Abstract
N-terminal acetylation is an irreversible protein modification that primarily occurs co-translationally, and is catalyzed by a highly conserved family of N-terminal acetyltransferases (NATs). The NatC complex (NAA30–NAA35–NAA38) is a major NAT enzyme, which was first described in yeast and estimated to N-terminally acetylate ∼20% of the proteome. The activity of NatC is crucial for the correct functioning of its substrates, which include translocation to the Golgi apparatus, the inner nuclear membrane as well as proper mitochondrial function. We show in comparative viability and growth assays that yeast cells lacking MAK3/NAA30 grow poorly in non-fermentable carbon sources and other stress conditions. By using two different experimental approaches and two yeast strains, we show that liquid growth assays are the method of choice when analyzing subtle growth defects, keeping loss of information to a minimum. We further demonstrate that human NAA30 can functionally replace yeast MAK3/NAA30. However, this depends on the genetic background of the yeast strain. These findings indicate that the function of MAK3/NAA30 is evolutionarily conserved from yeast to human. Our yeast system provides a powerful approach to study potential human NAA30 variants using a high-throughput liquid growth assay with various stress conditions.
Highlights
N-terminal acetylation is a common protein modification defined by the addition of an acetyl group to the start of a protein [1,2,3]
To study the impact of possible NatC variants in vivo, we have explored the consequences of deleting MAK3, the catalytic subunit of the NatC complex and the yeast homolog of human NAA30
Our results indicate that this ‘double stress’ caused by sodium acetate potentiates the effects caused by NaCl, and results in a growth defect that cannot be fully rescued by overexpression of human NAA30
Summary
N-terminal acetylation is a common protein modification defined by the addition of an acetyl group to the start of a protein [1,2,3]. Current estimates suggest that ∼80% of human proteins and ∼50–70% of yeast proteins are to varying degrees N-terminally acetylated [1,4,5,6]. From a clinical perspective, dysregulated N-terminal acetylation can be causative factor of intellectual disability, autism spectrum disorder, and congenital heart disease [8,9,10,11]. These clinical manifestations may arise from defects in the developmental programming of organ specifications. A group of enzymes called N-terminal acetyltransferases (NATs) catalyzes N-terminal acetylation, either during or after protein synthesis [12]. The NAT enzymes play essential roles in cell proliferation, apoptosis, protein trafficking, and several other biological processes [12,28]
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