Identification of a large complex containing the integrin alpha 6 beta 1 laminin receptor in neural retinal cells

1994 ◽  
Vol 107 (11) ◽  
pp. 3165-3172 ◽  
Author(s):  
I. de Curtis ◽  
G. Gatti
Keyword(s):  
Biochemical Properties ◽  
Cytoskeletal Proteins ◽  
Laminin Receptor ◽  
Retinal Neurons ◽  
Differential Distribution ◽  
Gradient Distribution ◽  
Large Complex ◽  
Beta 1 Integrin ◽  
Integrin Alpha 6 ◽  
And Function

Integrin alpha 6 beta 1 is a laminin receptor involved in adhesion and neurite extension of retinal neurons on laminin. The present study was carried out to identify potential interactions between the alpha 6 beta 1 receptor and cellular proteins that may be involved in integrin signaling and function. For this purpose we have used a biochemical approach based on the solubilization of retinal neurons cultured on laminin with nonionic detergents, followed by centrifugation on sucrose velocity gradients. Analysis of the distribution of the alpha 6 and beta 1 integrin subunits in the gradients showed that they migrate as a large complex after extraction of cells with octylglucoside, but not after Triton X-100 extraction. Cytoskeletal proteins known to localize in adhesion plaques did not comigrate with alpha 6 beta 1 in octylglucoside gradients, while a set of polypeptides whose tyrosine phosphorylation was enhanced by culture on laminin colocalized with alpha 6 beta 1 on the gradients after octylglucoside solubilization. Culture of retinal neurons on bovine serum albumin, a nonadhesive substratum, partially affected the gradient distribution of the receptor after octylglucoside extraction. Furthermore, analysis of the gradient distribution of two alternatively spliced isoforms of the alpha 6 subunit, alpha 6-cytoA and alpha 6-cytoB, showed that the effect of non-adhesion on the sedimentation properties of the two integrin alpha 6 isoforms was more dramatic for alpha 6-cytoB than alpha 6-cytoA. These differences in the sedimentation behaviour indicate distinct biochemical properties of the two alpha 6 isoforms that, together with previous observations on their differential distribution in the developing retina, may reflect functional specificities.

Differential distribution of two cytoplasmic variants of the alpha 6 beta 1 integrin laminin receptor in the ventral plasma membrane of embryonic fibroblasts

1995 ◽  
Vol 108 (9) ◽  
pp. 3067-3078 ◽  
Author(s):  
A. Cattelino ◽  
R. Longhi ◽  
I. de Curtis
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Polyclonal Antibodies ◽  
Focal Adhesions ◽  
Receptor Occupancy ◽  
Cell Types ◽  
Laminin Receptor ◽  
Differential Distribution ◽  
Beta 1 Integrin ◽  
Integrin Alpha 6

The integrin alpha 6 beta 1 is a receptor involved in the adhesion of several cell types to laminin. By using function-blocking antibodies, we have shown that alpha 6 beta 1 is a functional laminin receptor in chick embryo fibroblasts. We also found that these cells express two variants of the alpha 6 subunit, alpha 6A and alpha 6B, characterized by different cytoplasmic domains. By using indirect immunofluorescence with isoform-specific polyclonal antibodies, we showed that the two isoforms of the alpha 6 subunit distribute differently on the ventral plasma membrane of these cells cultured on laminin-coated substrates. In fact, while the alpha 6A subunit was found codistributing with vinculin in focal contacts, the alpha 6B subunit showed a homogeneously distributed punctate pattern. This difference was particularly evident when preparations of ventral plasma membranes were used for the immunolocalization. Furthermore, when cells were cultured on fibronectin, a substrate not recognized by the alpha 6 beta 1 laminin receptor, the distribution of the two alpha 6 isoforms was similar to that observed on laminin, with alpha 6A still colocalizing with vinculin in focal adhesions. Our results indicate that two forms of the alpha 6 beta 1 laminin receptor coexpressed in the same cells show distinctive distributions, and suggest that receptor occupancy by laminin is not essential for the accumulation of the alpha 6A beta 1 integrin in adhesion plaques.

scholarly journals The integrin alpha 6 beta 4 is a laminin receptor.

1992 ◽  
Vol 117 (3) ◽  
pp. 671-678 ◽  
Author(s):  
E C Lee ◽  
M M Lotz ◽  
G D Steele ◽  
A M Mercurio
Keyword(s):  
Cell Line ◽  
Human Cell ◽  
Specific Antibody ◽  
Colon Adenocarcinoma ◽  
Human Cell Line ◽  
Laminin Receptor ◽  
Receptor Function ◽  
A Cells ◽  
Beta 1 Integrin ◽  
Integrin Alpha 6

In this study, the putative laminin receptor function of the alpha 6 beta 4 integrin was assessed. For this purpose, we used a human cell line, referred to as clone A, that was derived from a highly invasive, colon adenocarcinoma. This cell line, which expresses the alpha 6 beta 4 integrin, adheres to the E8 and not to the P1 fragment of laminin. The adhesion of clone A cells to laminin is extremely rapid with half-maximal adhesion observed at 5 min after plating. Adhesion to laminin is blocked by GoH3, and alpha 6 specific antibody (60% inhibition), as well as by A9, a beta 4 specific antibody (30% inhibition). Most importantly, we demonstrate that alpha 6 beta 4 binds specifically to laminin-Sepharose columns in the presence of either Mg2+ or Mn2+ and it is eluted from these columns with EDTA but not with NaCl. The alpha 6 beta 4 integrin does not bind to collagen-Sepharose, but the alpha 2 beta 1 integrin does bind. Clone A cells do not express alpha 6 beta 1 as evidenced by the following observations: (a) no beta 1 integrin is detected in beta 1 immunoblots of GoH3 immunoprecipitates; and (b) no alpha 6 beta 1 integrin is seen in GoH3 immunoprecipitates of clone A extracts that had been immunodepleted of all beta 4 containing integrin using the A9 antibody. These data establish that laminin is a ligand for the alpha 6 beta 4 integrin and that this integrin can function as a laminin receptor independently of alpha 6 beta 1.

scholarly journals Laminin receptors in the retina: sequence analysis of the chick integrin alpha 6 subunit. Evidence for transcriptional and posttranslational regulation.

1991 ◽  
Vol 113 (2) ◽  
pp. 405-416 ◽  
Author(s):  
I de Curtis ◽  
V Quaranta ◽  
R N Tamura ◽  
L F Reichardt
Keyword(s):  
Retinal Ganglion Cells ◽  
Ganglion Cells ◽  
Developmental Regulation ◽  
Cell Types ◽  
Laminin Receptor ◽  
Retinal Ganglion ◽  
Retinal Neurons ◽  
Laminin Receptors ◽  
Chick Retina ◽  
Integrin Alpha 6

The integrin alpha 6 beta 1 is a prominent laminin receptor used by many cell types. In the present work, we isolate clones and determine the primary sequence of the chick integrin alpha 6 subunit. We show that alpha 6 beta 1 is a prominent integrin expressed by cells in the developing chick retina. Between embryonic days 6 and 12, both retinal ganglion cells and other retinal neurons lose selected integrin functions, including the ability to attach and extend neurites on laminin. In retinal ganglion cells, we show that this is correlated with a dramatic decrease in alpha 6 mRNA and protein, suggesting that changes in gene expression account for the developmental regulation of the interactions of these neurons with laminin. In other retinal neurons the expression of alpha 6 mRNA and protein remains high while function is lost, suggesting that the function of the alpha 6 beta 1 heterodimer in these cells is regulated by posttranslational mechanisms.

scholarly journals The activation dependent adhesion of macrophages to laminin involves cytoskeletal anchoring and phosphorylation of the alpha 6 beta 1 integrin.

1990 ◽  
Vol 110 (6) ◽  
pp. 2167-2174 ◽  
Author(s):  
L M Shaw ◽  
J M Messier ◽  
A M Mercurio
Keyword(s):  
Cell Surface ◽  
De Novo ◽  
Surface Expression ◽  
Laminin Receptor ◽  
Dynamic Matrix ◽  
Beta 1 Integrin ◽  
Macrophage Adhesion ◽  
Coated Surfaces ◽  
Integrin Alpha 6 ◽  
Triton X

Macrophages require activation with either PMA (Mercurio, A. M., and L. M. Shaw. 1988. J. Cell Biol. 107:1873-1880) or interferon-gamma (Shaw, L. M., and A. M. Mercurio. 1989. J. Exp. Med. 169:303-308) to adhere to a laminin substratum. In the present study, we identified an integrin laminin receptor on macrophages and characterized cellular changes that occur in response to PMA activation that facilitate laminin adhesion. A monoclonal antibody (GoH3) that recognizes the integrin alpha 6 subunit (Sonnenberg, A., H. Janssen, F. Hogervorst, J. Calafat, and J. Hilgers. 1987. J. Biol. Chem. 262:10376-10383) specifically inhibited adhesion to laminin-coated surfaces. This antibody precipitated an alpha 6 beta 1 heterodimer (Mr 130/110 kD) from 125I surface-labeled macrophages. The amount of radiolabeled receptor on the cell surface did not increase after PMA activation. Thus, the induction of laminin adhesion cannot be attributed to de novo or increased surface expression of alpha 6 beta 1. By initially removing the Triton X-100-soluble fraction of macrophages and then disrupting the remaining cytoskeletal framework, we observed that 75% of the alpha 6 beta 1 heterodimer on the cell surface is anchored to the cytoskeleton in macrophages that had adhered to a laminin substratum in response to PMA. Significant cytoskeletal anchoring of this receptor was not observed in macrophages that had adhered to fibronectin or tissue culture plastic, nor was it seen in nonadherent cells. PMA also induced phosphorylation of the cytoplasmic domain of the alpha 6 subunit, but not the beta 1 subunit. Phosphorylated alpha 6 was localized to the cytoskeletal fraction only in macrophages plated on a laminin substratum. In summary, our results support a mechanism for the regulation of macrophage adhesion to laminin that involves specific and dynamic matrix integrin-cytoskeletal interactions that may be facilitated by integrin phosphorylation.

Joints and connective tissue—structure and function

2020 ◽  
pp. 4379-4385
Author(s):  
Thomas Pap ◽  
Adelheid Korb-Pap ◽  
Christine Hartmann ◽  
Jessica Bertrand
Keyword(s):  
Articular Cartilage ◽  
Biochemical Properties ◽  
Structure And Function ◽  
Physical Interaction ◽  
Connective Tissues ◽  
Inflammatory Joint Diseases ◽  
Synovial Joints ◽  
Key Features ◽  
And Function ◽  
Connective Tissue Structure

Synovial joints are complex functional elements of the vertebrate body that provide animals with motion capabilities and hence the ability for locomotion and direct physical interaction with their environment. They are composed of different connective tissues structures that are derived from the same developmental structures in the embryo but have distinct cellular and biochemical properties. Articular cartilage and synovial membrane are key components of synovial joints and show several peculiarities that makes them different from other tissues. An in-depth knowledge of these features is important not only for understanding key features of articular function, but also providing explanations for important characteristics of both degenerative and inflammatory joint diseases. This chapter reviews the structure, biochemical composition, and function of articular cartilage and synovium, and points to important links between physiology and pathologic conditions, particularly arthritis.

scholarly journals Abnormal subcellular distribution of myosin and talin in Wistar Furth rat platelets [published erratum appears in Blood 1995 Aug 1;86(3):1242]

Blood ◽  
1995 ◽  
Vol 85 (9) ◽  
pp. 2436-2446 ◽  
Author(s):  
TI Pestina ◽  
CW Jackson ◽  
PE Stenberg
Keyword(s):  
Plasma Membranes ◽  
Cytoskeletal Proteins ◽  
Striking Difference ◽  
Membrane Association ◽  
Fourfold Increase ◽  
Membrane Blebbing ◽  
Normal Platelet ◽  
And Function ◽  
Platelet Formation ◽  
Rat Platelets

The roles of most cytoskeletal proteins in platelet formation and function remain largely undefined. We earlier detected megakaryocyte membrane blebbing and a unique antigenic determinant associated with a missense mutation in the cytoskeletal protein, talin, in an animal model of hereditary macrothrombocytopenia, the Wistar Furth (WF) rat, which led us to examine the distribution of talin and other cytoskeletal proteins in resting normal and WF rat platelets. In contrast to the conclusions of an earlier ultrastructural analysis, our biochemical and ultrastructural immunogold studies indicate a significant membrane-association of talin in both resting normal and WF rat platelets as found earlier for rat megakaryocytes. Talin was associated with plasma membranes, membranes of the surface-connected canalicular system, and with alpha-granule membranes of both normal and WF rat platelets, but as in WF megakaryocytes, talin was absent from the large membrane complexes of WF platelets. An even more striking difference was seen in the distribution of myosin in subcellular fractions of normal and WF rat platelets separated in density gradients, in which the proportion of myosin in the least dense WF rat platelet membrane fraction was one half that in the same normal platelet fraction. This difference was balanced by a fourfold increase in myosin in the most dense WF rat subcellular fraction, which is highly enriched for alpha-granules. These results support our hypothesis that the platelet abnormalities of the WF rat are related to defects in the megakaryocyte-platelet cytoskeleton.

Characterization of a butyrate kinase from Desulfovibrio vulgaris str. Hildenborough

2020 ◽  
Vol 367 (6) ◽  
Author(s):  
Maxwell J Bachochin ◽  
Jessica Castillo Venegas ◽  
Gundeep Singh ◽  
Liyang Zhang ◽  
Robert D Barber
Keyword(s):  
Gel Filtration ◽  
Binding Pocket ◽  
Biochemical Properties ◽  
Chain Fatty Acid ◽  
Desulfovibrio Vulgaris ◽  
Optimum Temperature ◽  
Butyrate Kinase ◽  
Branched Chain ◽  
And Function

ABSTRACT Short and branched chain fatty acid kinases participate in both bacterial anabolic and catabolic processes, including fermentation, through the reversible, ATP-dependent synthesis of acyl phosphates. This study reports biochemical properties of a predicted butyrate kinase from Desulfovibrio vulgaris str. Hildenborough (DvBuk) expressed heterologously and purified from Escherichia coli. Gel filtration chromatography indicates purified DvBuk is active as a dimer. The optimum temperature and pH for DvBuk activity is 44°C and 7.5, respectively. The enzyme displays enhanced thermal stability in the presence of substrates as observed for similar enzymes. Measurement of kcat and KM for various substrates reveals DvBuk exhibits the highest catalytic efficiencies for butyrate, valerate and isobutyrate. In particular, these measurements reveal this enzyme's apparent high affinity for C4 fatty acids relative to other butyrate kinases. These results have implications on structure and function relationships within the ASKHA superfamily of phosphotransferases, particularly regarding the acyl binding pocket, as well as potential physiological roles for this enzyme in Desulfovibrio vulgaris str. Hildenborough.

scholarly journals Much More Than a Scaffold: Cytoskeletal Proteins in Neurological Disorders

Cells ◽  
2020 ◽  
Vol 9 (2) ◽  
pp. 358 ◽  
Author(s):  
Diana C. Muñoz-Lasso ◽  
Carlos Romá-Mateo ◽  
Federico V. Pallardó ◽  
Pilar Gonzalez-Cabo
Keyword(s):  
Neurological Disorders ◽  
Actin Filaments ◽  
Molecular Mechanisms ◽  
Current Knowledge ◽  
Neurological Diseases ◽  
Three Dimensional ◽  
Cytoskeletal Proteins ◽  
Dimensional Lattice ◽  
New Treatments ◽  
And Function

Recent observations related to the structure of the cytoskeleton in neurons and novel cytoskeletal abnormalities involved in the pathophysiology of some neurological diseases are changing our view on the function of the cytoskeletal proteins in the nervous system. These efforts allow a better understanding of the molecular mechanisms underlying neurological diseases and allow us to see beyond our current knowledge for the development of new treatments. The neuronal cytoskeleton can be described as an organelle formed by the three-dimensional lattice of the three main families of filaments: actin filaments, microtubules, and neurofilaments. This organelle organizes well-defined structures within neurons (cell bodies and axons), which allow their proper development and function through life. Here, we will provide an overview of both the basic and novel concepts related to those cytoskeletal proteins, which are emerging as potential targets in the study of the pathophysiological mechanisms underlying neurological disorders.

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