Golgin45-Syntaxin5 Interaction Contributes to Structural Integrity of the Golgi Stack

Abstract

The unique stacked morphology of the Golgi apparatus had been a topic of intense investigation among the cell biologists over the years. We had previously shown that the two Golgin tethers (GM130 and Golgin45) could, to a large degree, functionally substitute for GRASP-type Golgi stacking proteins to sustain normal Golgi morphology and function in GRASP65/55-double depleted HeLa cells. However, compared to well-studied GM130, the exact role of Golgin45 in Golgi structure remains poorly understood. In this study, we aimed to further characterize the functional role of Golgin45 in Golgi structure and identified Golgin45 as a novel Syntaxin5-binding protein. Based primarily on a sequence homology between Golgin45 and GM130, we found that a leucine zipper-like motif in the central coiled-coil region of Golgin45 appears to serve as a Syntaxin5 binding domain. Mutagenesis study of this conserved domain in Golgin45 showed that a point mutation (D171A) can abrogate the interaction between Golgin45 and Syntaxin5 in pull-down assays using recombinant proteins, whereas this mutant Golgin45 binding to Rab2-GTP was unaffected in vitro . Strikingly, exogenous expression of this Syntaxin5 binding deficient mutant (D171A) of Golgin45 in HeLa cells resulted in frequent intercisternal fusion among neighboring Golgi cisterna, as readily observed by EM and EM tomography. Further, double depletion of the two Syntaxin5-binding Golgin tethers also led to significant intercisternal fusion, while double depletion of GRASP65/55 didn’t lead to this phenotype. These results suggest that certain tether-SNARE interaction within Golgi stack may play a role in inhibiting intercisternal fusion among neighboring cisternae, thereby contributing to structural integrity of the Golgi stack.

A protumorigenic secretory pathway activated by p53 deficiency in lung adenocarcinoma

“…Based on spectral counts of peptides identified by LC-MS analysis (Supplemental Figure 7C and Supplemental Table 2), G45 was the top hit among 88 proteins identified, an indication that G45 was a physiological binding partner of G55 in our model. G45 is a coiled-coil domain protein that facilitates SNARE-mediated vesicle fusion with Golgi membranes (12,27). High G45 levels were correlated with shorter survival in the TCGA LUAD cohort (Figure 5F), and G45 is a known driver of vesicle biogenesis in the Golgi ( 12), but the mechanism by which G45 drives vesicle biogenesis and the biological significance of G45dependent secretion in cancer remain unclear.…”

Loss of Profilin3 Impairs Spermiogenesis by Affecting Acrosome Biogenesis, Autophagy, Manchette Development and Mitochondrial Organization

“…In order to investigate the underlying mechanism of impaired acrosome biogenesis in Pfn3 -deficient mice, we performed immunofluorescence (IF) staining by using GM130 and TGN46 antibodies, markers for cis- and trans- Golgi network, respectively. GM130 plays a crucial role in vesicle tethering, fusion, and maintaining cis -Golgi structural integrity ( Tiwari et al, 2019 ), while TGN46 is important for formation of exocytic vesicles and secretion from the trans- part of the Golgi network ( Huang et al, 2019 ). IF revealed cis- and trans- Golgi predominantly concentrated at one pole of the Pfn3 +/+ ( Figures 6A,D ) and Pfn3 +/– ( Figures 6B,E ) spermatids, whereas the Pfn3 –/– spermatids showed defects and disorganization in cis- ( Figure 6C ) and trans -Golgi network ( Figure 6F ).…”

Tankyrase-1-mediated degradation of Golgin45 regulates glycosyltransferase trafficking and protein glycosylation in Rab2-GTP-dependent manner

“…We have previously shown that Golgin45 and GM130 play a partly redundant role in Golgi structure maintenance 12 . Although these two proteins appear to be unrelated phylogenetically, they share similar arrangement for binding partner interaction, such as GRASPs and Syntaxin5 8 , 10 . As N-terminal domain of GM130 mediates its binding to another vesicle-binding Golgin tether, p115, in a cell cycle-dependent manner via Cdc2 17 , we reasoned that Golgin45-TNKS1 interaction could be also subjected to Cdc2-dependent regulation.…”

“…On the other hand, Golgin45 is a Golgi structural protein that has been shown to directly interact with at least four Golgi-associated proteins, including ACBD3, GRASP55, Syntaxin5 and a small GTPase, Rab2-GTP 8 – 10 . Golgin45 is also called BLZF1, Basic Leucine Zipper Nuclear Factor 1, as it is known to function as a nuclear cofactor 11 .…”

“…ACBD3 recruits TBC1D22, a putative Rab33b-GTPase activating protein (GAP), to the large multi-protein complex, potentially leading to a unique membrane micro-domain organization at the cis- and medial -Golgi 9 . In addition, we have recently shown that Golgin45-Syntaxin5 interaction contributes to the structural integrity of the Golgi stack by inhibiting intercisternal fusion between adjacent Golgi cisterna 10 .…”