Active peptides from the carboxyl‐terminal globular domain of laminin α2 and <i>Drosophila</i> α chains

Nomizu, Motoyoshi; Song, Sang‐Yong; Kuratomi, Yuichiro; Tanaka, Masahiko; Woo, Ho Kim; Kleinman, Hynda K.; Yamada, Yoshihiko

doi:10.1016/0014-5793(96)01060-5

Cited by 34 publications

(36 citation statements)

References 35 publications

(42 reference statements)

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“…It is possible that the peptides were coupled to CNBr-activated Sepharose beads through their ⑀-amino groups in the lysine residues and this eliminated the cell attachment activity. We showed previously an Arg-Gly-Asp (RGD) (38) containing 12-mer peptide derived from fibronectin was active in the Sepharose bead assay, but this peptide was not active in the peptide-coated plate assay (29). The peptide-conjugated Sepharose bead assay is advantageous for determining the cell binding activity of small peptides, which might not bind to the plastic or have the Cell attachment assays using HT-1080 cells were performed on untreated controls (a), or in the presence of mouse preimmune IgG (2 g/ml) (b), anti-␤ 1 integrin monoclonal antibody ascites (P4C10, 1:33 dilution) (c), anti-␣ 2 integrin monoclonal antibody ascites (P1E6, 1:33 dilution) (d), anti-␣ 3 integrin monoclonal antibody ascites (P1B5, 1:33 dilution) (e), and anti-␣ 6 integrin monoclonal antibody (GoH3, 2 g/ml) (f).…”

Section: Discussionmentioning

confidence: 99%

“…Ethanolamine-coupled beads were prepared as a control. Amounts of coupled peptide were determined by amino acid analysis (10 -20 mol of peptides/g of Sepharose bead) (29).…”

Section: Methodsmentioning

confidence: 99%

“…After washing off the unattached cells, 200 l of 1% SDS was used to dissolve the stained cells and the optical density at 560 nm was measured in a Titertek Multiscan. Peptide-coated wells without cells were processed simultaneously to subtract the background staining of coated peptides (29).…”

Section: Methodsmentioning

confidence: 99%

“…As a control, laminin-1-and ethanolamine-conjugated Sepharose beads were prepared. We also prepared laminin ␣1 chain G-domain peptides, AG-10 and AG-73 (26), conjugated to Sepharose beads as additional controls as these had been shown previously to actively bind to cells (29). We evaluated cell adhesion to the covalently conjugated peptides on either Sepharose or polystyrene beads using both HT-1080 human fibrosarcoma cells and B16-F10 mouse melanoma cells.…”

Section: Cell Attachment Activities Of Synthetic Laminin ␥1mentioning

confidence: 99%

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Identification of Cell Binding Sequences in Mouse Laminin γ1 Chain by Systematic Peptide Screening

Nomizu¹,

Kuratomi²,

Song³

et al. 1997

Journal of Biological Chemistry

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111

103

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Laminin-1, a major component of basement membranes, consists of three different chains designated ␣1, ␤1, and ␥1 and has diverse biological functions. We have identified cell binding sites on the mouse laminin ␥1 chain, using systematic screening of 165 overlapping synthetic peptides covering the entire chain. We identified 12 cell binding sequences using HT-1080 human fibrosarcoma and B16-F10 mouse melanoma cells in two independent assays employing peptide-conjugated Sepharose beads and peptide-coated dishes. Four peptides (C-16, C-28, C-64, and C-68) located on the globular domains of the ␥1 chain were the most active and showed dose-dependent cell attachment. Cell attachment to C-68 was inhibited by EDTA and by anti-␣ 2 ␤ 1 integrin antibodies. Cell attachment to C-16 and C-64 was partially inhibited by EDTA but was not inhibited by anti-integrin antibodies. EDTA and anti-integrin antibodies did not affect cell attachment to C-28. The four peptides were tested in adhesion and differentiation assays with endothelial, neuronal, and human salivary gland cells. C-16 was the most active for all of the cells, whereas the other three peptides showed cell type specificity in their activities. The active core sequences of C-16, C-28, C-64, and C-68 are YVRL, IRVTLN, TTVKYIFR, and SIKIRGTY, respectively. These sequences are highly conserved among the different species and in the laminin ␥2 chain. These results suggest that the specific sequences on the laminin ␥1 chain are biologically active and interact with distinct cell surface receptors.Laminin-1, a major component of the basement membrane matrix, has multiple biological activities including promotion of cell adhesion, spreading, growth, neurite outgrowth, tumor metastasis, and collagenase IV secretion (1-4). There are at least 11 isoforms of laminin, each consisting of three different chains (5). The most extensively characterized laminin, laminin-1 (M r ϭ 900,000) from the mouse Engelbreth-Holm-Swarm tumor consists of ␣1, ␤1, and ␥1 chains, which assemble into a triple-stranded cross-like structure with a coiled-coil domain at the long arm (6).Several active sequences on laminin-1 have been identified using proteolytic fragments, recombinant proteins and synthetic peptides (7,8). The YIGSR sequence located on the ␤1 chain promotes cell adhesion and migration, and inhibits angiogenesis and tumor metastasis (9 -11). The PDSGR and F-9 (RYVVLPR) sequences located on the ␤1 chain also promote cell adhesion (12, 13). An IKVAV sequence located on the C-terminal end of the long arm of the ␣1 chain promotes cell adhesion, neurite outgrowth, experimental metastasis, collagenase IV activity, angiogenesis, plasminogen activator activation, cell growth, and tumor growth (14 -18). Recently, we identified five cell binding sequences in the mouse laminin ␣1 chain C-terminal globular domain, G domain (positions 2111-3060), by systematic peptide screening using overlapping peptide-polystyrene beads and free peptides.Although the laminin ␥1 chain is present in most laminin isoform...

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Section: Discussionmentioning

confidence: 99%

“…Ethanolamine-coupled beads were prepared as a control. Amounts of coupled peptide were determined by amino acid analysis (10 -20 mol of peptides/g of Sepharose bead) (29).…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Cell Attachment Activities Of Synthetic Laminin ␥1mentioning

confidence: 99%

See 2 more Smart Citations

Identification of Cell Binding Sequences in Mouse Laminin γ1 Chain by Systematic Peptide Screening

Nomizu¹,

Kuratomi²,

Song³

et al. 1997

Journal of Biological Chemistry

Self Cite

111

103

View full text Add to dashboard Cite

show abstract

“…IKVAV, on the ␣1 chain, promotes cell adhesion, neurite outgrowth, experimental metastasis, collagenase IV secretion, angiogenesis, and tumor growth (16,17). Systematic screening of the G-domain of the laminin ␣1 chain has identified five active peptides with cell adhesion and spreading activities (18,19). Testing these peptides for activity with neuronal cell lines identified peptide sequences from corresponding regions of the ␣1 and ␣2 laminin chains with cell type specificity for neurite outgrowth (20).…”

mentioning

confidence: 99%

Laminin-1 and Laminin-2 G-domain Synthetic Peptides Bind Syndecan-1 and Are Involved in Acinar Formation of a Human Submandibular Gland Cell Line

Hoffman

Nomizu

Roque

et al. 1998

Journal of Biological Chemistry

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179

173

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The culture of human submandibular gland (HSG) cells on laminin-1 induces acinar differentiation. We identified a site on laminin involved in acinar differentiation using synthetic peptides derived from the C-terminal G-domain of the laminin ␣1 and ␣2 chains. The ␣1 chain peptide AG73 (RKRLQVQLSIRT) decreases the size of acini formed on laminin-1. Cells cultured with either AG73 or the homologous ␣2 chain peptide MG73 (KNRLTIELEVRT) form structures that appear acinarlike, but the cell nuclei are not polarized to the basal surface and no lumen formation occurs, indicating that additional sites on laminin are required for complete differentiation. The G-domain of laminin-1 contains both integrin and heparin binding sites, and anti-␤ 1 -integrin antibodies disrupt acinar formation. Cell adhesion to the peptides and to E3, an elastase digest fragment of laminin-1 containing AG73, is specific, since other laminin peptides or EDTA do not compete the binding. Heparin and heparan sulfate decrease cell adhesion to AG73 and MG73 but anti-␤ 1 -integrin antibodies have no effect. Treating the cell surface with heparitinase inhibits adhesion to both AG73 and MG73. We isolated cell surface ligands using both peptide affinity chromatography and laminin-1 affinity chromatography. Treating the material bound to the affinity columns with heparitinase and chondroitinase enriches for a core protein identified as syndecan-1 by Western blot analysis, thus identifying a syndecan-1 binding site in the globular domain of laminin-1 and laminin-2. In summary, multiple interactions between laminin and HSG cells contribute to acinar differentiation, involving both ␤ 1 -integrins and syndecan-1.

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SHBG region of the anticoagulant cofactor protein S: Secondary structure prediction, circular dichroism spectroscopy, and analysis of naturally occurring mutations

et al. 1997

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Protein S (PS) and growth arrest specific factor 6 (GAS6) are vitamin K-dependent proteins with similar structures. They are mosaic proteins possessing a carboxyl-terminal region presenting sequence similarity with plasma sex hormone binding globulin (plasma SHBG), although apparently not involved in steroid binding. The SHBG-like modules have sequence similarity with the G repeats of the chain A of laminin. Laminin G repeats have been reported to contain mainly beta-strands (about 40-50%) but no or little alpha structure by circular dichroism (CD) spectroscopy. Secondary structure predictions carried out in the present work unexpectedly showed a 20 to 27% helices content in the SHBG region of PS/GAS6 (about 100 residues), while plasma SHBG and laminin G repeats had around 10% helices. CD measurements for human PS indicated also that its SHBG region had about 100 residues in alpha-helical structure. These data suggest that the SHBG region of PS/GAS6 on the one hand, and the laminin G repeats and possibly plasma SHBG on the other hand, could present important structural differences. Previously reported polymorphisms and point mutations leading to PS deficiency and thrombophilia have been analyzed with our structural predictions. We found a good agreement between these structural predictions, CD measurements, experimental and clinical data. This information allows us to gain insights into the three-dimensional structure of PS that will be helpful for the design of new experiments and future clinical investigations.

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Active peptides from the carboxyl‐terminal globular domain of laminin α2 and Drosophila α chains

Cited by 34 publications

References 35 publications

Identification of Cell Binding Sequences in Mouse Laminin γ1 Chain by Systematic Peptide Screening

Identification of Cell Binding Sequences in Mouse Laminin γ1 Chain by Systematic Peptide Screening

Laminin-1 and Laminin-2 G-domain Synthetic Peptides Bind Syndecan-1 and Are Involved in Acinar Formation of a Human Submandibular Gland Cell Line

SHBG region of the anticoagulant cofactor protein S: Secondary structure prediction, circular dichroism spectroscopy, and analysis of naturally occurring mutations

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