TY - JOUR
T1 - Mapping axon initial segment structure and function by multiplexed proximity biotinylation
AU - Hamdan, Hamdan
AU - Lim, Brian C.
AU - Torii, Tomohiro
AU - Joshi, Abhijeet
AU - Konning, Matthias
AU - Smith, Cameron
AU - Palmer, Donna J.
AU - Ng, Philip
AU - Leterrier, Christophe
AU - Oses-Prieto, Juan A.
AU - Burlingame, Alma L.
AU - Rasband, Matthew N.
N1 - Funding Information:
The work reported here was supported by the following research grants: NIH NS044916 (MNR); NIH NS069688 (MNR); GM103481 (ALB), CNRS ATIP AO2016 (CL), and by the Dr. Miriam and Sheldon G. Adelson Medical Research Foundation (ALB and MNR). We thank Dr. Jonathan Terman (UT Southwestern) for insightful suggestions on Mical3 functions. We thank Dr. Takeshi Yoshimura for construction of the α2 spectrin-GST fusion proteins.
Publisher Copyright:
© 2020, The Author(s).
PY - 2020/12/1
Y1 - 2020/12/1
N2 - Axon initial segments (AISs) generate action potentials and regulate the polarized distribution of proteins, lipids, and organelles in neurons. While the mechanisms of AIS Na+ and K+ channel clustering are understood, the molecular mechanisms that stabilize the AIS and control neuronal polarity remain obscure. Here, we use proximity biotinylation and mass spectrometry to identify the AIS proteome. We target the biotin-ligase BirA* to the AIS by generating fusion proteins of BirA* with NF186, Ndel1, and Trim46; these chimeras map the molecular organization of AIS intracellular membrane, cytosolic, and microtubule compartments. Our experiments reveal a diverse set of biotinylated proteins not previously reported at the AIS. We show many are located at the AIS, interact with known AIS proteins, and their loss disrupts AIS structure and function. Our results provide conceptual insights and a resource for AIS molecular organization, the mechanisms of AIS stability, and polarized trafficking in neurons.
AB - Axon initial segments (AISs) generate action potentials and regulate the polarized distribution of proteins, lipids, and organelles in neurons. While the mechanisms of AIS Na+ and K+ channel clustering are understood, the molecular mechanisms that stabilize the AIS and control neuronal polarity remain obscure. Here, we use proximity biotinylation and mass spectrometry to identify the AIS proteome. We target the biotin-ligase BirA* to the AIS by generating fusion proteins of BirA* with NF186, Ndel1, and Trim46; these chimeras map the molecular organization of AIS intracellular membrane, cytosolic, and microtubule compartments. Our experiments reveal a diverse set of biotinylated proteins not previously reported at the AIS. We show many are located at the AIS, interact with known AIS proteins, and their loss disrupts AIS structure and function. Our results provide conceptual insights and a resource for AIS molecular organization, the mechanisms of AIS stability, and polarized trafficking in neurons.
UR - https://www.scopus.com/pages/publications/85077480107
U2 - 10.1038/s41467-019-13658-5
DO - 10.1038/s41467-019-13658-5
M3 - Article
C2 - 31900387
AN - SCOPUS:85077480107
SN - 2041-1723
VL - 11
JO - Nature Communications
JF - Nature Communications
IS - 1
M1 - 100
ER -