ray scattering, we show that in the homodimeric state, the OH transitions between these two different packing states, with the predominant state comprising the OH packed against the BARAD, contrary to expectations based on known BECN1 interactions with heterologous partners. We confirmed this observation by comparing the impact of mutating four residues that mediate packing of the OH against both the CCD and BARAD on structure and stability of the CCD, the OH+BARAD, and the two-domain CCD–BARAD. Last, we used cellular assays to demonstrate that mutation of these OH-interface residues abrogates starvation-induced up-regulation of autophagy butmore » does not affect basal autophagy. In summary, we have identified a BECN1 helical region that transitions between packing as part of either one of two conserved domains (i.e. the CCD or the BARAD). Our findings have important implications for the relative stability of autophagy-inactive and autophagy-active BECN1 complexes.« less

Authors:
; ; ; ; ; ORCiD logo; ORCiD logo
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
NSFNIH
OSTI Identifier:
1404949
Resource Type:
Journal Article
Resource Relation:
Journal Name: Journal of Biological Chemistry; Journal Volume: 292; Journal Issue: 39
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES

Citation Formats

Glover, Karen, Li, Yue, Mukhopadhyay, Shreya, Leuthner, Zoe, Chakravarthy, Srinivas, Colbert, Christopher L., and Sinha, Sangita C.. Structural transitions in conserved, ordered Beclin 1 domains essential to regulating autophagy. United States: N. p., 2017. Web. doi:10.1074/jbc.M117.804195.
Glover, Karen, Li, Yue, Mukhopadhyay, Shreya, Leuthner, Zoe, Chakravarthy, Srinivas, Colbert, Christopher L., & Sinha, Sangita C.. Structural transitions in conserved, ordered Beclin 1 domains essential to regulating autophagy. United States. doi:10.1074/jbc.M117.804195.
Glover, Karen, Li, Yue, Mukhopadhyay, Shreya, Leuthner, Zoe, Chakravarthy, Srinivas, Colbert, Christopher L., and Sinha, Sangita C.. Thu . "Structural transitions in conserved, ordered Beclin 1 domains essential to regulating autophagy". United States. doi:10.1074/jbc.M117.804195.
@article{osti_1404949,
title = {Structural transitions in conserved, ordered Beclin 1 domains essential to regulating autophagy},
author = {Glover, Karen and Li, Yue and Mukhopadhyay, Shreya and Leuthner, Zoe and Chakravarthy, Srinivas and Colbert, Christopher L. and Sinha, Sangita C.},
abstractNote = {Beclin 1 (BECN1) is a key regulator of autophagy, a critical catabolic homeostasis pathway that involves sequestration of selected cytoplasmic components by multilayered vesicles called autophagosomes, followed by lysosomal fusion and degradation. BECN1 is a core component of class III phosphatidylinositol-3-kinase complexes responsible for autophagosome nucleation. Without heterologous binding partners, BECN1 forms an antiparallel homodimer via its coiled-coil domain (CCD). However, the last 16 CCD residues, composing an “overlap helix” (OH), have been crystallized in two mutually exclusive states: either as part of the CCD or packed against the C-terminal β-α repeated, autophagy-specific domain (BARAD). Here, using CD spectroscopy, isothermal titration calorimetry, and small-angle X-ray scattering, we show that in the homodimeric state, the OH transitions between these two different packing states, with the predominant state comprising the OH packed against the BARAD, contrary to expectations based on known BECN1 interactions with heterologous partners. We confirmed this observation by comparing the impact of mutating four residues that mediate packing of the OH against both the CCD and BARAD on structure and stability of the CCD, the OH+BARAD, and the two-domain CCD–BARAD. Last, we used cellular assays to demonstrate that mutation of these OH-interface residues abrogates starvation-induced up-regulation of autophagy but does not affect basal autophagy. In summary, we have identified a BECN1 helical region that transitions between packing as part of either one of two conserved domains (i.e. the CCD or the BARAD). Our findings have important implications for the relative stability of autophagy-inactive and autophagy-active BECN1 complexes.},
doi = {10.1074/jbc.M117.804195},
journal = {Journal of Biological Chemistry},
number = 39,
volume = 292,
place = {United States},
year = {Thu Aug 10 00:00:00 EDT 2017},
month = {Thu Aug 10 00:00:00 EDT 2017}
}