A role for human N-alpha acetyltransferase 30 (Naa30) in maintaining mitochondrial integrity

N-terminal acetylation (Nt-acetylation) by N-terminal acetyltransferases (NATs) is one of the most common protein modifications in eukaryotes. The NatC complex represents one of three major NATs of which the substrate profile remains largely unexplored. Here, we defined the in vivo human NatC Nt-acetylome on a proteome-wide scale by combining knockdown of its catalytic subunit Naa30 with positional proteomics. We identified 46 human NatC substrates, expanding our current knowledge on the substrate repertoire of NatC which now includes proteins harboring Met-Leu, Met-Ile, Met-Phe, Met-Trp, Met-Val, Met-Met, Met-His and Met-Lys N termini. Upon Naa30 depletion the expression levels of several organellar proteins were found reduced, in particular mitochondrial proteins, some of which were found to be NatC substrates. Interestingly, knockdown of Naa30 induced the loss of mitochondrial membrane potential and fragmentation of mitochondria. In conclusion, NatC N-tacetylates a large variety of proteins and is essential for mitochondrial integrity and function

LEDGF/p75 has increased expression in blasts from chemotherapy-resistant human acute myelogenic leukemia patients and protects leukemia cells from apoptosis in vitro

<p>Abstract</p> <p>Background</p> <p>Relapse due to chemoresistant residual disease is a major cause of death in acute myelogenous leukemia (AML). The present study was undertaken to elucidate the molecular mechanisms of chemoresistance by comparing differential gene expression in blasts from patients with resistant relapsing AML and chemosensitive AML.</p> <p>Results</p> <p>About 20 genes were identified as preferentially expressed in blasts pooled from patients with resistant disease, as compared to chemosensitive AML blasts, based on differential gene expression screening. Half of these genes encoded proteins related to protein translation, of these a novel protein related to the ribosomal stalk protein P0. Other upregulated mRNAs coded for cytochrome C oxidase III, the transcription factors ERF-2/TIS11d, and the p75 and p52 splice variants of Lens Epithelial Derived Growth Factor (LEDGF). Analysis of blasts from single patients disclosed that LEDGF/p75 was the most consistently upregulated mRNA in resistant AML. Transfection experiments demonstrated that LEDGF/p75 and p52b antagonized daunorubicin-induced and cAMP-induced apoptosis in an AML cell line. Also HEK-293 cells were protected against daunorubicin by LEDGF/p75 and p52b, whereas LEDGF/p52 splice variants lacking exon 6 had proapoptotic effects. Interestingly, full length LEDGF/p75 protected against truncated pro-apoptotic LEDGF/p75.</p> <p>Conclusion</p> <p>Our results provide evidence for an association between the overexpression of genes encoding survival proteins like LEDGF/p75 and chemo-resistance in acute myelogenous leukemia. LEDGF/p75 has previously not been shown to protect against chemotherapy, and is a potential drug target in AML.</p

Optimized bisubstrate inhibitors for the actin N-terminal acetyltransferase NAA80

Acetylation of protein N-termini is one of the most common protein modifications in the eukaryotic cell and is catalyzed by the N-terminal acetyltransferase family of enzymes. The N-terminal acetyltransferase NAA80 is expressed in the animal kingdom and was recently found to specifically N-terminally acetylate actin, which is the main component of the microfilament system. This unique animal cell actin processing is essential for the maintenance of cell integrity and motility. Actin is the only known substrate of NAA80, thus potent inhibitors of NAA80 could prove as important tool compounds to study the crucial roles of actin and how NAA80 regulates this by N-terminal acetylation. Herein we describe a systematic study toward optimizing the peptide part of a bisubstrate-based NAA80 inhibitor comprising of coenzyme A conjugated onto the N-terminus of a tetrapeptide amide via an acetyl linker. By testing various combinations of Asp and Glu which are found at the N-termini of β- and γ-actin, respectively, CoA-Ac-EDDI-NH2 was identified as the best inhibitor with an IC50 value of 120 nM

NAA10 dysfunction with normal NatA-complex activity in a girl with non-syndromic ID and a de novo NAA10 p.(V111G) variant – a case report

Abstract Background The NAA10-NAA15 (NatA) protein complex is an N-terminal acetyltransferase responsible for acetylating ~ 40% of eukaryotic proteins. In recent years, NAA10 variants have been found in patients with an X-linked developmental disorder called Ogden syndrome in its most severe form and, in other familial or de novo cases, with variable degrees of syndromic intellectual disability (ID) affecting both sexes. Case presentation Here we report and functionally characterize a novel and de novo NAA10 (NM_003491.3) c.332 T > G p.(V111G) missense variant, that was detected by trio-based whole exome sequencing in an 11 year old girl with mild/moderate non-syndromic intellectual disability. She had delayed motor and language development, but normal behavior without autistic traits. Her blood leukocyte X-inactivation pattern was within normal range (80/20). Functional characterization of NAA10-V111G by cycloheximide chase experiments suggests that NAA10-V111G has a reduced stability compared to NAA10-WT, and in vitro acetylation assays revealed a reduced enzymatic activity of monomeric NAA10-V111G but not for NAA10-V111G in complex with NAA15 (NatA enzymatic activity). Conclusions We show that NAA10-V111G has a reduced stability and monomeric catalytic activity, while NatA function remains unaltered. This is the first example of isolated NAA10 dysfunction in a case of ID, suggesting that the syndromic cases may also require a degree of compromised NatA function