Splice Variants of the Drosophila PS2 Integrins Differentially Interact with RGD-containing Fragments of the Extracellular Proteins Tiggrin, Ten-m, and D-Laminin α2

Graner, Michael W.; Bunch, Thomas A.; Baumgärtner, Stefan; Kerschen, Arthur; Brower, Danny L.

doi:10.1074/jbc.273.29.18235

Cited by 66 publications

(67 citation statements)

References 53 publications

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“…These results suggest that PGANT3 may glycosylate a subset of basal ECM proteins (including tiggrin) in the developing wing disc. The aforementioned ECM proteins are also known to specifically interact with ␣PS2 integrin to mediate cell adhesion in a number of developmental contexts (9,10,13,14). We postulate that glycosylation of ECM proteins in addition to tiggrin could also play a role in the wing blade adhesion.…”

Section: Discussionmentioning

confidence: 99%

A Mucin-type O-Glycosyltransferase Modulates Cell Adhesion during Drosophila Development

Zhang

Hagen

2008

Journal of Biological Chemistry

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Cell-cell and cell-matrix adhesion are crucial during many stages of eukaryotic development. Here, we provide the first example that mucin-type O-linked glycosylation is involved in a developmentally regulated cell adhesion event in Drosophila melanogaster. Mutations in one member of the evolutionarily conserved family of enzymes that initiates O-linked glycosylation alter epithelial cell adhesion in the Drosophila wing blade. A transposon insertion mutation in pgant3 or RNA interference to pgant3 resulted in blistered wings, a phenotype characteristic of genes involved in integrin-mediated cell interactions. Expression of wild type pgant3 in the mutant background rescued the wing blistering phenotype, whereas expression of another family member (pgant35A) did not, revealing a unique requirement for pgant3. pgant3 mutants displayed reduced O-glycosylation along the basal surface of larval wing imaginal discs, which was restored with wild type pgant3 expression, suggesting that reduced glycosylation of basal proteins is responsible for disruption of adhesion in the adult wing blade. Glycosylation reactions demonstrated that PGANT3 glycosylates certain extracellular matrix (ECM) proteins. Immunoprecipitation experiments revealed that PGANT3 glycosylates tiggrin, an ECM protein known to bind integrin. We propose that this glycosyltransferase is uniquely responsible for glycosylating tiggrin in the wing disc, thus modulating proper cell adhesion through integrin-ECM interactions. This study provides the first evidence for the role of O-glycosylation in a developmentally regulated, integrin-mediated, cell adhesion event and reveals a novel player in wing blade formation during Drosophila development.Cell interactions and adhesion are critical in many diverse processes, from events occurring during embryogenesis and organogenesis to wound healing and the alterations in cell adhesion seen upon tumor formation and metastasis (1). Drosophila wing development has been used as a model system to identify factors responsible for regulating cell adhesion (2-5). Aberrant adhesion of the two epithelial cell layers that comprise the adult wing blade results in separation of the cell layers, creating a localized blister shortly after eclosion. Mutations in the integrin family of cell surface receptors (2, 3, 5-7) as well as proteins that interact with integrins (4, 5, 8 -13) have been shown to produce wing blisters, highlighting the central role of integrin-mediated cell adhesion events during wing blade formation. Interestingly, many cell surface and extracellular matrix (ECM) 2 proteins that influence wing blistering also interact with integrins in other developmentally regulated cell adhesion events (2, 14, 15).Cell surface, secreted, and ECM proteins undergo a number of post-translational modifications as they transit the secretory apparatus to their final destinations. Although the roles of classical N-linked glycans and proteoglycans are widely appreciated, recent studies have elucidated crucial roles for other types of gly...

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Section: Discussionmentioning

confidence: 99%

A Mucin-type O-Glycosyltransferase Modulates Cell Adhesion during Drosophila Development

Zhang

Hagen

2008

Journal of Biological Chemistry

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“…For all cell culture experiments, the ␣PS2C (canonical) and ␤PS4A isoforms were used (Graner et al, 1998). In some experiments, these integrin-expressing S2/M3 cells also stably expressed Drosophila talin head-GFP chimeras (wild-type or R367A mutants) under the control of the yeast Gal4 UAS .…”

Section: Cell Lines and Culturementioning

confidence: 99%

Differences in Regulation ofDrosophilaand Vertebrate Integrin Affinity by Talin

Helsten

Bunch²,

Kato

et al. 2008

MBoC

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Integrin-mediated cell adhesion is essential for development of multicellular organisms. In worms, flies, and vertebrates, talin forms a physical link between integrin cytoplasmic domains and the actin cytoskeleton. Loss of either integrins or talin leads to similar phenotypes. In vertebrates, talin is also a key regulator of integrin affinity. We used a ligand-mimetic Fab fragment, TWOW-1, to assess talin's role in regulating Drosophila ␣PS2␤PS affinity. Depletion of cellular metabolic energy reduced TWOW-1 binding, suggesting ␣PS2␤PS affinity is an active process as it is for vertebrate integrins. In contrast to vertebrate integrins, neither talin knockdown by RNA interference nor talin head overexpression had a significant effect on TWOW-1 binding. Furthermore, replacement of the transmembrane or talin-binding cytoplasmic domains of ␣PS2␤PS with those of human ␣IIb␤3 failed to enable talin regulation of TWOW-1 binding. However, substitution of the extracellular and transmembrane domains of ␣PS2␤PS with those of ␣IIb␤3 resulted in a constitutively active integrin whose affinity was reduced by talin knockdown. Furthermore, wild-type ␣IIb␤3 was activated by overexpression of Drosophila talin head domain. Thus, despite evolutionary conservation of talin's integrin/cytoskeleton linkage function, talin is not sufficient to regulate Drosophila ␣PS2␤PS affinity because of structural features inherent in the ␣PS2␤PS extracellular and/or transmembrane domains. INTRODUCTIONIntegrin adhesion receptors couple the extracellular matrix with the actin cytoskeleton, allowing transmission of both mechanical force and biochemical signals across the plasma membrane (Hynes, 2002). The cytoskeletal protein talin, a product of the talin 1 (TLN1) gene, provides a key physical linkage between integrin ␤ cytoplasmic domains and F-actin (Critchley, 2004;Wiesner et al., 2005). Talin's N-terminal head region contains a FERM domain with a PTB subdomain capable of interacting with several proteins, including integrin ␤ cytoplasmic domains (Wegener et al., 2007). Talin's C-terminal rod domain contains binding sites for several cytoskeletal proteins, including actin (Calderwood, 2004;Critchley, 2004). Loss of talin in worms, flies, and mice leads to lethal phenotypes that closely resemble those caused by loss or mutation of integrins, indicating that talin's function overlaps that of integrins in the developing organism (Monkley et al., 2000;Brown et al., 2002;Brower, 2003;Cram et al., 2003;Wiesner et al., 2005). In fact, talin is essential for the normal development of organisms ranging in complexity from the multicellular slime mold Dictyostelium discoideum to vertebrates (Nuckolls et al., 1992;Albiges-Rizo et al., 1995;Bolton et al., 1997;Priddle et al., 1998;Tsujioka et al., 1999;Monkley et al., 2000;Cram et al., 2003). In vertebrates, it is also clear that talin plays a key role in regulating the affinity of integrins for adhesive ligands (Calderwood et al., 1999(Calderwood et al., , 2002Tadokoro et al., 2003;Ginsberg et al., 2005;Wegen...

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“…We have previously shown that an ECM protein, Tiggrin (3,18,19), is normally modified by the post-translational addition of the sugar GalNAc (20) through the action of a member of the UDP-GalNAc:polypeptide N-acetylgalactosaminyltransferase family of glycosyltranferases (EC 2.4.1.41). This type of evolutionarily conserved protein modification, known as mucin-type O-glycosylation (21)(22)(23), was shown to be required for proper cell adhesion between the epithelial cell layers comprising the Drosophila wing blade.…”

mentioning

confidence: 99%

An O-Glycosyltransferase Promotes Cell Adhesion during Development by Influencing Secretion of an Extracellular Matrix Integrin Ligand

Zhang

Tran

Hagen

2010

Journal of Biological Chemistry

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Protein secretion and localization are crucial during eukaryotic development, establishing local cell environments as well as mediating cell interactions, signaling, and adhesion. In this study, we demonstrate that the glycosyltransferase, pgant3, specifically modulates integrin-mediated cell adhesion by influencing the secretion and localization of the integrin ligand, Tiggrin. We demonstrate that Tiggrin is normally O-glycosylated and localized to the basal matrix where the dorsal and ventral cell layers adhere in wild type Drosophila wings. In pgant3 mutants, Tiggrin is no longer O-glycosylated and fails to be properly secreted to this basal cell layer interface, resulting in disruption of integrin-mediated cell adhesion in the wing. pgant3-mediated effects are dependent on enzymatic activity, as mutations that form a stable protein yet abrogate O-glycosyltransferase activity result in Tiggrin accumulation within the dorsal and ventral cells comprising the wing. Our results provide the first in vivo evidence for the role of O-glycosylation in the secretion of specific extracellular matrix proteins, thus altering the composition of the cellular "microenvironment" and thereby modulating developmentally regulated cell adhesion events. As alterations in cell adhesion are a hallmark of cancer progression, this work provides insight into the long-standing association between aberrant O-glycosylation and tumorigenesis.

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Splice Variants of the Drosophila PS2 Integrins Differentially Interact with RGD-containing Fragments of the Extracellular Proteins Tiggrin, Ten-m, and D-Laminin α2

Cited by 66 publications

References 53 publications

A Mucin-type O-Glycosyltransferase Modulates Cell Adhesion during Drosophila Development

A Mucin-type O-Glycosyltransferase Modulates Cell Adhesion during Drosophila Development

Differences in Regulation ofDrosophilaand Vertebrate Integrin Affinity by Talin

An O-Glycosyltransferase Promotes Cell Adhesion during Development by Influencing Secretion of an Extracellular Matrix Integrin Ligand

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