Limits...
Novel perspectives on the transcytotic route in osteoclasts.

Hirvonen MJ, Fagerlund K, Lakkakorpi P, Väänänen HK, Mulari MT - Bonekey Rep (2013)

Bottom Line: Helix pomatia lectin binding indicated that transcytotic vesicles expose aberrant N-acetylgalactosamine glycoconjugates, which is associated with a poor prognosis for a range of metastasizing human adenocarcinomas.Transcytotic vesicles fuse with the autophagosomal compartments and represent raft concentrates.Finally, our data suggest that the targeting of these membrane pathways may be determined by a novel F-actin-containing and FSD-circumscribing molecular barrier.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Turku, Finland.

ABSTRACT
We analyzed the characteristics of degraded bone matrix-delivering vesicles along the transcytotic route from the ruffled border to the functional secretory domain (FSD) in bone-penetrating osteoclasts. Cells of rat or human origin were cultured on bovine bone slices and analyzed via confocal microscopy. Helix pomatia lectin binding indicated that transcytotic vesicles expose aberrant N-acetylgalactosamine glycoconjugates, which is associated with a poor prognosis for a range of metastasizing human adenocarcinomas. Transcytotic vesicles fuse with the autophagosomal compartments and represent raft concentrates. Furthermore, the results of a vertical vesicle analysis suggest that multiple vesicle populations arise from the ruffled border and that some of these vesicles undergo a maturation process along the transcytotic route. Finally, our data suggest that the targeting of these membrane pathways may be determined by a novel F-actin-containing and FSD-circumscribing molecular barrier.

No MeSH data available.


Related in: MedlinePlus

The FSD is surrounded by F-actin and is colonized by microtubules but not by vimentin filaments. (a) A field emission scanning electron microscopy image of a rat osteoclast viewed from above. Villous FSD is observed on the right and the peripheral non-bone-facing plasma membrane on the left. Note the clearly demarcated border between the two membrane domains. (b, d and f) Represent single horizontal sections from the FSD level of three separate osteoclasts. Phalloidin staining reveals a ring-like F-actin structure at the top of the osteoclast. (c) WGA-lectin that binds to the transcytosed bone matrix accumulates inside the actin boundaries. (e) Vimentin filaments are absent from the interior of the ring structure, while numerous microtubules (g) are present. Immunolabelings were performed after permeabilization with Triton X-100. Bars, 10 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC3722746&req=5

f4: The FSD is surrounded by F-actin and is colonized by microtubules but not by vimentin filaments. (a) A field emission scanning electron microscopy image of a rat osteoclast viewed from above. Villous FSD is observed on the right and the peripheral non-bone-facing plasma membrane on the left. Note the clearly demarcated border between the two membrane domains. (b, d and f) Represent single horizontal sections from the FSD level of three separate osteoclasts. Phalloidin staining reveals a ring-like F-actin structure at the top of the osteoclast. (c) WGA-lectin that binds to the transcytosed bone matrix accumulates inside the actin boundaries. (e) Vimentin filaments are absent from the interior of the ring structure, while numerous microtubules (g) are present. Immunolabelings were performed after permeabilization with Triton X-100. Bars, 10 μm.

Mentions: We have previously reported that when osteoclasts are infected with Influenza virus, and the behavior of haemagglutinin at the FSD is monitored, a strictly demarcated border between the HA-labeled FSD and the peripheral non-bone-facing plasma membrane is present in 66.7% (n=120) of the resorbing osteoclasts.17 In 31.7% of the resorptive cells, there is a clear increase in the number of budding virions toward the FSD but no distinct borderline was observed. In the rest of the cells (1.6%), HA is spread throughout the non-bone-facing plasma membrane and shows no preference for the FSD. Similar results were obtained with human osteoclasts (our unpublished data). This pattern of FSD diversity is also observed with scanning electron microscopy, which reveals osteoclasts similarly with and without fully developed FSD as well as osteoclasts with microvilli throughout the non-bone-facing plasma membrane.10 The existence of osteoclasts with a strictly bordered FSD in most resorbing osteoclasts suggest the presence of a cytoskeletal machinery maintaining such a membranous structure (Figure 4a). Indeed, during numerous independent experiments, it became apparent that a thin actin circle structure at the top of a subset of resorbing osteoclasts exists (Figures 4b). WGA-lectin, which labels transcytotic degraded bone matrix and has been previously used to determine the FSD in osteoclasts,11 accumulates inside this structure (Figure 4c). There are only few vimentin filaments at this area (Figure 4e), but dense bundles of microtubules, which we have previously suggested to be involved in transcytosis of degraded bone matrix, are abundantly present (Figure 4g). The osteoclast thus harbors two actin ring structures, namely, a robust ring structure surrounding the ruffled border282930 and a less prominent ring-like structure surrounding the FSD.


Novel perspectives on the transcytotic route in osteoclasts.

Hirvonen MJ, Fagerlund K, Lakkakorpi P, Väänänen HK, Mulari MT - Bonekey Rep (2013)

The FSD is surrounded by F-actin and is colonized by microtubules but not by vimentin filaments. (a) A field emission scanning electron microscopy image of a rat osteoclast viewed from above. Villous FSD is observed on the right and the peripheral non-bone-facing plasma membrane on the left. Note the clearly demarcated border between the two membrane domains. (b, d and f) Represent single horizontal sections from the FSD level of three separate osteoclasts. Phalloidin staining reveals a ring-like F-actin structure at the top of the osteoclast. (c) WGA-lectin that binds to the transcytosed bone matrix accumulates inside the actin boundaries. (e) Vimentin filaments are absent from the interior of the ring structure, while numerous microtubules (g) are present. Immunolabelings were performed after permeabilization with Triton X-100. Bars, 10 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3722746&req=5

f4: The FSD is surrounded by F-actin and is colonized by microtubules but not by vimentin filaments. (a) A field emission scanning electron microscopy image of a rat osteoclast viewed from above. Villous FSD is observed on the right and the peripheral non-bone-facing plasma membrane on the left. Note the clearly demarcated border between the two membrane domains. (b, d and f) Represent single horizontal sections from the FSD level of three separate osteoclasts. Phalloidin staining reveals a ring-like F-actin structure at the top of the osteoclast. (c) WGA-lectin that binds to the transcytosed bone matrix accumulates inside the actin boundaries. (e) Vimentin filaments are absent from the interior of the ring structure, while numerous microtubules (g) are present. Immunolabelings were performed after permeabilization with Triton X-100. Bars, 10 μm.
Mentions: We have previously reported that when osteoclasts are infected with Influenza virus, and the behavior of haemagglutinin at the FSD is monitored, a strictly demarcated border between the HA-labeled FSD and the peripheral non-bone-facing plasma membrane is present in 66.7% (n=120) of the resorbing osteoclasts.17 In 31.7% of the resorptive cells, there is a clear increase in the number of budding virions toward the FSD but no distinct borderline was observed. In the rest of the cells (1.6%), HA is spread throughout the non-bone-facing plasma membrane and shows no preference for the FSD. Similar results were obtained with human osteoclasts (our unpublished data). This pattern of FSD diversity is also observed with scanning electron microscopy, which reveals osteoclasts similarly with and without fully developed FSD as well as osteoclasts with microvilli throughout the non-bone-facing plasma membrane.10 The existence of osteoclasts with a strictly bordered FSD in most resorbing osteoclasts suggest the presence of a cytoskeletal machinery maintaining such a membranous structure (Figure 4a). Indeed, during numerous independent experiments, it became apparent that a thin actin circle structure at the top of a subset of resorbing osteoclasts exists (Figures 4b). WGA-lectin, which labels transcytotic degraded bone matrix and has been previously used to determine the FSD in osteoclasts,11 accumulates inside this structure (Figure 4c). There are only few vimentin filaments at this area (Figure 4e), but dense bundles of microtubules, which we have previously suggested to be involved in transcytosis of degraded bone matrix, are abundantly present (Figure 4g). The osteoclast thus harbors two actin ring structures, namely, a robust ring structure surrounding the ruffled border282930 and a less prominent ring-like structure surrounding the FSD.

Bottom Line: Helix pomatia lectin binding indicated that transcytotic vesicles expose aberrant N-acetylgalactosamine glycoconjugates, which is associated with a poor prognosis for a range of metastasizing human adenocarcinomas.Transcytotic vesicles fuse with the autophagosomal compartments and represent raft concentrates.Finally, our data suggest that the targeting of these membrane pathways may be determined by a novel F-actin-containing and FSD-circumscribing molecular barrier.

View Article: PubMed Central - PubMed

Affiliation: Department of Cell Biology and Anatomy, Institute of Biomedicine, University of Turku , Turku, Finland.

ABSTRACT
We analyzed the characteristics of degraded bone matrix-delivering vesicles along the transcytotic route from the ruffled border to the functional secretory domain (FSD) in bone-penetrating osteoclasts. Cells of rat or human origin were cultured on bovine bone slices and analyzed via confocal microscopy. Helix pomatia lectin binding indicated that transcytotic vesicles expose aberrant N-acetylgalactosamine glycoconjugates, which is associated with a poor prognosis for a range of metastasizing human adenocarcinomas. Transcytotic vesicles fuse with the autophagosomal compartments and represent raft concentrates. Furthermore, the results of a vertical vesicle analysis suggest that multiple vesicle populations arise from the ruffled border and that some of these vesicles undergo a maturation process along the transcytotic route. Finally, our data suggest that the targeting of these membrane pathways may be determined by a novel F-actin-containing and FSD-circumscribing molecular barrier.

No MeSH data available.


Related in: MedlinePlus