Glycoproteins of the lysosomal membrane.

Lewis, Elfed; Green, S A; Marsh, Mark; Vihko, Pirkko; Helenius, Ari; Mellman, Ira

doi:10.1083/jcb.100.6.1839

Cited by 281 publications

(178 citation statements)

References 43 publications

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“…Our results show a significant overlap of EEA1 and RasGEF1b, suggesting that a main localization of RasGEF1b occurs at the level of early endosomes (Figure 4c). Further, we observed no colocalization of ectopic RasGEF1b and LAMP-1, a lysosome marker 17 (Supplementary Figure 5), suggesting that RasGEF1b may shuttle between early and late endosomes.…”

Section: Cloning Expression and Subcellular Localization Of Rasgef1mentioning

confidence: 83%

Early endosome localization and activity of RasGEF1b, a toll-like receptor-inducible Ras guanine-nucleotide exchange factor

et al. 2010

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Guanine-nucleotide exchange factors (GEFs) stimulate the intrinsic GDP/GTP exchange activity of Ras and promote the formation of active Ras-GTP, which in turn controls diverse signalling networks important for the regulation of cell proliferation, survival, differentiation, vesicular trafficking, and gene expression. RasGEF1b is a GEF, whose expression is induced in macrophages on stimulation with toll-like receptor (TLR) agonists. Here, we showed that in vitro RasGEF1b expression by macrophages is mostly induced by TLR3 (poly I:C) and TLR4 (lipopolysaccharyde) through the MyD88-independent pathway. In vivo infection with the protozoan parasites Trypanosoma cruzi and Plasmodium chabaudi induced RasGEF1b in an MyD88-, TRIF-, and IFN-g-dependent manner. Ectopically expressed RasGEF1b was found, mostly, in the heavy membrane fraction of HEK 293T, and by confocal microscopy, it was found to be located at early endosomes. Computational modelling of the RasGEF1b-Ras interaction revealed that RasGEF1b interacts with the binding domain site of Ras, a critical region for interacting with GEFs involved in the activation of Ras-Raf-MEK-ERK pathway. More important, RasGEF1b was found to be closely associated with Ras in live cells and to trigger Ras activity. Altogether, these results indicate that on TLR activation, RasGEF1b may trigger Ras-like proteins and regulate specific biological activities described for this subtype of GTPases.

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Section: Cloning Expression and Subcellular Localization Of Rasgef1mentioning

confidence: 83%

Early endosome localization and activity of RasGEF1b, a toll-like receptor-inducible Ras guanine-nucleotide exchange factor

et al. 2010

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“…a, Gold particles are associated with small vesicles which interact with the outer melanosomal membrane (arrows) and also with Golgi cisternae which possibly give rise to them (panel a, insert (Figure 9). This provides evidence that the antigen contains 3 to 5 N-linked oligosaccharide chains, based on a reported 2-4 kDa shift per N-linked saccharide moiety on SDS-PAGE gels (Lewis et al, 1985). Early in our studies several factors had pointed towards the possibility of the HMSA-5 antigen being the enzyme tyrosinase.…”

Section: Pulse-chase Experimentsmentioning

confidence: 95%

A murine monoclonal antibody, MoAb HMSA-5, against a melanosomal component highly expressed in early stages, and common to normal and neoplastic melanocytes

Der

Dixon²,

Jimbow³

et al. 1993

Br J Cancer

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Summary The melanosome is a secretory organelle unique to the melanocyte and its neoplastic counterpart, malignant melanoma. The synthesis and assembly of these intracytoplasmic organelles is not yet fully understood. We have developed a murine monoclonal antibody (MoAb) against melanosomes isolated from human melanocytes (newborn foreskin) cultured in the presence of 12-0 tetradecanoyl phorbol-13-acetate (TPA). This MoAb, designated HMSA-5 (Human Melanosome-Specific Antigen-5) (IgG1), recognised a cytoplasmic antigen in both normal human melanocytes and neoplastic cells, such as common and dysplastic melanocytic nevi, and malignant melanoma. None of the carcinoma or sarcoma specimens tested showed positive reactivity with MoAb HMSA-5. Under immunoelectron microscopy, immuno-gold deposition was seen on microvesicles associated with melanosomes, and a portion of the ER-Golgi complexes. Radioimmunoprecipitation analysis showed that the HMSA-5 reactive antigen was a glycoprotein of M, 69 to 73 kDa. A pulse-chase time course study showed that the amount of antigen detected by MoAb HMSA-5 decreased over a 24 h period without significant expression on the cell surface, or corresponding appearance of the antigen in the culture supernatant. This glycoprotein appears to play a role in the early stages of melanosomal development, and the HMSA-5 reactive epitope may be lost during subsequent maturation processes. Importantly, HMSA-5 can be identified in all forms of human melanocytes, hence it can be considered a new common melanocytic marker even on routine paraffin sections.

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“…If all four sites for N-linked glycosylation are used, then the monomer molecular weight is expected to be ϳ40 -44 kDa assuming a mass of ϳ3-4 kDa for each sugar side chain (28). To test the contribution of N-linked carbohydrate, we treated lysates containing baculo DϩC with endoglycosidase F. As seen in Fig.…”

Section: Expression Of Disintegrin-like Domain-containing Constructs mentioning

confidence: 99%

Sequence-specific Interaction between the Disintegrin Domain of Mouse ADAM 2 (Fertilin β) and Murine Eggs

Bigler¹,

Takahashi²,

Chen³

et al. 2000

Journal of Biological Chemistry

105

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Little is yet known about the biological and biochemical properties of the disintegrin-like domains of ADAM (a disintegrin and metalloprotease) proteins. Mouse ADAM 2 (mADAM 2; fertilin ␤) is a sperm surface protein involved in murine fertilization. We produced recombinant proteins containing the disintegrin-like domain of mADAM 2 in both insect cells and in bacteria. The protein produced in insect cells (baculo D؉C) contained a signal sequence followed by the disintegrin-like and cysteine-rich domains; it was purified from the medium of recombinant baculovirus-infected cells. A bacterial construct containing the disintegrin-like domain was produced in Escherichia coli as a glutathione S-transferase chimera. Baculo D؉C, as well as the D domain of the bacterial construct (released with thrombin), bound to the microvillar surface of murine eggs. Using concentrations in the range of 1 to 5 M, both recombinant proteins strongly inhibited sperm-egg binding and fusion; the baculovirus-produced protein exhibited a somewhat greater extent of inhibition (ϳ75 versus ϳ55% maximal inhibition). Substitution of alanine for each of the five charged residues within the disintegrin loop of mADAM 2 revealed a critical importance for the aspartic acid at position nine. Binding of both recombinant proteins to the egg was inhibited by the function blocking anti-␣ 6 monoclonal antibody, GoH3, but not by a nonfunctionblocking anti-␣ 6 monoclonal antibody. Binding was also inhibited by a peptide analogue of, and with an antibody against, the disintegrin loop of mADAM 2. ADAMs1 are a large group of type I integral membrane glycoproteins that contain a disintegrin and a metalloprotease domain (1, 2). Following their metalloprotease and disintegrinlike domains, they contain a cysteine-rich domain, an EGF repeat, a transmembrane domain, and a cytoplasmic tail. Their closest relatives are the P-III snake venom metalloproteases (SVMPs), which are secreted proteins that contain a disintegrin-like and a metalloprotease domain as well as a cysteinerich domain. P-II SVMPs contain metalloprotease and disintegrin domains but lack the cysteine-rich (and other) domains.The disintegrin domains of the P-II SVMPs have a 13-amino acid loop containing, at their tips, sequences such as RGD (kistrin and echistatin), KGD (barbourin), and MVD (atrolysin E). Several P-II snake disintegrins have been shown to bind to the platelet integrin, ␣ IIb ␤ 3, thereby preventing binding of fibrinogen and inhibiting platelet aggregation. Inhibition of platelet aggregation accounts, in part, for the severe hemorrhagic response in snakebite victims. The disintegrin-like domains of P-III SVMPs differ from their P-II counterparts in having two additional cysteine residues and a 14-residue predicted loop that aligns with the 13-amino acid "RGD" loop found in the P-II disintegrins. One of the additional cysteine residues is near the center of the disintegrin loop. The disintegrin-like domain of the P-III SVMP atrolysin A has been characterized. Atrolysin A purified from snake ...

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Glycoproteins of the lysosomal membrane.

Cited by 281 publications

References 43 publications

Early endosome localization and activity of RasGEF1b, a toll-like receptor-inducible Ras guanine-nucleotide exchange factor

Early endosome localization and activity of RasGEF1b, a toll-like receptor-inducible Ras guanine-nucleotide exchange factor

A murine monoclonal antibody, MoAb HMSA-5, against a melanosomal component highly expressed in early stages, and common to normal and neoplastic melanocytes

Sequence-specific Interaction between the Disintegrin Domain of Mouse ADAM 2 (Fertilin β) and Murine Eggs

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