Dataset Information

An organellar Nα-acetyltransferase Naa60 acetylates cytosolic N-termini of transmembrane proteins and maintains Golgi integrity

ABSTRACT: N-terminal acetylation is a major and vital protein modification catalysed by N-terminal acetyltransferases (NATs). NatF or Nα-acetyltransferase 60 (Naa60) was recently identified as a NAT in higher eukaryotes. Here, we found that Naa60 differs from all other known NATs by its Golgi localization. A new membrane topology assay named PROMPT and a selective membrane permeabilization assay established Naa60 to face the cytosolic side of intracellular membranes. An Nt-acetylome analysis of NAA60 knockdown cells revealed that Naa60, as opposed to other NATs, specifically acetylates transmembrane proteins, and that is has a preference for N-termini facing the cytosol. Moreover, NAA60 knockdown caused Golgi fragmentation, indicating an important role in maintenance of Golgi structural integrity. This work uncovers the first NAT associated with membranous compartments and establishes N-terminal acetylation as a common modification among transmembrane proteins, a thus far poorly characterized part of the N-terminal acetylome.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Jonathan Vandenbussche

LAB HEAD: Kris Gevaert

PROVIDER: PXD001155 | Pride | 2019-06-04

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	R7647_Petra_807_siRNA_hNaa60p_A432_28-32-1.RAW	Raw
	R7648_Petra_807_siRNA_hNaa60p_A432_28-32-2.RAW	Raw
	R7649_Petra_807_siRNA_hNaa60p_A432_28-32-3.RAW	Raw
	R7650_Petra_807_siRNA_hNaa60p_A432_28-32-4.RAW	Raw
	R7651_Petra_807_siRNA_hNaa60p_A432_28-32-5.RAW	Raw

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An organellar nα-acetyltransferase, naa60, acetylates cytosolic N termini of transmembrane proteins and maintains Golgi integrity.

Aksnes Henriette H Van Damme Petra P Goris Marianne M Starheim Kristian K KK Marie Michaël M Støve Svein Isungset SI Hoel Camilla C Kalvik Thomas Vikestad TV Hole Kristine K Glomnes Nina N Furnes Clemens C Ljostveit Sonja S Ziegler Mathias M Niere Marc M Gevaert Kris K Arnesen Thomas T

Cell reports 20150226 8

N-terminal acetylation is a major and vital protein modification catalyzed by N-terminal acetyltransferases (NATs). NatF, or Nα-acetyltransferase 60 (Naa60), was recently identified as a NAT in multicellular eukaryotes. Here, we find that Naa60 differs from all other known NATs by its Golgi localization. A new membrane topology assay named PROMPT and a selective membrane permeabilization assay established that Naa60 faces the cytosolic side of intracellular membranes. An Nt-acetylome analysis of ...[more]

PMID: 25732826

Similar Datasets

Project description:Co-translational N-terminal (Nt-) acetylation of nascent polypeptides is catalyzed by N-terminal acetyltransferases (NATs). The very N-terminal amino acid sequence is the major factor determining whether or not a given protein is Nt-acetylated. In humans, six different NATs, denoted NatA-NatF, are identified. In the current study we used N-terminal COFRADIC analysis to define the in vivo substrate specificity of Naa50 (Nat5)/NatE as this has long remained elusive. Three yeast strains were generated; a control strain endogenously expressing yNaa50, a deletion strain depleted of yNaa50 and a strain deleted of yNaa50 but ectopically expressing human Naa50. When comparing the Nt-acetylation status of different N-termini in the control strain with the deletion strain, a reduction in Nt-acetylation for several yeast proteins was observed. To our surprise, these substrates were not of the predicted NatE-type substrates, but rather canonical NatA substrates. Further, ectopic expression of hNaa50 mainly resulted in the Nt-acetylation of a selected class of otherwise Nt-free yeast N-termini besides increasing the degree of Nt-acetylation of several other yeast proteins, and as such (partially) complementing those N-termini displaying reduced Nt-acetylation upon yNAA50-deletion. The preferred substrates of hNaa50 were predominantly Met-starting N-termini including Met-Lys-, Met-Val-, Met-Ala-, Met-Tyr-, Met-Phe-, Met-Leu-, Met-Ser- and Met-Thr, and highly overlapped with the previously identified human Naa60/NatF substrate specificity profile. Identification of several hNaa50 substrates with a small amino acid in the second position also revealed a potential interplay between the NATs and methionine aminopeptidases (MetAPs). The initiator Methionine (iMet) is normally cleaved off by MetAPs when the second amino acid is small, but our in vitro data suggest that in contrast to a free iMet, an Nt-acetylated iMet is not hydrolyzed by MetAPs. Thus, Naa50-mediated Nt-acetylation may potentially act as a mechanism to retain the iMet of proteins with a small amino acid at the second position that normally would be hydrolyzed.