The main finding in this study is that although monomeric MANI-FM localizes to cis-medial Golgi cisternae and is present in peri-Golgi vesicles and tubules, the same construct, upon shifting to a polymeric state, moves from the rims to the center of the cisternae, disappears from peri-Golgi carriers, and moves from the cis-Golgi to the trans-Golgi cisternae at a rate reported for cisternal progression (Bonfanti et al., 1998
). The simplest interpretation of these data are that monomeric MANI-FM normally recycles backward in the stack in lockstep with the progression of the cisternae (and of the cargo) via retrograde transport intermediates to maintain its cis location and that, when its recycling is inhibited by polymerization, it progresses through the stack along with the cisternal flow. Notably, monomeric MANI-FM mimics the localization, and hence presumably the behavior, of the corresponding endogenous resident enzyme. Alternative interpretations of the forward progression of MANI-FM polymers and, in particular, interpretations based on models by which cargo moves across stable cisternae via anterograde carriers must assume that polymerization induces MANI-FM to move forward in the stack by promoting its entry into anterograde transport intermediates. However, this is just the opposite of what is observed experimentally: polymerization induces a nearly complete disappearance of MANI-FM from all types of peri-Golgi carriers (both vesicles and tubules), as well as a shift of this construct from the rims to the center of the cisternae, a location that is not suitable for export. Another recently proposed stable cisternae model proposes that cargo moves forward by shifting across heterologous connected cisternae in adjacent stacks of the ribbon (Pfeffer, 2010
). Also, this model is inconsistent with our results, which were obtained using physically separated ministacks. The findings in this study are therefore consistent with the cisternal progression maturation mechanism and not with stable cisternae models.
A further line of evidence in favor of cisternal maturation is that when MANI-FM is depolymerized in the trans-cisterna it rapidly reenters cisternal rims as well as vesicles/tubules and shifts back toward its cis-Golgi location. These data represent evidence for the rapid recycling of Golgi resident enzymes through the stack in mammals. Notably, a Golgi enzyme, GAlNacT2, has been recently reported to cycle rapidly between the Golgi and the intermediate compartment in mammals (Jarvela and Linstedt, 2012
). This finding does not relate to the progression/maturation of the cisternae through the stack. Rather, it reflects the formation of new cis-cisternae from the intermediate compartment (Jarvela and Linstedt, 2012
Regarding the mechanism of recycling, the MANI-FM–containing carriers that proliferate upon MANI-FM depolymerization appear to be COP-I–coated tubules and vesicles. Discordant conclusions have been proposed in the past regarding the question of whether vesicles or tubules (which might be dissociated or connected with adjacent cisternae) are involved in recycling (Orci et al., 2000
; Lanoix et al., 2001
; Martinez-Menárguez et al., 2001
; Kweon et al., 2004
; Trucco et al., 2004
). Our current data, and the recent proposal that Golgi vesicles and tubules are strongly mechanistically related (Yang et al., 2011
), might explain the previous discrepancies.
Do these findings show that cisternal maturation is the only mode of intra-Golgi transport? As is often stated in the literature, transport mechanisms are not mutually exclusive (Emr et al., 2009
). Thus, although our data provide evidence for cisternal maturation, they do not exclude that other transport principles might coexist with the maturation process in the Golgi complex, such as vesicular trafficking (Rothman and Wieland, 1996
; Malsam et al., 2005
) or diffusional trafficking via continuities (Trucco et al., 2004
), with the prevailing mechanism perhaps depending on the type of cargo being transported, the pathophysiological conditions, the cell type, and/or the trafficking step being examined. Examples of the eclectic nature of the transport apparatus have been provided by studies of the endocytic pathway, where different transport principles have been shown to coexist (Pryor and Luzio, 2009
; De Matteis and Luini, 2011
). Membrane trafficking has central roles in many cellular processes (Mellman and Warren, 2000
; De Matteis and Luini, 2011
). A better understanding of the full repertoire of eukaryotic trafficking principles will provide valuable insight into key aspects of cell physiology and pathology.