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Partitioning and Exocytosis of Secretory Granules during Division of PC12 Cells.

Bukoreshtliev NV, Hodneland E, Eichler TW, Eifart P, Rustom A, Gerdes HH - Int J Cell Biol (2012)

Bottom Line: By combining ultrastructural analyses and time-lapse microscopy, we here show that, in dividing PC12 cells, the prominent peripheral localization of secretory granules is retained during prophase but clearly reduced during prometaphase, ending up with only few peripherally localized secretory granules in metaphase cells.During anaphase and telophase, secretory granules exhibited a pronounced movement towards the cell midzone and, evidently, their tracks colocalized with spindle microtubules.During cytokinesis, secretory granules were excluded from the midbody and accumulated at the bases of the intercellular bridge.

View Article: PubMed Central - PubMed

Affiliation: Interdisciplinary Center for Neurosciences (IZN), Department of Neurobiology, University of Heidelberg, INF 364, 69120 Heidelberg, Germany.

ABSTRACT
The biogenesis, maturation, and exocytosis of secretory granules in interphase cells have been well documented, whereas the distribution and exocytosis of these hormone-storing organelles during cell division have received little attention. By combining ultrastructural analyses and time-lapse microscopy, we here show that, in dividing PC12 cells, the prominent peripheral localization of secretory granules is retained during prophase but clearly reduced during prometaphase, ending up with only few peripherally localized secretory granules in metaphase cells. During anaphase and telophase, secretory granules exhibited a pronounced movement towards the cell midzone and, evidently, their tracks colocalized with spindle microtubules. During cytokinesis, secretory granules were excluded from the midbody and accumulated at the bases of the intercellular bridge. Furthermore, by measuring exocytosis at the single granule level, we showed, that during all stages of cell division, secretory granules were competent for regulated exocytosis. In conclusion, our data shed new light on the complex molecular machinery of secretory granule redistribution during cell division, which facilitates their release from the F-actin-rich cortex and active transport along spindle microtubules.

No MeSH data available.


Distribution of SGs during early stages of PC12 cell division. Single confocal planes of representative immunostained PC12 cells at interphase ((A), (B), (C)), prophase ((D), (E), (F)), prometaphase ((G), (H), (I)), and metaphase ((J), (K), (L)) are shown as indicated. In interphase and prophase cells, SGs accumulate in the cell periphery ((A) and (D), arrows), while the number of nonperipherally localized SGs is significantly lower ((A) and (D), arrowheads,). During prometaphase, the number of peripheral SGs is reduced ((G), arrows), concurrent with an increase in the number of SGs in the cytoplasm ((G), arrowheads). SGs exhibited a nearly homogeneous distribution in cells at metaphase ((J), arrowheads) and no accumulation of SGs in the cell periphery could be detected. Scale bars, 5 μm.
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fig3: Distribution of SGs during early stages of PC12 cell division. Single confocal planes of representative immunostained PC12 cells at interphase ((A), (B), (C)), prophase ((D), (E), (F)), prometaphase ((G), (H), (I)), and metaphase ((J), (K), (L)) are shown as indicated. In interphase and prophase cells, SGs accumulate in the cell periphery ((A) and (D), arrows), while the number of nonperipherally localized SGs is significantly lower ((A) and (D), arrowheads,). During prometaphase, the number of peripheral SGs is reduced ((G), arrows), concurrent with an increase in the number of SGs in the cytoplasm ((G), arrowheads). SGs exhibited a nearly homogeneous distribution in cells at metaphase ((J), arrowheads) and no accumulation of SGs in the cell periphery could be detected. Scale bars, 5 μm.

Mentions: In order to distinguish between cells in prophase and prometaphase, we examined the distribution of SGs during early mitosis with immunofluorescence combined with confocal microscopy. Secretogranin II (SgII), a member of the granin family of regulated secretory proteins, which is efficiently sorted in SGs of PC12 cells, was used as an endogenous marker of SGs [20]. The pattern of distribution of SGs in interphase and prophase cells was very similar. The majority of SGs in these cells was peripherally localized in accumulations (Figures 3(A), 3(D), arrows) with only a small number of SGs found in the cytoplasm (Figures 3(A), 3(D), arrowheads). Interestingly, peripheral SGs in prometaphase cells were markedly reduced in frequency (Figure 3(G), arrows), paralleled by an increase of the number of SGs deeper inside the cell (Figure 3(G), arrowheads). SGs that remained in the periphery appeared as single puncta rather than as accumulations of punctuated signals, as observed in interphase and prophase (compare Figures 3(G)with 3(A) and 3(D)). In metaphase cells, the fraction of peripherally localized SGs was further reduced and the majority of SGs was almost homogeneously distributed across the cytoplasm, except those parts occupied by the chromosomes (Figures 3(J) and 3(L)). The latter is consistent with the ultrastructural analysis of metaphase PC12 cells.


Partitioning and Exocytosis of Secretory Granules during Division of PC12 Cells.

Bukoreshtliev NV, Hodneland E, Eichler TW, Eifart P, Rustom A, Gerdes HH - Int J Cell Biol (2012)

Distribution of SGs during early stages of PC12 cell division. Single confocal planes of representative immunostained PC12 cells at interphase ((A), (B), (C)), prophase ((D), (E), (F)), prometaphase ((G), (H), (I)), and metaphase ((J), (K), (L)) are shown as indicated. In interphase and prophase cells, SGs accumulate in the cell periphery ((A) and (D), arrows), while the number of nonperipherally localized SGs is significantly lower ((A) and (D), arrowheads,). During prometaphase, the number of peripheral SGs is reduced ((G), arrows), concurrent with an increase in the number of SGs in the cytoplasm ((G), arrowheads). SGs exhibited a nearly homogeneous distribution in cells at metaphase ((J), arrowheads) and no accumulation of SGs in the cell periphery could be detected. Scale bars, 5 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3375158&req=5

fig3: Distribution of SGs during early stages of PC12 cell division. Single confocal planes of representative immunostained PC12 cells at interphase ((A), (B), (C)), prophase ((D), (E), (F)), prometaphase ((G), (H), (I)), and metaphase ((J), (K), (L)) are shown as indicated. In interphase and prophase cells, SGs accumulate in the cell periphery ((A) and (D), arrows), while the number of nonperipherally localized SGs is significantly lower ((A) and (D), arrowheads,). During prometaphase, the number of peripheral SGs is reduced ((G), arrows), concurrent with an increase in the number of SGs in the cytoplasm ((G), arrowheads). SGs exhibited a nearly homogeneous distribution in cells at metaphase ((J), arrowheads) and no accumulation of SGs in the cell periphery could be detected. Scale bars, 5 μm.
Mentions: In order to distinguish between cells in prophase and prometaphase, we examined the distribution of SGs during early mitosis with immunofluorescence combined with confocal microscopy. Secretogranin II (SgII), a member of the granin family of regulated secretory proteins, which is efficiently sorted in SGs of PC12 cells, was used as an endogenous marker of SGs [20]. The pattern of distribution of SGs in interphase and prophase cells was very similar. The majority of SGs in these cells was peripherally localized in accumulations (Figures 3(A), 3(D), arrows) with only a small number of SGs found in the cytoplasm (Figures 3(A), 3(D), arrowheads). Interestingly, peripheral SGs in prometaphase cells were markedly reduced in frequency (Figure 3(G), arrows), paralleled by an increase of the number of SGs deeper inside the cell (Figure 3(G), arrowheads). SGs that remained in the periphery appeared as single puncta rather than as accumulations of punctuated signals, as observed in interphase and prophase (compare Figures 3(G)with 3(A) and 3(D)). In metaphase cells, the fraction of peripherally localized SGs was further reduced and the majority of SGs was almost homogeneously distributed across the cytoplasm, except those parts occupied by the chromosomes (Figures 3(J) and 3(L)). The latter is consistent with the ultrastructural analysis of metaphase PC12 cells.

Bottom Line: By combining ultrastructural analyses and time-lapse microscopy, we here show that, in dividing PC12 cells, the prominent peripheral localization of secretory granules is retained during prophase but clearly reduced during prometaphase, ending up with only few peripherally localized secretory granules in metaphase cells.During anaphase and telophase, secretory granules exhibited a pronounced movement towards the cell midzone and, evidently, their tracks colocalized with spindle microtubules.During cytokinesis, secretory granules were excluded from the midbody and accumulated at the bases of the intercellular bridge.

View Article: PubMed Central - PubMed

Affiliation: Interdisciplinary Center for Neurosciences (IZN), Department of Neurobiology, University of Heidelberg, INF 364, 69120 Heidelberg, Germany.

ABSTRACT
The biogenesis, maturation, and exocytosis of secretory granules in interphase cells have been well documented, whereas the distribution and exocytosis of these hormone-storing organelles during cell division have received little attention. By combining ultrastructural analyses and time-lapse microscopy, we here show that, in dividing PC12 cells, the prominent peripheral localization of secretory granules is retained during prophase but clearly reduced during prometaphase, ending up with only few peripherally localized secretory granules in metaphase cells. During anaphase and telophase, secretory granules exhibited a pronounced movement towards the cell midzone and, evidently, their tracks colocalized with spindle microtubules. During cytokinesis, secretory granules were excluded from the midbody and accumulated at the bases of the intercellular bridge. Furthermore, by measuring exocytosis at the single granule level, we showed, that during all stages of cell division, secretory granules were competent for regulated exocytosis. In conclusion, our data shed new light on the complex molecular machinery of secretory granule redistribution during cell division, which facilitates their release from the F-actin-rich cortex and active transport along spindle microtubules.

No MeSH data available.