Cytoskeleton in Myelin-Basic Protein-Deficient Shiverer Oligodendrocytes

Dyer, Charissa A.; Philibotte, Thomas; Billings‐Gagliardi, Susan; Wolf, Merrill K.

doi:10.1159/000111273

Cited by 50 publications

(55 citation statements)

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“…Classic MBP has also been shown to stabilize microtubules from depolymerizing in the cold in vitro, and in cultured OLGs [28,29]. In cultured OLGs isolated from the shiverer mutant mouse, in which MBP is lacking, the microtubules and actin filaments were abnormal in size and distribution, and production of processes and membrane sheets was abnormal [30]. These studies support an important role for interaction of MBP with the cytoskeleton in OLG development and myelination [31][32][33].…”

Section: Introductionmentioning

confidence: 73%

“…MBP is co-localized with these cytoskeletal veins of actin filaments and microtubules, and also with cortical actin filaments in cultured OLGs. MBP has also been shown to be important for formation of the cytoskeleton and for stabilizing microtubules in the cold in primary OLGs [29,30]. In OLGs from the shiverer mutant mouse lacking MBP, the microtubules and actin filaments were abnormal in size and distribution, and production of processes and membrane sheets was abnormal, indicating a role for MBP-cytoskeletal interactions for myelination in vivo [30].…”

Section: Discussionmentioning

confidence: 99%

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Classic 18.5- and 21.5-kDa Myelin Basic Protein Isoforms Associate with Cytoskeletal and SH3-Domain Proteins in the Immortalized N19-Oligodendroglial Cell Line Stimulated by Phorbol Ester and IGF-1

et al. 2012

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The 18.5-kDa classic myelin basic protein (MBP) is an intrinsically disordered protein arising from the Golli (Genes of Oligodendrocyte Lineage) gene complex and is responsible for compaction of the myelin sheath in the central nervous system. This MBP splice isoform also has a plethora of post-translational modifications including phosphorylation, deimination, methylation, and deamidation, that reduce its overall net charge and alter its protein and lipid associations within oligodendrocytes (OLGs). It was originally thought that MBP was simply a structural component of myelin; however, additional investigations have demonstrated that MBP is multifunctional, having numerous protein-protein interactions with Ca 2+ -calmodulin, actin, tubulin, and proteins with SH3-domains, and it can tether these proteins to a lipid membrane in vitro. Here, we have examined cytoskeletal interactions of classic 18.5-kDa MBP, in vivo, using early developmental N19-OLGs transfected with fluorescently-tagged MBP, actin, tubulin, and zonula occludens 1 (ZO-1). We show that MBP redistributes to distinct 'membrane-ruffled' regions of the plasma membrane where it co-localizes with actin and tubulin, and with the SH3-domaincontaining proteins cortactin and ZO-1, when stimulated with PMA, a potent activator of the protein kinase C pathway. Moreover, using phospho-specific antibody staining, we show an increase in phosphorylated Thr98 MBP (human sequence numbering) in membrane-ruffled OLGs. CIHR Author ManuscriptCIHR Author Manuscript CIHR Author ManuscriptPreviously, Thr98 phosphorylation of MBP has been shown to affect its conformation, interactions with other proteins, and tethering of other proteins to the membrane in vitro. Here, MBP and actin were also co-localized in new focal adhesion contacts induced by IGF-1 stimulation in cells grown on laminin-2. This study supports a role for classic MBP isoforms in cytoskeletal and other protein-protein interactions during membrane and cytoskeletal remodeling in OLGs.

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Section: Introductionmentioning

confidence: 73%

Section: Discussionmentioning

confidence: 99%

Classic 18.5- and 21.5-kDa Myelin Basic Protein Isoforms Associate with Cytoskeletal and SH3-Domain Proteins in the Immortalized N19-Oligodendroglial Cell Line Stimulated by Phorbol Ester and IGF-1

et al. 2012

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“…Oligodendrocyte-enriched shake-off cultures were prepared from 2-day-old (SJL 3 B10)F 1 newborn mice by using a modified version of the method of McCarthy and de Vellis (1980) as described by Dyer et al (1995). Live cultures were treated with undiluted hybridoma supernatant, followed by addition of appropriate fluorescein isothiocyanate (FITC; Boehringer Mannheim Corp.) or TRITC (Jackson Immunoresearch) goat anti-mouse IgG or IgM secondary antibody diluted 1:40.…”

Section: Immunofluorescent Staining Of Mouse Oligodendrocytesmentioning

confidence: 99%

“…To detect internal epitopes, the cells were fixed in 4% paraformaldehyde for 5 min and permeabilized with 0.05% saponin for 15 min prior to staining with the supernatant or primary mAb. The O4 hybridoma was grown and purified as described elsewhere (Dyer et al, 1995). SMI-99 mouse mAb reactive with myelin basic protein (MBP) was purchased from Sternberger Monoclonals (Baltimore, MD).…”

Section: Immunofluorescent Staining Of Mouse Oligodendrocytesmentioning

confidence: 99%

Monoclonal antibodies to distinct regions of human myelin proteolipid protein simultaneously recognize central nervous system myelin and neurons of many vertebrate species

Greenfield¹,

Reddy²,

Lees³

et al. 2006

J of Neuroscience Research

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Myelin proteolipid protein (PLP), the major protein of mammalian CNS myelin, is a member of the proteolipid gene family (pgf). It is an evolutionarily conserved polytopic integral membrane protein and a potential autoantigen in multiple sclerosis (MS). To analyze antibody recognition of PLP epitopes in situ, monoclonal antibodies (mAbs) specific for different regions of human PLP (50–69, 100–123, 139–151, 178–191, 200–219, 264–276) were generated and used to immunostain CNS tissues of representative vertebrates. mAbs to each region recognized whole human PLP on Western blots; the anti‐100–123 mAb did not recognize DM‐20, the PLP isoform that lacks residues 116–150. All of the mAbs stained fixed, permeabilized oligodendrocytes and mammalian and avian CNS tissue myelin. Most of the mAbs also stained amphibian, teleost, and elasmobranch CNS myelin despite greater diversity of their pgf myelin protein sequences. Myelin staining was observed when there was at least 40% identity of the mAb epitope and known pgf myelin proteins of the same or related species. The pgf myelin proteins of teleosts and elasmobranchs lack 116–150; the anti‐100–123 mAb did not stain their myelin. In addition to myelin, the anti‐178–191 mAb stained many neurons in all species; other mAbs stained distinct neuron subpopulations in different species. Neuronal staining was observed when there was at least approximately 30% identity of the PLP mAb epitope and known pgf neuronal proteins of the same or related species. Thus, anti‐human PLP epitope mAbs simultaneously recognize CNS myelin and neurons even without extensive sequence identity. Widespread anti‐PLP mAb recognition of neurons suggests a novel potential pathophysiologic mechanism in MS patients, i.e., that anti‐PLP antibodies associated with demyelination might simultaneously recognize pgf epitopes in neurons, thereby affecting their functions. © 2006 Wiley‐Liss, Inc.

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“…The PLP antisense has no apparent significant effect on either AS or OL proliferation, as compared with control and sense-treated cultures. demonstrated further by comparing the phenotype of shiverer (shi) OLs in culture that are deficient in MBP (Dyer et al, 1995). These MBP-deficient OLs often have membrane sheets that directly surround the cell body but generally lack the long processes seen with the PLP-antisense-treated OLs.…”

Section: The Effects Of Blocking Plp Synthesis On Myelin Sheet Formatmentioning

confidence: 99%

Proteolipid Protein Regulates the Survival and Differentiation of Oligodendrocytes

Yang

Skoff

1997

J. Neurosci.

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Proteolipid protein (PLP) has been postulated to play a critical role in the early differentiation of oligodendrocytes (OLs) in addition to its known role as a structural component of myelin.To identify this early function, we blocked the synthesis of PLP in glial cultures with antisense oligodeoxynucleotides that targeted the PLP initiation codon. Primary glial cultures were incubated with phosphorothioate-protected oligodeoxynucleotides (S-ODNs) for up to 11 d. PLP in OLs was reduced Ͼ90%. OLs treated with antisense S-ODNs appeared strikingly healthy as judged by (1) immunocytochemical staining for myelin glycolipids and myelin basic protein, (2) their prolonged survival compared with untreated cultures, and (3) their ability to reestablish membrane sheets after removal of the S-ODNs.Our studies show that PLP is required for elaboration and stability of the myelin membrane sheets made by most OLs, but it is not necessary for the network of processes established by OLs. More importantly, the number of OLs in the antisensetreated cultures was nearly sevenfold greater after a 10 -11 d incubation with S-ODNs than in control cultures. The number of proliferating OL progenitors was not increased in the antisensetreated cultures, indicating that the increase in the number of OLs was attributable to prolonged OL survival. The tissue culture studies reveal that the absence of PLP/DM20 has the positive effect of promoting OL survival but the negative effect of preventing their full differentiation. This finding clarifies many of the paradoxical findings seen in the PLP mutants, the PLP overexpressers, and the PLP Ϫ animals.

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Cytoskeleton in Myelin-Basic Protein-Deficient Shiverer Oligodendrocytes

Cited by 50 publications

References 0 publications

Classic 18.5- and 21.5-kDa Myelin Basic Protein Isoforms Associate with Cytoskeletal and SH3-Domain Proteins in the Immortalized N19-Oligodendroglial Cell Line Stimulated by Phorbol Ester and IGF-1

Classic 18.5- and 21.5-kDa Myelin Basic Protein Isoforms Associate with Cytoskeletal and SH3-Domain Proteins in the Immortalized N19-Oligodendroglial Cell Line Stimulated by Phorbol Ester and IGF-1

Monoclonal antibodies to distinct regions of human myelin proteolipid protein simultaneously recognize central nervous system myelin and neurons of many vertebrate species

Proteolipid Protein Regulates the Survival and Differentiation of Oligodendrocytes

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