The Interaction of Two Tethering Factors, p115 and COG complex, is Required for Golgi Integrity

Department of Cell Biology, Fukuoka University School of Medicine, Jonan-ku, Fukuoka 814-0180, Japan.
Traffic (Impact Factor: 4.35). 04/2007; 8(3):270-84. DOI: 10.1111/j.1600-0854.2006.00530.x
Source: PubMed


The vesicle-tethering protein p115 functions in endoplasmic reticulum-Golgi trafficking. We explored the function of homologous region 2 (HR2) of the p115 head domain that is highly homologous with the yeast counterpart, Uso1p. By expression of p115 mutants in p115 knockdown (KD) cells, we found that deletion of HR2 caused an irregular assembly of the Golgi, which consisted of a cluster of mini-stacked Golgi fragments, and gathered around microtubule-organizing center in a microtubule-dependent manner. Protein interaction analyses revealed that p115 HR2 interacted with Cog2, a subunit of the conserved oligomeric Golgi (COG) complex that is known another putative cis-Golgi vesicle-tethering factor. The interaction between p115 and Cog2 was found to be essential for Golgi ribbon reformation after the disruption of the ribbon by p115 KD or brefeldin A treatment and recovery by re-expression of p115 or drug wash out, respectively. The interaction occurred only in interphase cells and not in mitotic cells. These results strongly suggested that p115 plays an important role in the biogenesis and maintenance of the Golgi by interacting with the COG complex on the cis-Golgi in vesicular trafficking.

Download full-text


Available from: Shin-ichiro Yoshimura, Oct 16, 2014
  • Source
    • "VOLUME 288 • NUMBER 6 • FEBRUARY 8 , 2013 Immunoprecipitation—GFP - TMF expression plasmids were transfected into HEK - 293 cells stably expressing HA - tagged Cog1 using the GeneCellin transfection reagent ( BioCellChal - lenge ) . For COG - COPI co - immunoprecipitations YFP - ␤COP and HA - Cog2 ( Sohda et al . , 2007 ) were co - transfected into HeLa cells . After 36 – 48 h , cells were washed with IP buffer ( 20 mM Hepes , 110 mM KOAc , 1 mM MgCl 2 , pH 7 . 4 ) then scraped into 1 ml of IP lysis buffer ( IP buffer with 1% Triton X - 100 ) . All steps were performed at room temperature . The cell slurry was rotated for 30 min prior to 30 min of cent"
    [Show abstract] [Hide abstract]
    ABSTRACT: Protein sorting between eukaryotic compartments requires vesicular transport, wherein tethering provides the first contact between vesicle and target membranes. Here we map and start to functionally analyze the interaction network of the conserved oligomeric Golgi (COG) complex that mediates retrograde tethering at the Golgi. The interactions of COG subunits with members of transport factor families assign the individual subunits as specific interaction hubs. Functional analysis of selected interactions suggests a mechanistic tethering model. We find that the COG complex interacts with two different Rabs in addition to each end of the golgin “TATA element modulatory factor” (TMF). This allows COG to potentially bridge the distance between the distal end of the golgin and the target membrane thereby promoting tighter docking. Concurrently we show that the central portion of TMF can bind to Golgi membranes that are liberated of their COPI cover. This latter interaction could serve to bring vesicle and target membranes into close apposition prior to fusion. A target selection mechanism, in which a hetero-oligomeric tethering factor organizes Rabs and coiled transport factors to enable protein sorting specificity, could be applicable to vesicle targeting throughout eukaryotic cells.
    Journal of Biological Chemistry 12/2012; 288(6). DOI:10.1074/jbc.M112.426767 · 4.57 Impact Factor
  • Source
    • "Until now, only the H1, H2 and CC1 have been implicated in p115 function (An et al., 2009; Guo et al., 2008; Puthenveedu and Linstedt, 2004; Sohda et al., 2007). The role of the AD is controversial, with one study suggesting that it is required for Fig. 8. Interactions of mutant p115 with cellular proteins. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The tethering factor p115 (known as Uso1p in yeast) has been shown to facilitate Golgi biogenesis and membrane traffic in cells in culture. However, the role of p115 within an intact animal is largely unknown. Here, we document that depletion of p115 by using RNA interference (RNAi) in C. elegans causes accumulation of the 170 kD soluble yolk protein (YP170) in the body cavity and retention of the yolk receptor RME-2 in the ER and the Golgi within oocytes. Structure-function analyses of p115 have identified two homology regions (H1 and H2) within the N-terminal globular head and the coiled-coil 1 (CC1) domain as essential for p115 function. We identify a new C-terminal domain of p115 as necessary for Golgi ribbon formation and cargo trafficking. We show that p115 mutants that lack the fourth CC domain (CC4) act in a dominant-negative manner to disrupt Golgi and prevent cargo trafficking in cells containing endogenous p115. Furthermore, using RNAi of p115 and the subsequent transfection with p115 deletion mutants, we show that CC4 is necessary for Golgi ribbon formation and membrane trafficking in cells depleted of endogenous p115. p115 has been shown to bind a subset of ER-Golgi SNAREs through CC1 and CC4 domains (Shorter et al., 2002). Our findings show that CC4 is required for p115 function, and suggest that both the CC1 and the CC4 SNARE-binding motifs participate in p115-mediated membrane tethering.
    Journal of Cell Science 02/2012; 125(Pt 8):1896-909. DOI:10.1242/jcs.090571 · 5.43 Impact Factor
  • Source
    • "This suggests the involvement of a two-phased tethering mechanism: a first long range in which golgins locate and loosely attach a vesicle to the membrane , after which the COG complex, or another multimeric tethering complex, is recruited for intimate tethering and induction of fusion via assembly of the SNARE complex (Figure 3A). This observation was supported by the identification of a direct interaction of COG2 with p115, another protein of the golgin family (Sohda et al. 2007). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Protein glycosylation is one of the major biosynthetic functions occurring in the endoplasmic reticulum and Golgi compartments. It requires an amazing number of enzymes, chaperones, lectins and transporters whose actions delicately secure the fidelity of glycan structures. Over the past 30 years, glycobiologists hammered that glycan structures are not mere decorative elements but serve crucial cellular functions. This becomes dramatically illustrated by a group of mostly severe, inherited human disorders named congenital disorders of glycosylation (CDG). To date, many types of CDG have been defined genetically and most of the time the defects impair the biosynthesis, transfer and remodeling of N-glycans. Recently, the identification of the several types of CDG caused by deficiencies in the conserved oligomeric Golgi (COG) complex, a complex involved in vesicular Golgi trafficking, expanded the field of CDG but also brought novel insights in glycosylation. The molecular mechanisms underlying the complex pathway of N-glycosylation in the Golgi are far from understood. The availability of COG-deficient CDG patients and patients' cells offered a new way to study how COG, and its different subunits, could influence the Golgi N-glycosylation machinery and localization. This review summarizes the recent findings on the implication of COG in Golgi glycosylation. It highlights the need for a dynamic, finely tuned balance between anterograde and retrograde trafficking for the correct localization of Golgi enzymes to assure the stepwise maturation of N-glycan chains.
    Glycobiology 11/2010; 21(7):853-63. DOI:10.1093/glycob/cwq179 · 3.15 Impact Factor
Show more