Abstract
The organization of the Golgi apparatus (GA) is tightly regulated. Golgi stack scattering is observed in cellular processes such as apoptosis and mitosis, and has also been associated with disruption of cellular lipid metabolism and neurodegenerative diseases. Our studies show that depletion of the human N-α-acetyltransferase 30 (hNaa30) induces fragmentation of the Golgi stack in HeLa and CAL-62 cell lines. The GA associated GTPase ADP ribosylation factor related protein 1 (ARFRP1) was previously shown to require N-terminal acetylation for membrane association and based on its N-terminal sequence, it is likely to be a substrate of hNaa30. ARFRP1 is involved in endosome-to-trans-Golgi network (TGN) traffic. We observed that ARFRP1 shifted from a predominantly cis-Golgi and TGN localization to localizing both Golgi and non-Golgi vesicular structures in hNaa30-depleted cells. However, we did not observe loss of membrane association of ARFRP1. We conclude that hNaa30 depletion induces Golgi scattering and induces aberrant ARFRP1 Golgi localization.
Highlights
The structure and functionality of the Golgi apparatus (GA) is maintained by at least four systems: microtubule-associated transport proteins, the actin-associated cytoskeleton, Golgi matrix proteins and proteins involved in targeting and fusion of vesicles such as GTPases and SNARE proteins [1]
Results human N-α-acetyltransferase 30 (hNaa30) knockdown leads to scattering of GA and trans-Golgi network (TGN) in HeLa and anaplasmic thyroid carcinoma cell line (CAL-62) cell lines hNaa30 was depleted by siRNA-mediated knockdown of hNAA30 gene expression
Western blot analysis was routinely performed to confirm knockdown efficiency of the siRNA constructs on hNaa30 protein levels (Figure 1). siRNA knockdown will not give a complete depletion of protein levels, and all results need to be evaluated as consequences of protein reduction rather than complete loss of protein
Summary
The structure and functionality of the Golgi apparatus (GA) is maintained by at least four systems: microtubule-associated transport proteins, the actin-associated cytoskeleton, Golgi matrix proteins and proteins involved in targeting and fusion of vesicles such as GTPases and SNARE proteins [1]. Disruption of any of these systems can lead to changes in GA organization, such as for example GA collapse or disassembly and fragmentation [2,3,4,5]. The Arfs bind to their target membranes through an N-terminal membrane anchor and an N-terminal amphipathic helix. The Arfs contain a glycine in the second position and are subjected to N-myristoylation [12,13]. A subset of the Arfs are not myristoylated, but rather N-terminally acetylated (Nt-acetylated) [14]
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