The Spindle Matrix Protein, Chromator, Is a Novel Tubulin Binding Protein That Can Interact with Both Microtubules and Free Tubulin

The role of the latent TGF-beta binding protein (LTBP) is unclear. In cultures of fetal rat calvarial cells, which form mineralized bonelike nodules, both LTBP and the TGF-beta 1 precursor localized to large fibrillar structures in the extracellular matrix. The appearance of these fibrillar structures preceded the appearance of type I collagen fibers. Plasmin treatment abolished the fibrillar staining pattern for LTBP and released a complex containing both LTBP and TGF-beta. Antibodies and antisense oligonucleotides against LTBP inhibited the formation of mineralized bonelike nodules in long-term fetal rat calvarial cultures. Immunohistochemistry of fetal and adult rat bone confirmed a fibrillar staining pattern for LTBP in vivo. These findings, together with the known homology of LTBP to the fibrillin family of proteins, suggest a novel function for LTBP, in addition to its role in matrix storage of latent TGF-beta, as a structural matrix protein that may play a role in bone formation.

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C4b Binding Protein Acts as an Innate Immune Effector Against Influenza A Virus

Frontiers in Immunology ◽

10.3389/fimmu.2020.585361 ◽

2021 ◽

Vol 11 ◽

Author(s):

Praveen M. Varghese ◽

Valarmathy Murugaiah ◽

Nazar Beirag ◽

Nigel Temperton ◽

Haseeb A. Khan ◽

...

Keyword(s):

Influenza A Virus ◽

Influenza A ◽

Matrix Protein ◽

Binding Protein ◽

Fluid Phase ◽

Factor H ◽

Epithelial Cell Line ◽

Mrna Levels ◽

Dependent Manner ◽

Independent Manner

C4b Binding Protein (C4BP) is a major fluid phase inhibitor of the classical and lectin pathways of the complement system. Complement inhibition is achieved by binding to and restricting the role of activated complement component C4b. C4BP functions as a co-factor for factor I in proteolytic inactivation of both soluble and cell surface-bound C4b, thus restricting the formation of the C3-convertase, C4b2a. C4BP also accelerates the natural decay/dissociation of the C3 convertase. This makes C4BP a prime target for exploitation by pathogens to escape complement attack, as seen in Streptococcus pyogenes or Flavivirus. Here, we examined whether C4BP can act on its own in a complement independent manner, against pathogens. C4BP bound H1N1 and H3N2 subtypes of Influenza A Virus (IAV) most likely via multiple sites in Complement Control Protein (CCP) 1-2, 4-5, and 7-8 domains of its α-chain. In addition, C4BP CCP1-2 bound H3N2 better than H1N1. C4BP bound three IAV envelope proteins: Haemagglutinin (~70 kDa), Neuraminidase (~55 kDa), and Matrix protein 1 (~25kDa). C4BP suppressed H1N1 subtype infection into the lung epithelial cell line, A549, while it promoted infection by H3N2 subtype. C4BP restricted viral entry for H1N1 but had the opposite effect on H3N2, as evident from experiments using pseudo-typed viral particles. C4BP downregulated mRNA levels of pro-inflammatory IFN-α, IL-12, and NFκB in the case of H1N1, while it promoted a pro-inflammatory immune response by upregulating IFN- α, TNF-α, RANTES, and IL-6 in the case of H3N2. We conclude that C4BP differentially modulates the efficacy of IAV entry, and hence, replication in a target cell in a strain-dependent manner, and acts as an entry inhibitor for H1N1. Thus, CCP containing complement proteins such as factor H and C4BP may have additional defense roles against IAV that do not rely on the regulation of complement activation.

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Developmental association of the beta-galactoside-binding protein galectin-1 with the nuclear matrix of rat calvarial osteoblasts

Journal of Cell Science ◽

10.1242/jcs.111.20.3035 ◽

1998 ◽

Vol 111 (20) ◽

pp. 3035-3043 ◽

Cited By ~ 1

Author(s):

J.Y. Choi ◽

A.J. van Wijnen ◽

F. Aslam ◽

J.D. Leszyk ◽

J.L. Stein ◽

...

Keyword(s):

Nuclear Matrix ◽

Osteoblast Differentiation ◽

Matrix Protein ◽

Binding Protein ◽

Protein Composition ◽

Matrix Proteins ◽

Specific Gene ◽

Two Dimensional Gel Electrophoresis ◽

Nuclear Matrix Proteins ◽

Differential Retention

The protein composition of the nuclear matrix changes significantly as the osteoblast matures from a proliferating pre-osteoblast to an osteocyte embedded in a mineralized matrix. These matrix protein are the result of developmental stage-specific gene expression during osteoblast differentiation. To isolate nuclear matrix proteins unique to the bone phenotype we analyzed nuclear matrix preparations from cultures of rat calvarial osteoblasts by high resolution two-dimensional gel electrophoresis at two different stages: proliferation (day 3) and differentiation (day 18, mineralized). We characterized one protein (14 kDa; pI 5.0), that was detectable only in the nuclear matrix of differentiated osteoblasts. By mass spectrometry and microsequencing, this protein was identified as the beta -galactoside-binding protein galectin-1. Both immunofluorescence staining of nuclear matrix preparations with the galectin-1 antibody and western blot analysis of subcellular fractions confirmed that galectin-1 is only associated with the nuclear matrix in differentiated osteoblasts as the result of differential retention. Galectin-1 protein and mRNA are present throughout osteoblast differentiation. Galectin-1 is present in the cytoplasmic and nuclear fractions in both proliferating and differentiated osteoblasts. However, its only stable binding is to the nuclear matrix of the differentiated osteoblast; but, in proliferating osteoblasts, galectin-1 is not retained in the nuclear matrix. Taken together, our results suggest that developmental association of galectin-1 with the nuclear matrix reflects differential subnuclear binding of galectin-1 during osteoblast differentiation.

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The growth-related, translationally controlled protein P23 has properties of a tubulin binding protein and associates transiently with microtubules during the cell cycle

Journal of Cell Science ◽

10.1242/jcs.112.8.1257 ◽

1999 ◽

Vol 112 (8) ◽

pp. 1257-1271 ◽

Cited By ~ 2

Author(s):

Y. Gachet ◽

S. Tournier ◽

M. Lee ◽

A. Lazaris-Karatzas ◽

T. Poulton ◽

...

Keyword(s):

Cell Cycle ◽

Mutational Analysis ◽

Binding Protein ◽

Binding Domain ◽

Dependent Manner ◽

Cell Extracts ◽

Tubulin Binding ◽

Cycle Dependent ◽

Almost All

The translationally controlled protein P23 was discovered by the early induction of its rate of synthesis after mitogenic stimulation of mouse fibroblasts. P23 is expressed in almost all mammalian tissues and it is highly conserved between animals, plants and yeast. Based on its amino acid sequence, P23 cannot be attributed to any known protein family, and its cellular function remains to be elucidated. Here, we present evidence that P23 has properties of a tubulin binding protein that associates with microtubules in a cell cycle-dependent manner. (1) P23 is a cytoplasmic protein that occurs in complexes of 100–150 kDa, and part of P23 can be immunoprecipitated from HeLa cell extracts with anti-tubulin antibodies. (2) In immunolocalisation experiments we find P23 associated with microtubules during G1, S, G2 and early M phase of the cell cycle. At metaphase, P23 is also bound to the mitotic spindle, and it is detached from the spindle during metaphase-anaphase transition. (3) A GST-P23 fusion protein interacts with alpha- and beta-tubulin, and recombinant P23 binds to taxol-stabilised microtubules in vitro. The tubulin binding domain of P23 was identified by mutational analysis; it shows similarity to part of the tubulin binding domain of the microtubule-associated protein MAP-1B. (4) Overexpression of P23 results in cell growth retardation and in alterations of cell morphology. Moreover, elevation of P23 levels leads to microtubule rearrangements and to an increase in microtubule mass and stability.

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RanBP10 Is a Cytoplasmic GDP/GTP Exchange Factor That Modulates Microtubule Dynamics in Late Megakaryocytes.

Blood ◽

10.1182/blood.v106.11.738.738 ◽

2005 ◽

Vol 106 (11) ◽

pp. 738-738

Author(s):

Harald Schulze ◽

Marei Dose ◽

Manav Korpal ◽

Joseph E. Italiano ◽

Ramesh A. Shivdasani

Keyword(s):

Binding Protein ◽

Blood Platelets ◽

Nucleotide Exchange ◽

Exchange Factor ◽

Guanine Nucleotide Exchange ◽

Nucleotide Exchange Factor ◽

Tubulin Binding ◽

Organizing Center ◽

Mammalian Genomes ◽

Β Tubulin

Abstract Megakaryocytes are large cells within the bone marrow that undergo complex fragmentation to release up to thousands of virtually identical blood platelets into the periphery. Each platelet contains a characterisitic microtubule (MT) marginal band that is derived from MT filaments present in long protrusion-like intermediate structures, designated proplatelets, that are immediate precursors of platelets. These MT filaments are generated in the MK periphery, where they require massive mobilization that is supposed to be different from either normal interphase MT nucleation that commonly depends on γ-tubulin in the MT-organizing center. or from MTs in the mitotic spindle that require Ran·GTP, which is generated along condensed chromosomes by the chromatin-asociated guanine nucleotide exchange factor (GEF) RCC1. We first demonstrated that γ-tubulin is absent in most of the mature or proplatelet-forming MKs, where it is therefore unlikely to attribute to the total MT nucleation. MTs are tubular cytoskeletal structures that contain polymerized α- or β-tubulin subunits. Mammalian genomes share 5–6 β-tubulin isotypes of which β1-tubulin is the most divergent, especially in its C-terminal domain. β1-tubulin expression is restricted to late MKs and platelets, where it accounts for most of the β-tubulin in MT filaments. Its ablation in the mouse results in thrombocytopenia, spherocytosis and attenuated platelet function. We therefore sought to identify proteins that bind to β1-tubulin and performed a yeast two-hybrid screen using a MK-derived cDNA library. We identified a cytoplasmic Ran-binding protein, RanBP10, as a factor that associates with cellular MTs and unexpectedly harbors GEF activity toward Ran. Loss of RanBP10 in cultured MKs disrupts MT organization and its overexpression drives accumulation of extranuclear Ran and assembly of thick and abnormally long MTs. RanBP10 thus functions as a localized β-tubulin binding protein that harbors GEF activity toward Ran in the cytoplasm, much like RCC1 in the nucleus. Our results suggest that spatiotemporally restricted generation of Ran·GTP in the cytoplasm organizes specialized MTs required for thrombopoiesis and that RanBP10 provides a molecular link between Ran and non-centrosomal MTs.

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