Promotion of Tyrosinase Folding in Cos 7 Cells by Calnexin

Toyofuku, Kazutomo; Wada, Ikuo; Hirosaki, Kuninori; Park, Jong Sung; Hori, Yoshiaki; Jimbow, Kowichi

doi:10.1093/oxfordjournals.jbchem.a022272

Cited by 68 publications

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“…For example, if glycosylation is prevented or if glucosidases are inhibited, complexes are not detected between CNX or CRT and the ␣ and ␤ subunits of the T cell receptor (51), influenza hemagglutinin (2), vesicular stomatitis virus G glycoprotein (52), ribonuclease B (35), myeloperoxidase (9), cruzipain (53), and tyrosinase (13). However, complexes can readily be detected at normal or reduced levels under conditions of deglycosylation or glucosidase inhibition with the ⑀ and ␦ subunits of the T cell receptor (20,25), P glycoprotein (23), erythrocyte AE1 (54), acid phosphatase (27), major histocompatibility complex class II-␣ and -␤ chains (19), major histocompatibility complex class II invariant chain (24), major histocompatibility complex class I H chain (this study), and human immunodeficiency virus gp160 (10).…”

Section: Calnexin Associates With Many Substrate Proteins When the Fomentioning

confidence: 99%

The Lectin Chaperone Calnexin Utilizes Polypeptide-based Interactions to Associate with Many of Its Substrates in Vivo

Danilczyk¹,

Williams

2001

Journal of Biological Chemistry

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Calnexin and calreticulin are molecular chaperones of the endoplasmic reticulum that bind to newly synthesized glycoproteins in part through a lectin site specific for monoglucosylated (Glc 1 Man 7-9 GlcNAc 2 ) oligosaccharides. In addition to this lectin-oligosaccharide interaction, in vitro studies have demonstrated that calnexin and calreticulin can bind to polypeptide segments of both glycosylated and nonglycosylated proteins. However, the in vivo relevance of this latter interaction has been questioned. We examined whether polypeptidebased interactions occur between calnexin and its substrates in vivo using the glucosidase inhibitor castanospermine or glucosidase-deficient cells to prevent the formation of monoglucosylated oligosaccharides. We show that if care is taken to preserve weak interactions, the block in lectin-oligosaccharide binding leads to the loss of some calnexin-substrate complexes, but many others remain readily detectable. Furthermore, we demonstrate that calnexin is capable of associating in vivo with a substrate that completely lacks Asn-linked oligosaccharides. The binding of calnexin to proteins that lack monoglucosylated oligosaccharides could not be attributed to nonspecific adsorption nor to its inclusion in protein aggregates. We conclude that both lectin-oligosaccharide and polypeptide-based interactions occur between calnexin and diverse proteins in vivo and that the strength of the latter interaction varies substantially between protein substrates. Glycoprotein folding within the endoplasmic reticulum (ER)1 is facilitated in part by the membrane-bound chaperone calnexin (CNX) and its soluble homolog calreticulin (CRT) (1). These proteins are unique among molecular chaperones in that they utilize a lectin site as a means to associate with unfolded glycoproteins (2-4). The lectin site is specific for Glc 1 Man 7-9 GlcNAc 2 oligosaccharides, which exist transiently as intermediates during the processing of Asn-linked glycoproteins. It is a widely held view that the removal and re-addition of the terminal glucose residue of these oligosaccharides, catalyzed, respectively, by the ER enzymes glucosidase II and UDPglucose:glycoprotein glucosyltransferase, regulate cycles of CNX and CRT binding to glycoproteins (5, 6). In this model, CNX and CRT do not function as classical molecular chaperones that prevent aggregation by binding to hydrophobic polypeptide segments. Rather, they are thought to promote folding by recruiting other chaperones and folding enzymes, such as the thiol oxidoreductase ERp57 (7,8), to the glycoprotein substrate.The concept that CNX and CRT associate with glycoproteins solely through lectin-oligosaccharide interactions is based primarily on experiments wherein cultured cells were treated with tunicamycin to block Asn-linked oligosaccharide addition or with the glucosidase I and II inhibitors castanospermine or deoxynojirimycin to prevent the conversion of the Glc 3 Man 9 GlcNAc 2 precursor to the monoglucosylated Glc 1 Man 9 GlcNAc 2 species. Subsequent immunoprec...

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Section: Calnexin Associates With Many Substrate Proteins When the Fomentioning

confidence: 99%

The Lectin Chaperone Calnexin Utilizes Polypeptide-based Interactions to Associate with Many of Its Substrates in Vivo

Danilczyk¹,

Williams

2001

Journal of Biological Chemistry

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“…Tyrp1 in humans (36) and in mice (44) is synthesized as a precursor polypeptide of approximately 55-69 kDa in the endoplasmic reticulum (ER). This nascent polypeptide undergoes rapid post-translation folding via di-sulfide bonds into its initial native conformation assisted by molecular chaperones like calnexin, a lectin chaperone, and BiP, the immunoglobulin heavy chain binding protein (both of which are resident ER chaperones) (45,46). Formation of disulfide bonds during the folding of Tyrp1 is crucial for its transport through both secretory pathways in the ER and its subsequent processing/maturation in the Golgi apparatus.…”

Section: Tyrp1 the Proteinmentioning

confidence: 99%

Tyrp1 and Oculocutaneous Albinism Type 3

Sarangarajan

Boissy

2001

Pigment Cell Research

113

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addition to its role in eumelanin synthesis, Tyrp1 is involved in Tyrosinase-related protein 1 (Tyrp1) is a melanocyte-specific maintaining stability of tyrosinase protein and modulating its gene product involved in eumelanin synthesis. Mutations in the catalytic activity. Tyrp1 is also involved in maintenance of mouse Tyrp1 gene are associated with brown pelage, and in the human TYRP1 gene with oculocutaneous albinism type 3 melanosome ultrastructure and affects melanocyte prolifera-(OCA3). In the murine system, Tyrp1 expresses significant tion and melanocyte cell death. The current review is an dihydroxyindole carboxylic acid oxidase (i.e. DHICA oxattempt to consolidate our understanding of the role of Tyrp1 idase) activity. However, in humans, TYRP1 is enigmatic in in the melanocyte. that despite extensive efforts focused on the study of its Key words: Pigmentation, Tyrosine hydroxylase, Brown/Rufunction, its actual role in the human melanocyte is still fous Albinism, Brown locus, Protein trafficking unclear. There is mounting evidence demonstrating that in Mutations in the genes encoding some of the enzymes and regulatory proteins involved in melanin synthesis result in various forms of oculocutaneous albinism (OCA). OCA1 correlates with mutations in the tyrosinase (TYR) gene (7). Most individuals with OCA1 have completely amelanotic skin, hair and eyes with the inability to tan (i.e. OCA1A). However, some nucleotide lesions of the TYR gene result in a partially functional enzyme so that individuals with this subtype of OCA1 can exhibit moderate levels of skin and hair pigmentation and the ability to tan (i.e. OCA1B). OCA2 correlates with mutations in the P gene (8). Individuals with OCA2 present with minimal to moderate amounts of pigment remaining in the skin, hair and eyes, many of whom can develop pigmented freckles, lentigines and/or nevi with age. OCA3 correlates with mutations in the TYRP1 gene (9). Individuals with OCA3 present with minimal pigment reduction in the skin, hair and eyes. This form of albinism was previously referred to as Rufous and possibly some forms of Brown albinism.

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“…Endoplasmic reticulum (ER) processing of Tyr requires the presence of the chaperone calnexin, which is thought to increase ER retention time for Tyr Toyofuku et al, 1999). The most common Tyr mutations are associated with ER retention of the protein, presumably the result of misfolding (Berson et al, 2000;Halaban et al, 2000b;Toyofuku et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

Pink-eyed Dilution Protein Controls the Processing of Tyrosinase

Chen

Manga

Orlow

2002

MBoC

128

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The processing of tyrosinase, which catalyzes the limiting reaction in melanin synthesis, was investigated in melan-p1 melanocytes, which are null at the p locus. Endoglycosidase H digestion showed that a significant fraction of tyrosinase was retained in the endoplasmic reticulum. This retention could be rescued either by transfection of melan-p1 cells with an epitope-tagged wild-type p transcript or by treatment with either bafilomycin A1 or ammonium chloride. We found that the endoplasmic reticulum contains a significant amount of p protein, thus supporting a role for p within this compartment. Using immunofluoresence, we showed that most mature full-length tyrosinase in melan-p1 cells was located in the perinuclear area near the Golgi, in contrast to its punctate melanosomal pattern in wild-type melanocytes. Expression of p in melan-p1 cells restored tyrosinase to melanosomes. Triton X-114 phase separation revealed that an increased amount of tyrosinase was proteolyzed in melan-p1 cells compared with wild-type melanocytes. The proteolyzed tyrosinase was no longer membrane bound, but remained enzymatically active and a large proportion was secreted into the culture medium of melan-p1 cells. We conclude that p regulates posttranslational processing of tyrosinase, and hypopigmentation in melan-p1 cells is the result of altered tyrosinase processing and trafficking.

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Promotion of Tyrosinase Folding in Cos 7 Cells by Calnexin

Cited by 68 publications

References 0 publications

The Lectin Chaperone Calnexin Utilizes Polypeptide-based Interactions to Associate with Many of Its Substrates in Vivo

The Lectin Chaperone Calnexin Utilizes Polypeptide-based Interactions to Associate with Many of Its Substrates in Vivo

Tyrp1 and Oculocutaneous Albinism Type 3

Pink-eyed Dilution Protein Controls the Processing of Tyrosinase

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