EDTA treatment alters protein glycosylation in the cellular slime mold Dictyostelium discoideum

West, Christopher M.; Brownstein, Steven A.

doi:10.1016/0014-4827(88)90252-2

Cited by 7 publications

(9 citation statements)

References 19 publications

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“…8). 2) Expression of the catalytic domain of pp ␣-GlcNAc-T2 with a cleavable signal peptide known to direct secretion (30) rescues pp ␣-GlcNAc-T2 deficiency (Fig. 9). 3) Treatment of cells with EDTA inhibits pp ␣-GlcNAc-T2-dependent O-glycosylation (29), consistent with the presence of pp ␣-GlcNAc-T2 in an endosome-accessible compartment and its dependence, shown here, on MnCl 2 (Fig. 5A).…”

Section: Figsupporting

confidence: 84%

“…The others, A87V and F243S, occur in less well conserved regions of the protein. These substitutions are consistent with partial function of the modB allele in this strain, which exhibits a ladder of lower M r underglycosylated glycoforms of SP29 (29).…”

Section: Figsupporting

confidence: 80%

“…HU2421 is a non-axenic strain also carrying the HL220 modB allele (28). HW11 is an axenic strain that carries a modB mutant allele, which appears to partially retain function (29). Cis4c is a cisplatin-resistant strain (27) that contains an insertion of the bsr Ϫ marker in the cis4c(gntB) gene described in this report.…”

Section: Cellsmentioning

confidence: 99%

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Initiation of Mucin-type O-Glycosylation in Dictyostelium Is Homologous to the Corresponding Step in Animals and Is Important for Spore Coat Function

Wang

Metcalf

Wel

et al. 2003

Journal of Biological Chemistry

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Like animal cells, many unicellular eukaryotes modify mucin-like domains of secretory proteins with multiple O-linked glycans. Unlike animal mucin-type glycans, those of some microbial eukaryotes are initiated by ␣-linked GlcNAc rather than ␣-GalNAc. Based on sequence similarity to a recently cloned soluble polypeptide hydroxyproline GlcNAc-transferase that modifies Skp1 in the cytoplasm of the social ameba Dictyostelium, we have identified an enzyme, polypeptide ␣-N-acetylglucosaminyltransferase (pp ␣-GlcNAc-T2), that attaches GlcNAc to numerous secretory proteins in this organism. Unlike the Skp1 GlcNAc-transferase, pp ␣-GlcNAc-T2 is predicted to be a type 2 transmembrane protein. A highly purified, soluble, recombinant fragment of pp ␣-GlcNAc-T2 efficiently transfers GlcNAc from UDP-GlcNAc to synthetic peptides corresponding to mucin-like domains in two proteins that traverse the secretory pathway. pp ␣-GlcNAc-T2 is required for addition of GlcNAc to peptides in cell extracts and to the proteins in vivo. Mass spectrometry and Edman degradation analyses show that pp ␣-GlcNAc-T2 attaches GlcNAc in ␣-linkage to the Thr residues of all the synthetic mucin repeats. pp ␣-GlcNAc-T2 is encoded by the previously described modB locus defined by chemical mutagenesis, based on sequence analysis and complementation studies. This finding establishes that the many phenotypes of modB mutants, including a permeability defect in the spore coat, can now be ascribed to defects in mucin-type O-glycosylation. A comparison of the sequences of pp ␣-GlcNAc-T2 and the animal pp ␣-GalNAc-transferases reveals an ancient common ancestry indicating that, despite the different N-acetylhexosamines involved, the enzymes share a common mechanism of action.

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Section: Figsupporting

confidence: 84%

Section: Figsupporting

confidence: 80%

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Initiation of Mucin-type O-Glycosylation in Dictyostelium Is Homologous to the Corresponding Step in Animals and Is Important for Spore Coat Function

Wang

Metcalf

Wel

et al. 2003

Journal of Biological Chemistry

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“…As sialic acid has not been reported in Dictyosteliurn glycoproteins, type I1 sugars presumably contain GlcNAc. Another group of glycoantigens that fall into the type I1 category are GAXX and GAXI, recognized by mAbs 54.2 and 16.1, of West and co-workers (West & Erdos, 1988;West & Brownstein, 1988).…”

Section: Discussionmentioning

confidence: 99%

Immunodominant carbohydrate determinants in the multicellular stages of Dictyostelium discoideum

Champion

Callaghan

et al. 1991

Journal of General Microbiology

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Two families of glycoprotein are defined in DictyosteZium discoideum by the presence of different glycoconjugates, both of which are highly immunogenic in mice. The previously described monoclonal antibodies MUD50 and MUD62 recognize the glycoconjugates and identify the respective glycoprotein families. Both types of glycosylation occur on vegetative and developmentally regulated glycoproteins. The immunodominant components of both families are reportedly 0-linked sugars, but Western blots do not identify any glycoprotein that has both 0-glycans, suggesting that there are two independently processed types of 0-linked glycosylation in D. discoideum.The synthesis of the two 0-glycan families is affected by glycosylation-defective mutations. Strains with a mutation at the modB locus lack one of these glycosylation types (that recognized by MUDSO) and this mutation alters the size of two minor glycoproteins in the second family. Two new mutants, HU2470 (mod-352) and HU2471 (mod-353), lack the epitope recognized by MUD62. The two mutations map to different chromosomes. The mod-353 mutation also affects the size of PsA, a cell surface glycoprotein carrying the modB-dependent 0-glycan.

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“…Strain HW10 was derived from M4-3 in the same screen used to find HW11 (renamed from HM11; ref. 31) and is phenotypically normal. WS584, classified by morphological criteria as Dictyostelium discoideum, is a wild-type isolate previously reported to lack the monoclonal antibody (mAb) MUD1 epitope (32) and found here also to lack the mAb 59.9 epitope.…”

Section: Methodsmentioning

confidence: 99%

A glycosylation mutation affects cell fate in chimeras of Dictyostelium discoideum.

Houle

Balthazar

West

1989

Proc. Natl. Acad. Sci. U.S.A.

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Prestalk and prespore cells form a simple pattern in the pseudoplasmodium of the cellular slime mold Dictyostelium discoideum. Prestalk cells are distinguished from prespore cells by a low level of expression of a glycoantigen on their surfaces and by reduced intercellular cohesion. We examined the possible significance of these differences, using the modB mutation, which eliminates this glycoantigen genetically, leading to reduced intercellular cohesion. The formation of patterns of cells during development is essential to tissue morphogenesis. The cells in the pseudoplasmodium (or slug) of the cellular slime mold Dictyostelium form a pattern consisting of prestalk cells, which lie in the anterior 20-30% of the slug, and prespore cells, which lie in the remaining portion (1, 2). Mixed with the prespore cells is a small proportion (ca. 10%) of cells that most resemble prestalk cells but do not behave in the same way; these are referred to as anterior-like cells (3). Slugs form as the result of aggregation of cells after starvation and their rearrangement into a slug-shaped structure. During this phase, all cells appear to be the same biochemically and morphologically. However, evidence has accumulated that cells are determined to become prestalk or prespore cells at the time of starvation according to the phase of the cell cycle (4-6) and their glycogen content (7). These findings support the general model that cells first differentiate (8) and then rearrange themselves (3, 9-12), either by "self-assembly" (13) or with the help of a tip-localized organizer (14, 15). The mechanism of this rearrangement remains obscure, with evidence for both differential cell adhesion (16, 17) and cheniotaxis (18) having been presented.Determination in Dictyostelium is reversible. For example, raising the temperature in strains carrying a temperaturesensitive (ts) allele of stkA directs prespore cells to transdifferentiate into prestalk cells (19,20). Simple amputations cause morphallactic transdifferentiation of prestalk into prespore cells and vice versa (21). A diffusible substance (DIF) appears to act in the slug to influence prestalk-cell differentiation (22). In Dictyostelium mucoroides, prespore cells continuously transdifferentiate into prestalk cells as they traverse a boundary near the anterior end of the slug (23). Perhaps an initial process of deterministic differentiation and cell sorting serves to set up a provisional pattern, which is followed by a regulative phase guided by a positional component.In this report we examine the behavior of modB mutant cells introduced into normal slugs and vice versa. The modB mutation exerts a relatively limited effect on protein glycosylation: only two of the six glycoantigens (GAgs; defined at least in part by carbohydrate) we have screened are affected, and reactivity with mannose-and GIcNAc-reactive lectins is not dramatically altered (refs. 24 and 25; data not shown). The affected GAgs (GAg-XI and GAg-XX) are normally expressed only in aggregation stage cells and later...

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EDTA treatment alters protein glycosylation in the cellular slime mold Dictyostelium discoideum

Cited by 7 publications

References 19 publications

Initiation of Mucin-type O-Glycosylation in Dictyostelium Is Homologous to the Corresponding Step in Animals and Is Important for Spore Coat Function

Initiation of Mucin-type O-Glycosylation in Dictyostelium Is Homologous to the Corresponding Step in Animals and Is Important for Spore Coat Function

Immunodominant carbohydrate determinants in the multicellular stages of Dictyostelium discoideum

A glycosylation mutation affects cell fate in chimeras of Dictyostelium discoideum.

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