Localization and targeting of SCG10 to the trans‐Golgi apparatus and growth cone vesicles

Lütjens, Robert; Igarashi, Michihiro; Pellier, Véronique; Blasey, Horst; Paolo, Gilbert Di; Ruchti, Evelyne; Pfulg, Claudine; Staple, Julie; Catsicas, Stefan; Grenningloh, Gabriele

doi:10.1046/j.1460-9568.2000.00112.x

Cited by 76 publications

(82 citation statements)

References 35 publications

(54 reference statements)

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“…In order to identify essential motifs involved in this specific targeting, we examined the subcellular distribution of various subdomains derived from domain A of SCG10 fused with GFP. We show that the Golgi localization of SCG10 results from the cooperation of two motifs: a membrane-anchoring palmitoylation motif and a of growth cones (Stein et al, 1988;Di Paolo et al, 1997a;Gavet et al, 1998;Lutjens et al, 2000;Gavet et al, 2002). SCG10 was also shown to associate with lipid rafts (Maekawa et al, 2001).…”

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confidence: 63%

“…Stathmin-related proteins are peripheral membrane-bound proteins localized to the Golgi complex and on vesicular structures present in the cell body, along neurites, and concentrated within the growth cones of neurons (Stein et al, 1988; Di Paolo et al, 1997a;Lutjens et al, 2000;Gavet et al, 2002). Domain A, common to all stathmin family proteins other than stathmin, was shown here to be sufficient to target GFP to the Golgi complex and vesicles in primary hippocampal neurons, but also in HeLa and MDCK cells, in agreement with previous studies (Di Paolo et al, 1997b;Lutjens et al, 2000).…”

Section: Discussionmentioning

confidence: 99%

“…More generally we provide new information on essential mechanisms of functional protein subcellular targeting. (Stein et al, 1988; Di Paolo et al, 1997a;Gavet et al, 1998;Lutjens et al, 2000;Gavet et al, 2002). SCG10 was also shown to associate with lipid rafts (Maekawa et al, 2001).…”

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confidence: 98%

“…By contrast, domain A, which is the most Nterminal, is conserved in all stathmin-related proteins (60-70% identity) (Stein et al, 1988;Ozon et al, 1997;Ozon et al, 1998) and has been identified as a key element to target SCG10 to the Golgi complex (Di Paolo et al, 1997b). Indeed, it was shown that not only the deletion of domain A in SCG10 redistributed the protein to the cytosol (Di Paolo et al, 1997b), but also that the addition of this same domain to heterologous proteins was able to target the fusion protein to the Golgi complex in COS-7 cells and in neurons (Di Paolo et al, 1997b;Lutjens et al, 2000). Two palmitoylation sites have been identified in domain A of SCG10, but their mutation did not result in the complete solubilization of the protein (Di Paolo et al., 1997b), suggesting that other signals must be involved in the subcellular localization of stathmin-related proteins to the Golgi complex.…”

mentioning

confidence: 99%

“…By characterizing the subcellular localization mediated by various subdomains derived from domain A of SCG10 in fusion with GFP, we delineated and characterized two targeting motifs acting in a cooperative fashion, with membrane-anchoring and Golgi-specifying activities respectively. Altogether, our results open new perspectives concerning the mechanisms by which stathmin family proteins are targeted to their functional compartments and, more generally, concerning specific determinants for Golgi targeting of peripheral membrane proteins.Stathmin-related proteins are peripheral membrane-bound proteins localized to the Golgi complex and on vesicular structures present in the cell body, along neurites, and concentrated within the growth cones of neurons (Stein et al, 1988; Di Paolo et al, 1997a;Lutjens et al, 2000;Gavet et al, 2002). Domain A, common to all stathmin family proteins other than stathmin, was shown here to be sufficient to target GFP to the Golgi complex and vesicles in primary hippocampal neurons, but also in HeLa and MDCK cells, in agreement with previous studies (Di Paolo et al, 1997b;Lutjens et al, 2000).…”

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confidence: 99%

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Two separate motifs cooperate to target stathmin-related proteins to the Golgi complex

Charbaut

Chauvin

Enslen

et al. 2005

Journal of Cell Science

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newly identified Golgi-specifying sequence. The latter displayed no targeting activity by itself, but retained a Golgi-specifying activity when associated with another membrane-anchoring palmitoylation motif derived from the protein GAP-43. We further identified critical residues for the specific Golgi targeting of domain A. Altogether, our results give new insight into the regulation of the subcellular localization of stathmin family proteins, an important feature of their physiological functions in differentiating and mature neural cells. More generally we provide new information on essential mechanisms of functional protein subcellular targeting. (Stein et al., 1988; Di Paolo et al., 1997a;Gavet et al., 1998;Lutjens et al., 2000;Gavet et al., 2002). SCG10 was also shown to associate with lipid rafts (Maekawa et al., 2001). This specific subcellular distribution has led to proposals that stathmin-related proteins are involved in the local regulation of cellular processes, including microtubule dynamics and possibly others in relation with their other identified partners (Nixon et al., 2002;Liu et al., 2002;Greka et al., 2003). The membrane localization of stathmin-related proteins is achieved by their N-terminal extension composed of one or several specific domains (Fig. 1). Domains A′ and A′′, with unknown functions, are specific to the splice variants RB3, RB3′ and RB3′′. By contrast, domain A, which is the most Nterminal, is conserved in all stathmin-related proteins (60-70% identity) (Stein et al., 1988;Ozon et al., 1997;Ozon et al., 1998) and has been identified as a key element to target SCG10 to the Golgi complex (Di Paolo et al., 1997b). Indeed, it was shown that not only the deletion of domain A in SCG10 redistributed the protein to the cytosol (Di Paolo et al., 1997b), but also that the addition of this same domain to heterologous proteins was able to target the fusion protein to the Golgi complex in COS-7 cells and in neurons (Di Paolo et al., 1997b;Lutjens et al., 2000). Two palmitoylation sites have been identified in domain A of SCG10, but their mutation did not result in the complete solubilization of the protein (Di Paolo et al., 1997b), suggesting that other signals must be involved in the subcellular localization of stathmin-related proteins to the Golgi complex.In this study, we investigated the molecular mechanisms involved in the Golgi targeting of stathmin family proteins. By characterizing the subcellular distribution mediated by various subdomains derived from domain A of SCG10 fused to GFP, we show that it is achieved by the combination of two adjacent motifs, one being responsible for membrane association and the other specifying targeting to the Golgi complex. In addition, we demonstrate that this second motif is able to cooperate with another palmitoylated membrane-anchoring sequence for Golgi targeting and we identified critical residues for its functional integrity. Materials and Methods Plasmid constructsStandard recombinant DNA techniques were carried out as described (Sambroo...

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mentioning

confidence: 63%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 98%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Two separate motifs cooperate to target stathmin-related proteins to the Golgi complex

Charbaut

Chauvin

Enslen

et al. 2005

Journal of Cell Science

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The control of microtubule stability in vitro and in transfected cells by MAP1B and SCG10

Bondallaz

Barbier

Soehrman

et al. 2006

Cell Motil. Cytoskeleton

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In neurons, the regulation of microtubules plays an important role for neurite outgrowth, axonal elongation, and growth cone steering. SCG10 family proteins are the only known neuronal proteins that have a strong destabilizing effect, are highly enriched in growth cones and are thought to play an important role during axonal elongation. MAP1B, a microtubule-stabilizing protein, is found in growth cones as well, therefore it was important to test their effect on microtubules in the presence of both proteins. We used recombinant proteins in microtubule assembly assays and in transfected COS-7 cells to analyze their combined effects in vitro and in living cells, respectively. Individually, both proteins showed their expected activities in microtubule stabilization and destruction respectively. In MAP1B/SCG10 double-transfected cells, MAP1B could not protect microtubules from SCG10-induced disassembly in most cells, in particular not in cells that contained high levels of SCG10. This suggests that SCG10 is more potent to destabilize microtubules than MAP1B to rescue them. In microtubule assembly assays, MAP1B promoted microtubule formation at a ratio of 1 MAP1B per 70 tubulin dimers while a ratio of 1 SCG10 per two tubulin dimers was needed to destroy microtubules. In addition to its known binding to tubulin dimers, SCG10 binds also to purified microtubules in growth cones of dorsal root ganglion neurons in culture. In conclusion, neuronal microtubules are regulated by antagonistic effects of MAP1B and SCG10 and a fine tuning of the balance of these proteins may be critical for the regulation of microtubule dynamics in growth cones.

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Tau interacts with Golgi membranes and mediates their association with microtubules

Farah

Perreault

Liazoghli

et al. 2006

Cell Motil. Cytoskeleton

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Tau, a microtubule-associated protein enriched in the axon, is known to stabilize and promote the formation of microtubules during axonal outgrowth. Several studies have reported that tau was associated with membranes. In the present study, we further characterized the interaction of tau with membranous elements by examining its distribution in subfractions enriched in either Golgi or endoplasmic reticulum membranes isolated from rat brain. A subfraction enriched with markers of the medial Golgi compartment, MG160 and mannosidase II, presented a high tau content indicating that tau was associated with these membranes. Electron microscope morphometry confirmed the enrichment of this subfraction with Golgi membranes. Double-immunogold labeling experiments conducted on this subfraction showed the direct association of tau with vesicles labeled with either an antibody directed against MG160 or TGN38. The association of tau with the Golgi membranes was further confirmed by immunoisolating Golgi membranes with an anti-tau antibody. Immunogold labeling confirmed the presence of tau on the Golgi membranes in neurons in vivo. Overexpression of human tau in primary hippocampal neurons induced the formation of large Golgi vesicles that were found in close vicinity to tau-containing microtubules. This suggested that tau could serve as a link between Golgi membranes and microtubules. Such role for tau was demonstrated in an in vitro reconstitution assay. Finally, our results showed that some tau isoforms present in the Golgi subfraction were phosphorylated at the sites recognized by the phosphorylation-dependent antibodies PHF-1 and AT-8.

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Localization and targeting of SCG10 to the trans‐Golgi apparatus and growth cone vesicles

Cited by 76 publications

References 35 publications

Two separate motifs cooperate to target stathmin-related proteins to the Golgi complex

Two separate motifs cooperate to target stathmin-related proteins to the Golgi complex

The control of microtubule stability in vitro and in transfected cells by MAP1B and SCG10

Tau interacts with Golgi membranes and mediates their association with microtubules

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