Identification and characterisation of the cryptic Golgi Apparatus in Naegleria gruberi

Although the Golgi apparatus has a conserved morphology of flattened stacked cisternae in the vast majority of eukaryotes, the organelle has lost the stacked organization in several eukaryotic lineages raising the question of what range of morphologies is possible for the Golgi. In order to understand this range of organellar diversity, it is necessary to characterise the Golgi in many different lineages. Here we identify the Golgi apparatus in Naegleria, the first description of an unstacked Golgi organelle in a non-parasitic eukaryote, other than fungi. We provide a comprehensive list of Golgi-associated membrane trafficking genes encoded in two separate species of Naegleria and transcriptomic support to show that nearly all are expressed in mouse-passaged N. fowleri cells. We then study distribution of the Golgi marker NgCOPB by fluorescence, identifying membranous structures that can be disrupted by Brefeldin A treatment consistent with Golgi localisation. Confocal and immuno-electron microscopy revealed that NgCOPB is localized to membranous structures consistent with tubules. Our data not only identify the Golgi organelle for the first time in this major eukaryotic lineage, but also provide the rare example of a tubular form of the organelle representing an important sampling point for the comparative understanding of Golgi organellar diversity.