Podosomes Display Actin Turnover and Dynamic Self-Organization in Osteoclasts Expressing Actin-Green Fluorescent Protein
01 Feb 2003-Molecular Biology of the Cell (American Society for Cell Biology)-Vol. 14, Iss: 2, pp 407-416
TL;DR: A mechanism for the patterning of podosomes in osteoclasts is provided and a turnover of actin inside the podosome is revealed, showing that an additional step of differentiation, requiring microtubule integrity, stabilizes the podsomes at the periphery of the cell in large circular patterns.
Abstract: Podosomes, small actin-based adhesion structures, differ from focal adhesions in two aspects: their core structure and their ability to organize into large patterns in osteoclasts. To address the mechanisms underlying these features, we imaged live preosteoclasts expressing green fluorescent protein-actin during their differentiation. We observe that podosomes always form inside or close to podosome groups, which are surrounded by an actin cloud. Fluorescence recovery after photobleaching shows that actin turns over in individual podosomes in contrast to cortactin, suggesting a continuous actin polymerization in the podosome core. The observation of podosome assemblies during osteoclast differentiation reveals that they evolve from simple clusters into rings that expand by the continuous formation of new podosomes at their outer ridge and inhibition of podosome formation inside the rings. This self-organization of podosomes into dynamic rings is the mechanism that drives podosomes at the periphery of the cell in large circular patterns. We also show that an additional step of differentiation, requiring microtubule integrity, stabilizes the podosome circles at the cell periphery to form the characteristic podosome belt pattern of mature osteoclasts. These results therefore provide a mechanism for the patterning of podosomes in osteoclasts and reveal a turnover of actin inside the podosome.
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TL;DR: Recent progress on molecular mechanisms of formation of invasive protrusions used by tumor cells, such as lamellipodia and invadopodia, with regard to the functions of key regulatory proteins of the actin cytoskeleton; WASP family proteins, Arp2/3 complex, LIM-kinase, cofilin, and cortactin are summarized.
1,017 citations
Cites background from "Podosomes Display Actin Turnover an..."
...Because these structures are shown to require dynamic rearrangement and continuous assembly of actin filaments [41], WASP/N-WASP may contribute to these processes by inducing actin filament nucleation through activation of theArp2/3 complex....
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TL;DR: Understanding of the regulatory and functional aspects of podosome and invadopodium biology and their role in human disease is improved.
Abstract: Podosomes and invadopodia are actin-based dynamic protrusions of the plasma membrane of metazoan cells that represent sites of attachment to - and degradation of - the extracellular matrix. The key proteins in these structures include the actin regulators cortactin and neural Wiskott-Aldrich syndrome protein (N-WASP), the adaptor proteins Tyr kinase substrate with four SH3 domains (TKS4) and Tyr kinase substrate with five SH3 domains (TKS5), and the metalloprotease membrane type 1 matrix metalloprotease (MT1MMP; also known as MMP14). Many cell types can produce these structures, including invasive cancer cells, vascular smooth muscle and endothelial cells, and immune cells such as macrophages and dendritic cells. Recently, progress has been made in our understanding of the regulatory and functional aspects of podosome and invadopodium biology and their role in human disease.
953 citations
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...In osteoclasts cultured on glass, individual podosomes form transient circular rings that appear to fuse and form a podosome belt...
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TL;DR: In vitro assays performed on primary murine bone cells confirmed the dual action of strontium ranelate in vivo as an anabolic agent on bone remodeling, which stimulates bone formation through its positive action on osteoblast differentiation and function, and decreases osteoclast differentiation as well as function by disrupting actin cytoskeleton organization.
735 citations
Cites background or methods from "Podosomes Display Actin Turnover an..."
...[13]; spleens of 6– 8 week old male OF1 mice were separated on a cushion of leukocyte separation medium (Eurobio, les Ulis, France) at 1800×g for 20 min at 20 °C, and leukocytes were collected....
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...They exhibit several features such as podosomes, usually involved in cell invasion that can be implicated in their recruitment to the bone surface [13,19]....
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TL;DR: These results, combined with new findings from in vitro studies, have led to new insights into the molecular mechanisms of cell protrusive activity and chemotactic migration during invasion and metastasis.
723 citations
TL;DR: The known structural, regulatory and functional features of podosomes are summarized and discussed, the aim being to stimulate further research into these unique structures.
623 citations
References
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TL;DR: Findings indicate that, in addition to their effects on OC precursors, OPGL and OPG have profound and direct effects on mature OCs and indicate that the OC receptor, RANK, mediates OPGl's effects.
Abstract: Osteoprotegerin (OPG) and OPG-ligand (OPGL) potently inhibit and stimulate, respectively, osteoclast differentiation (Simonet, W.S., D.L. Lacey, C.R. Dunstan, M. Kelley, M.-S. Chang, R. Luethy, H.Q. Nguyen, S. Wooden, L. Bennett, T. Boone, et al. 1997. Cell. 89:309–319; Lacey, D.L., E. Timms, H.-L. Tan, M.J. Kelley, C.R. Dunstan, T. Burgess, R. Elliott, A. Colombero, G. Elliott, S. Scully, et al. 1998. Cell. 93: 165–176), but their effects on mature osteoclasts are not well understood. Using primary cultures of rat osteoclasts on bone slices, we find that OPGL causes approximately sevenfold increase in total bone surface erosion. By scanning electron microscopy, OPGL-treated osteoclasts generate more clusters of lacunae on bone suggesting that multiple, spatially associated cycles of resorption have occurred. However, the size of individual resorption events are unchanged by OPGL treatment. Mechanistically, OPGL binds specifically to mature OCs and rapidly (within 30 min) induces actin ring formation; a marked cytoskeletal rearrangement that necessarily precedes bone resorption. Furthermore, we show that antibodies raised against the OPGL receptor, RANK, also induce actin ring formation. OPGL-treated mice exhibit increases in blood ionized Ca++ within 1 h after injections, consistent with immediate OC activation in vivo. Finally, we find that OPG blocks OPGL's effects on both actin ring formation and bone resorption. Together, these findings indicate that, in addition to their effects on OC precursors, OPGL and OPG have profound and direct effects on mature OCs and indicate that the OC receptor, RANK, mediates OPGL's effects.
718 citations
TL;DR: Osteoclasts have developed an efficient machinery for dissolving crystalline hydroxyapatite and degrading organic bone matrix rich in collagen fibers that allows osteoclasts to remove large amounts of matrix-degradation products without losing their tight attachment to underlying bone.
Abstract: Osteoclasts are multinucleated cells responsible for bone resorption. They have developed an efficient machinery for dissolving crystalline hydroxyapatite and degrading organic bone matrix rich in collagen fibers. When initiating bone resorption, osteoclasts become polarized, and three distinct membrane domains appear: a ruffled border, a sealing zone and a functional secretory domain. Simultaneously, the cytoskeleton undergoes extensive re-organisation. During this process, the actin cytoskeleton forms an attachment ring at the sealing zone, the membrane domain that anchors the resorbing cell to bone matrix. The ruffled border appears inside the sealing zone, and has several characteristics of late endosomal membrane. Extensive vesicle transport to the ruffled border delivers hydrochloric acid and proteases to an area between the ruffled border and the bone surface called the resorption lacuna. In this extracellular compartment, crystalline hydroxyapatite is dissolved by acid, and a mixture of proteases degrades the organic matrix. The degradation products of collagen and other matrix components are endocytosed, transported through the cell and exocytosed through a functional secretory domain. This transcytotic route allows osteoclasts to remove large amounts of matrix-degradation products without losing their tight attachment to underlying bone. It also facilitates further processing of the degradation products intracellularly during the passage through the cell.
690 citations
"Podosomes Display Actin Turnover an..." refers background in this paper
...…Cell Biology 407 thought to evolve into the sealing zone in actively resorbing osteoclasts (Lakkakorpi et al., 1989), forming a large circular band of actin that provides tight attachment to the bone and seals off the resorption pit where proteases and protons are secreted (Vaananen et al., 2000)....
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..., 1989), forming a large circular band of actin that provides tight attachment to the bone and seals off the resorption pit where proteases and protons are secreted (Vaananen et al., 2000)....
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Book•
04 Jun 1997
TL;DR: This ebook presents an utter edition of this ebook in doc, ePub, DjVu, PDF, txt forms, and on this site you can reading manuals and other artistic eBooks online, or downloading theirs.
Abstract: If you are looking for the book Cells, Embryos and Evolution by Jon Gerhart;Marc Kirschner in pdf format, in that case you come on to the right website. We present utter edition of this ebook in doc, ePub, DjVu, PDF, txt forms. You can read Cells, Embryos and Evolution online by Jon Gerhart;Marc Kirschner either load. In addition to this ebook, on our site you can reading manuals and other artistic eBooks online, or downloading theirs. We like invite your consideration what our site does not store the book itself, but we give reference to site whereat you may downloading either read online. So that if need to download Cells, Embryos and Evolution pdf by Jon Gerhart;Marc Kirschner , then you've come to faithful website. We own Cells, Embryos and Evolution txt, PDF, doc, DjVu, ePub forms. We will be happy if you return us anew.
532 citations
"Podosomes Display Actin Turnover an..." refers background in this paper
...The microtubule network, which is also organized at the scale of the whole cell, is thought to be self-organized as defined by Gerhart and Kirschner (1997)....
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...The formation of large structures by the interplay of their basic constituents refers to the process of self-organization (Gerhart and Kirschner, 1997), which we propose to control the podosome rings....
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TL;DR: It is shown here that estrogen can suppress receptor activator of NF-kappaB ligand and macrophage colony-stimulating factor-induced differentiation of myelomonocytic precursors into multinucleated tartrate-resistant acid phosphatase-positive osteoclasts through an estrogen receptor-dependent mechanism that does not require mediation by stromal cells.
Abstract: Loss of ovarian function following menopause results in a substantial increase in bone turnover and a critical imbalance between bone formation and resorption. This imbalance leads to a progressive loss of trabecular bone mass and eventually osteoporosis, in part the result of increased osteoclastogenesis. Enhanced formation of functional osteoclasts appears to be the result of increased elaboration by support cells of osteoclastogenic cytokines such as IL-1, tumor necrosis factor, and IL-6, all of which are negatively regulated by estrogens. We show here that estrogen can suppress receptor activator of NF-κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF)-induced differentiation of myelomonocytic precursors into multinucleated tartrate-resistant acid phosphatase-positive osteoclasts through an estrogen receptor-dependent mechanism that does not require mediation by stromal cells. This suppression is dose-dependent, isomer-specific, and reversed by ICI 182780. Furthermore, the bone-sparing analogues tamoxifen and raloxifene mimic estrogen's effects. Estrogen blocks RANKL/M-CSF-induced activator protein-1-dependent transcription, likely through direct regulation of c-Jun activity. This effect is the result of a classical nuclear activity by estrogen receptor to regulate both c-Jun expression and its phosphorylation by c-Jun N-terminal kinase. Our results suggest that estrogen modulates osteoclast formation both by down-regulating the expression of osteoclastogenic cytokines from supportive cells and by directly suppressing RANKL-induced osteoclast differentiation.
465 citations
"Podosomes Display Actin Turnover an..." refers background in this paper
...This cell line differentiated into functional osteoclasts upon exposure to RANK-L and M-CSF, with similar kinetics to primary mouse splenocytes, i.e., in 8 d, as verified by TRAP staining, a marker for osteoclasts (Figure 1A); and by the ability to resorb bone (Figure 1B)....
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...To study the transition between these two patterns, we induced mouse splenocytes with RANK-L and M-CSF to differentiate into multinucleated, fully mature osteoclasts and double stained them at each day of differentiation for actin and vinculin....
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...The murine macrophage RAW cell line can differentiate into osteoclasts in the presence of RANK-L (Shevde et al., 2000)....
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...To study the transition from podosome clusters to rings, actin-GFP–expressing RAW cells stimulated with RANL-L and M-CSF were observed by confocal time-lapse microscopy after 4–6 d of stimulation with RANK-L and M-CSF....
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...Actin-GFP RAW cells were stimulated daily with RANKL/M-CSF to induce differentiation....
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TL;DR: It is found that WASp colocalizes with CDC42Hs and actin in the core of podosomes, a highly dynamic adhesion structure of human blood-derived macrophages, which indicates that Wasp controls podosome assembly and, in cooperation with CDC 42Hs,Podosome disassembly in primary human macrophage.
Abstract: Wiskott-Aldrich syndrome protein (WASp) is a hematopoietic-specific, multidomain protein whose mutation is responsible for the immunodeficiency disorder Wiskott-Aldrich syndrome. WASp contains a binding motif for the Rho GTPase CDC42Hs as well as verprolin/cofilin-like actin-regulatory domains, but no specific actin structure regulated by CDC42Hs-WASp has been identified. We found that WASp colocalizes with CDC42Hs and actin in the core of podosomes, a highly dynamic adhesion structure of human blood-derived macrophages. Microinjection of constitutively active V12CDC42Hs or a constitutively active WASp fragment consisting of the verprolin/cofilin-like domains led to the disassemly of podosomes. Conversely, macrophages from patients expressing truncated forms of WASp completely lacked podosomes. These findings indicate that WASp controls podosome assembly and, in cooperation with CDC42Hs, podosome disassembly in primary human macrophages.
432 citations
"Podosomes Display Actin Turnover an..." refers background in this paper
...This is consistent with the presence of a complex of protein regulating actin polymerization composed of cdc42, WASP, and the Arp2/3 complex ( Linder et al., 1999, 2000 )....
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...This is consistent with the presence of a complex of protein regulating actin polymerization composed of cdc42, WASP, and the Arp2/3 complex (Linder et al., 1999, 2000)....
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