scholarly journals The laminin-binding activity of the alpha 7 integrin receptor is defined by developmentally regulated splicing in the extracellular domain.

1997 ◽  
Vol 8 (9) ◽  
pp. 1723-1734 ◽  
Author(s):  
B L Ziober ◽  
Y Chen ◽  
R H Kramer
Keyword(s):  
Muscle Development ◽  
Variable Region ◽  
Extracellular Domain ◽  
Binding Activity ◽  
Developmentally Regulated ◽  
Dependent Cell ◽  
Beta 1 Integrin ◽  
Alpha 7 ◽  
A Site ◽  
Laminin Binding

The expression pattern of the laminin-binding alpha 7 beta 1 integrin is developmentally regulated in skeletal, cardiac, and smooth muscle. The X1/X2 alternative splicing in the extracellular domain of alpha 7 is found in the variable region between conserved alpha-chain homology repeat domains III and IV, a site implicated in ligand binding. To assess differences in X1/X2 isoform activity, we generated MCF-7 cell lines transfected with alpha 7-X1/X2 cDNAs. Transfectants expressing the alpha 7-X2 variant adhered rapidly to laminin 1, whereas those expressing alpha 7-X1 failed to attach. That alpha 7-X1 exists in an inactive state was established in assays using an activating beta 1 antibody that induced X1-dependent cell adhesion and spreading. Furthermore, the activation of alpha 7-X1 was cell type specific, and when expressed in HT1080 cells, the integrin was converted into a fully functional receptor capable of promoting adhesion. Thus, the expression of the alpha 7-X1/X2 integrin is a novel mechanism that regulates receptor affinity states in a cell-specific context and may modulate integrin-dependent events during muscle development and repair.

Laminins promote the locomotion of skeletal myoblasts via the alpha 7 integrin receptor

1996 ◽  
Vol 109 (13) ◽  
pp. 3139-3150 ◽  
Author(s):  
C.C. Yao ◽  
B.L. Ziober ◽  
A.E. Sutherland ◽  
D.L. Mendrick ◽  
R.H. Kramer
Keyword(s):  
Cytoplasmic Domain ◽  
Matrix Assembly ◽  
Developmentally Regulated ◽  
Laminin 5 ◽  
Alternative Mrna Splicing ◽  
Beta 1 Integrin ◽  
Alpha 7 ◽  
Myoblast Cell ◽  
And Migration

The alpha 7 beta 1 integrin is specifically expressed by skeletal and cardiac muscles, and its expression and alternative mRNA splicing at the cytoplasmic domain are developmentally regulated. We analyzed the role of alpha 7 integrin in mediating myoblast adhesion and motility on different laminin isoforms. Mouse C2C12 and MM14 myoblast cell lines were found by flow cytometry and immunoprecipitation to express high levels of the alpha 7 integrin. Overall expression of alpha 7 increased as the C2C12 myoblasts differentiated; myoblasts expressed only the alpha 7B cytoplasmic variant whereas in differentiating myotubes alpha 7A increased markedly. Function-perturbing monoclonal antibodies generated to alpha 7 integrin efficiently blocked both adhesion and migration of MM14 and C2C12 mouse myoblasts on laminin 1. Other studies with MM14 myoblasts showed that alpha 7 is also a receptor for laminin 2/4 (human placental merosins) but not for epithelial-cell-specific laminin 5. Blocking antibody to alpha 7 only partially inhibited adhesion to laminin 2/4 but almost completely blocked motility on this substrate. Finally, to assess the potential role of the alpha 7 cytoplasmic domain, CHO cells were stably transfected to expressed chimeric alpha 5 cDNA constructs containing the wild-type alpha 5 or the alpha 7A or alpha 7B cytoplasmic domain; all forms of the integrin showed identical activities for adhesion, migration, proliferation, and matrix assembly on fibronectin substrates. These results established that alpha 7 beta 1 receptor can promote myoblast adhesion and motility on a restricted number of laminin isoforms and may be important in myogenic precursor recruitment during regeneration and differentiation.

scholarly journals On the Need to Tell Apart Fraternal Twins eEF1A1 and eEF1A2, and Their Respective Outfits

2021 ◽  
Vol 22 (13) ◽  
pp. 6973
Author(s):  
Alberto Mills ◽  
Federico Gago
Keyword(s):  
De Novo ◽  
Elongation Factor ◽  
Missense Mutations ◽  
Developmentally Regulated ◽  
High Sequence Identity ◽  
80S Ribosome ◽  
Cellular Effects ◽  
Cellular Processes ◽  
Eukaryotic Elongation Factor ◽  
A Site

eEF1A1 and eEF1A2 are paralogous proteins whose presence in most normal eukaryotic cells is mutually exclusive and developmentally regulated. Often described in the scientific literature under the collective name eEF1A, which stands for eukaryotic elongation factor 1A, their best known activity (in a monomeric, GTP-bound conformation) is to bind aminoacyl-tRNAs and deliver them to the A-site of the 80S ribosome. However, both eEF1A1 and eEF1A2 are endowed with multitasking abilities (sometimes performed by homo- and heterodimers) and can be located in different subcellular compartments, from the plasma membrane to the nucleus. Given the high sequence identity of these two sister proteins and the large number of post-translational modifications they can undergo, we are often confronted with the dilemma of discerning which is the particular proteoform that is actually responsible for the ascribed biochemical or cellular effects. We argue in this review that acquiring this knowledge is essential to help clarify, in molecular and structural terms, the mechanistic involvement of these two ancestral and abundant G proteins in a variety of fundamental cellular processes other than translation elongation. Of particular importance for this special issue is the fact that several de novo heterozygous missense mutations in the human EEF1A2 gene are associated with a subset of rare but severe neurological syndromes and cardiomyopathies.

Role of Cysteine Residues in the N-Terminal Extracellular Domain of the Human VIP 1 Receptor for Ligand Binding A Site-directed Mutagenesis Studya

2006 ◽  
Vol 805 (1) ◽  
pp. 585-589 ◽  
Author(s):  
PASCALE GAUDIN ◽  
ALAIN COUVINEAU ◽  
JEAN-JOSÉ MAORET ◽  
CHRISTIANE ROUYER-FESSARD ◽  
MARC LABURTHE
Keyword(s):  
Ligand Binding ◽  
Extracellular Domain ◽  
Site Directed Mutagenesis ◽  
Cysteine Residues ◽  
Directed Mutagenesis ◽  
A Site

Structural features and functional domains of amassin-1, a cell-binding olfactomedin protein

2007 ◽  
Vol 85 (5) ◽  
pp. 552-562 ◽  
Author(s):  
Brian J. Hillier ◽  
Victor D. Vacquier
Keyword(s):  
Disulfide Bonds ◽  
Biological Activities ◽  
Structural Features ◽  
Body Cavity ◽  
Binding Activity ◽  
Coiled Coils ◽  
Cell Binding ◽  
Calcium Dependent ◽  
Dependent Cell ◽  
A Cell

Amassin-1 mediates a rapid cell adhesion that tightly adheres sea urchin coelomocytes (body cavity immunocytes) together. Three major structural regions exist in amassin-1: a short β region, 3 coiled coils, and an olfactomedin domain. Amassin-1 contains 8 disulfide-bonded cysteines that, upon reduction, render it inactive. Truncated forms of recombinant amassin-1 were expressed and purified from Pichia pastoris and their disulfide bonding and biological activities investigated. Expressed alone, the olfactomedin domain contained 2 intramolecular disulfide bonds, existed in a monomeric state, and inhibited amassin-1-mediated clotting of coelomocytes by a calcium-dependent cell-binding activity. The N-terminal β region, containing 3 cysteines, was not required for clotting activity. The coiled coils may dimerize amassin-1 in a parallel orientation through a homodimerizing disulfide bond. Neither amassin-1 fragments that were disulfide-linked as dimers or that were engineered to exist as dimers induced coelomocytes clotting. Clotting required higher multimeric states of amassin-1, possibly tetramers, which occurred through the N-terminal β region and (or) the first segment of coiled coils.

Identification of a protein complex that binds to a dodecamer sequence found at the 3' ends of yeast mitochondrial mRNAs

1993 ◽  
Vol 13 (7) ◽  
pp. 4167-4173
Author(s):  
J Min ◽  
H P Zassenhaus
Keyword(s):  
Binding Activity ◽  
Nucleoside Triphosphate ◽  
Yeast Mitochondria ◽  
Mobility Shift ◽  
Saccharomyces Cerevisiae Mitochondria ◽  
Molecular Masses ◽  
Labeled Rna ◽  
Gel Mobility Shift ◽  
A Site

An activity from Saccharomyces cerevisiae mitochondria was identified that specifically bound to a 12-nucleotide sequence, AAUAA(U/C)AUUCUU, that is a site for processing of pre-mRNAs so as to generate the mature 3' ends of mRNAs. Because processing occurs 3' to the end of the dodecamer site, all mRNAs in yeast mitochondria terminate with that sequence. RNase T1 digestion fragments which terminated precisely at their 3' ends with the dodecamer sequence bound the activity, indicating that mRNAs in vivo would be capable of binding. Gel mobility shift analyses using RNA oligonucleotides showed that binding was reduced by a U-to-A substitution at position 3 of the dodecamer sequence; a C-to-A substitution at position 10 eliminated binding. UV cross-linking identified three polypeptides with approximate molecular masses of 19, 60, and 70 kDa as constituents of the binding activity. These estimates included the contribution of the 32P-labeled RNA oligonucleotide used to tag these polypeptides. An oligonucleotide with a UA-->AU substitution at positions 3 and 4 of the dodecamer site formed complexes deficient in the 19-kDa species, suggesting that binding specificity was inherent to the higher-molecular-weight polypeptides. Assembly of the complex at a dodecamer site on an RNA protected sequences located 5' to the dodecamer site from digestion by a nucleoside triphosphate-dependent 3' exoribonuclease found in yeast mitochondria. Since mitochondrial mRNAs terminate with an intact dodecamer sequence, the binding activity may function in the stabilization of mRNAs in addition to 3'-end formation of mRNAs.

Expression of alpha 7 integrin cytoplasmic domains during skeletal muscle development: alternate forms, conformational change, and homologies with serine/threonine kinases and tyrosine phosphatases

1993 ◽  
Vol 106 (4) ◽  
pp. 1139-1152 ◽  
Author(s):  
W.K. Song ◽  
W. Wang ◽  
H. Sato ◽  
D.A. Bielser ◽  
S.J. Kaufman
Keyword(s):  
Skeletal Muscle ◽  
Untranslated Region ◽  
Muscle Development ◽  
Myogenic Differentiation ◽  
Cytoplasmic Domain ◽  
Cytoplasmic Domains ◽  
Coding Region ◽  
Common Coding ◽  
Alternate Forms ◽  
Alpha 7

We recently reported the cloning and sequencing of the alpha 7 integrin chain and its regulated expression during the development of skeletal muscle (Song et al. (1992) J. Cell Biol. 117, 643–657). The alpha 7 chain is expressed during the development of the myogenic lineage and on adult muscle fibers and this suggests that it participates in multiple and diverse functions at different times during muscle development. One interesting portion of this isoform is its cytoplasmic domain; comprised of 77 amino acids it is the largest in the alpha chains thus reported. In these experiments we begin to study the potential functions of the alpha 7 cytoplasmic domain by analyzing homologies between the rat and human sequences, by immunologic studies using an anti-cytoplasmic domain antiserum, and by identifying two alternate forms. In keeping with the nomenclature used to describe the alpha 3 and alpha 6 alternate cytoplasmic domains, we refer to the originally reported species as alpha 7B and the two additional forms as alpha 7A and alpha 7C. These three cytoplasmic domains likely arise as a consequence of alternate splicing. A splice site at the junctions of the transmembrane and cytoplasmic domains is used to generate the alpha 3, alpha 6 and alpha 7 A and B forms. The alpha 7A form RNA contains an additional 113 nucleotides compared to the B form, and a common coding region in the A and B form RNAs is used in alternate reading frames. Part of the coding region of alpha 7B appears to be used as the 3′-untranslated region of the alpha 7A form. The alpha 7C mRNA is 595 nucleotides smaller than the alpha 7B RNA and part of the 3′-untranslated region of the alpha 7B isoform is used as coding sequence in alpha 7C. There is developmental specificity in expression of these alternate mRNAs: alpha 7A and alpha 7C transcripts are found upon terminal myogenic differentiation whereas alpha 7B is present earlier in replicating cells and diminishes upon differentiation. We suggest this selective expression of the alpha 7 cytoplasmic domains underlies the diversity in function of the alpha 7 beta 1 integrin at different stages of muscle development. Immunochemical analyses indicate that the alpha 7B cytoplasmic domain undergoes a change in conformation in response to binding laminin or upon crosslinking initiated with antibody reactive with the integrin extracellular domain. Crosslinking also promotes association of the integrin with the cell cytoskeleton.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Expression of a Large Single-Chain 13F6 Antibody with Binding Activity against Ebola Virus-Like Particles in a Plant System

2020 ◽  
Vol 21 (19) ◽  
pp. 7007 ◽  
Author(s):  
Sohee Lim ◽  
Do-Sun Kim ◽  
Kisung Ko
Keyword(s):  
Ebola Virus ◽  
Virus Disease ◽  
Variable Region ◽  
Binding Activity ◽  
Single Chain ◽  
Plant System ◽  
Gene Insertion ◽  
C Terminus ◽  
Er Retention ◽  
Human Viruses

Pathogenic animal and human viruses present a growing and persistent threat to humans worldwide. Ebola virus (EBOV) causes zoonosis in humans. Here, two structurally different anti-Ebola 13F6 antibodies, recognizing the heavily glycosylated mucin-like domain (MLD) of the glycoprotein (GP), were expressed in transgenic Nicotiana tabacum plants and designed as inexpensive and effective diagnostic antibodies against Ebola virus disease (EVD). The first was anti-EBOV 13F6 full size antibody with heavy chain (HC) and light chain (LC) (monoclonal antibody, mAb 13F6-FULL), while the second was a large single-chain (LSC) antibody (mAb 13F6-LSC). mAb 13F6-LSC was constructed by linking the 13F6 LC variable region (VL) with the HC of mAb 13F6-FULL using a peptide linker and extended to the C-terminus using the endoplasmic reticulum (ER) retention motif KDEL. Agrobacterium-mediated plant transformation was employed to express the antibodies in N. tabacum. PCR, RT-PCR, and immunoblot analyses confirmed the gene insertion, transcription, and protein expression of these antibodies, respectively. The antibodies tagged with the KDEL motif displayed high-mannose type N-glycan structures and efficient binding to EBOV-like particles (VLPs). Thus, various forms of anti-EBOV plant-derived mAbs 13F6-FULL and LSC with efficient binding affinity to EBOV VLP can be produced in the plant system.

scholarly journals Direct visualization of the extracellular binding structure of E-cadherins in liquid

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Teiko Shibata-Seki ◽  
Masato Nagaoka ◽  
Mitsuaki Goto ◽  
Eiry Kobatake ◽  
Toshihiro Akaike
Keyword(s):  
Chimeric Protein ◽  
Chelating Agent ◽  
Extracellular Domain ◽  
Cell Morphogenesis ◽  
X Ray ◽  
Dependent Cell ◽  
And Function ◽  
Direct Confirmation ◽  
Good Agreement ◽  
E Cadherin

Abstract E-cadherin is a key Ca-dependent cell adhesion molecule, which is expressed on many cell surfaces and involved in cell morphogenesis, embryonic development, EMT, etc. The fusion protein E-cad-Fc consists of the extracellular domain of E-cadherin and the IgG Fc domain. On plates coated with this chimeric protein, ES/iPS cells are cultivated particularly well and induced to differentiate. The cells adhere to the plate via E-cad-Fc in the presence of Ca2+ and detach by a chelating agent. For the purpose of clarifying the structures of E-cad-Fc in the presence and absence of Ca2+, we analyzed the molecular structure of E-cad-Fc by AFM in liquid. Our AFM observations revealed a rod-like structure of the entire extracellular domain of E-cad-Fc in the presence of Ca2+ as well as trans-binding of E-cad-Fc with adjacent molecules, which may be the first, direct confirmation of trans-dimerization of E-cadherin. The observed structures were in good agreement with an X-ray crystallographic model. Furthermore, we succeeded in visualizing the changes in the rod-like structure of the EC domains with and without calcium. The biomatrix surface plays an important role in cell culture, so the analysis of its structure and function may help promote cell engineering based on cell recognition.

POMT1 mutation results in defective glycosylation and loss of laminin-binding activity in  -DG

Neurology ◽  
2004 ◽  
Vol 62 (6) ◽  
pp. 1009-1011 ◽  
Author(s):  
D. -S. Kim ◽  
Y. K. Hayashi ◽  
H. Matsumoto ◽  
M. Ogawa ◽  
S. Noguchi ◽  
...  
Keyword(s):  
Binding Activity ◽  
Laminin Binding

scholarly journals Regulation of fibronectin and laminin binding activity in cultured human lymphoblastic cell lines

1993 ◽  
Vol 154 (3) ◽  
pp. 593-600 ◽  
Author(s):  
L. M. Stoolman ◽  
Tai-Ling Wang ◽  
Rui Situ ◽  
J. Varani
Keyword(s):  
Cell Lines ◽  
Binding Activity ◽  
Laminin Binding
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