Limits...
The de novo centriole assembly pathway in HeLa cells: cell cycle progression and centriole assembly/maturation.

La Terra S, English CN, Hergert P, McEwen BF, Sluder G, Khodjakov A - J. Cell Biol. (2005)

Bottom Line: Here, we show that removal of resident centrioles (by laser ablation or needle microsurgery) does not impede cell cycle progression in HeLa cells.This maturation is not simply a time-dependent phenomenon, because de novo-formed centrioles do not mature if they are assembled in S phase-arrested cells.By selectively ablating only one centriole at a time, we find that the presence of a single centriole inhibits the assembly of additional centrioles, indicating that centrioles have an activity that suppresses the de novo pathway.

View Article: PubMed Central - PubMed

Affiliation: Wadsworth Center, New York State Department of Health, Albany, NY 12201, USA.

ABSTRACT
It has been reported that nontransformed mammalian cells become arrested during G1 in the absence of centrioles (Hinchcliffe, E., F. Miller, M. Cham, A. Khodjakov, and G. Sluder. 2001. Science. 291:1547-1550). Here, we show that removal of resident centrioles (by laser ablation or needle microsurgery) does not impede cell cycle progression in HeLa cells. HeLa cells born without centrosomes, later, assemble a variable number of centrioles de novo. Centriole assembly begins with the formation of small centrin aggregates that appear during the S phase. These, initially amorphous "precentrioles" become morphologically recognizable centrioles before mitosis. De novo-assembled centrioles mature (i.e., gain abilities to organize microtubules and replicate) in the next cell cycle. This maturation is not simply a time-dependent phenomenon, because de novo-formed centrioles do not mature if they are assembled in S phase-arrested cells. By selectively ablating only one centriole at a time, we find that the presence of a single centriole inhibits the assembly of additional centrioles, indicating that centrioles have an activity that suppresses the de novo pathway.

Show MeSH

Related in: MedlinePlus

Pedigree of a cell born without centrosome. In this particular experiment, the acentrosomal cell formed two centrioles during the first cell cycle (compare 22:00 with 29:00). When this cell underwent mitosis, both de novo–formed centrioles were distributed to one of the two progeny (a), leaving the other one acentrosomal (b). This cell activated the de novo pathway, which now resulted in the formation of eight new centrioles (b, compare 46:00 with 53:45). During third mitosis these 8 centrioles were equally distributed between the two progeny (ba and bb). Cell a, which inherited two de novo–formed centrioles as the result of the second mitosis, replicated these centrioles during the second cell cycle so that both progeny of this cell (aa and ab) inherited two centrioles (one mother and one daughter). Control cell (sister of the cell born without a centrosome) and its progeny exhibited the expected orderly replication of centrioles in both first and second cell cycles. Time is shown in hours:minutes. Bar, 5 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2171814&req=5

fig5: Pedigree of a cell born without centrosome. In this particular experiment, the acentrosomal cell formed two centrioles during the first cell cycle (compare 22:00 with 29:00). When this cell underwent mitosis, both de novo–formed centrioles were distributed to one of the two progeny (a), leaving the other one acentrosomal (b). This cell activated the de novo pathway, which now resulted in the formation of eight new centrioles (b, compare 46:00 with 53:45). During third mitosis these 8 centrioles were equally distributed between the two progeny (ba and bb). Cell a, which inherited two de novo–formed centrioles as the result of the second mitosis, replicated these centrioles during the second cell cycle so that both progeny of this cell (aa and ab) inherited two centrioles (one mother and one daughter). Control cell (sister of the cell born without a centrosome) and its progeny exhibited the expected orderly replication of centrioles in both first and second cell cycles. Time is shown in hours:minutes. Bar, 5 μm.

Mentions: The difference between mature (mother) and immature (daughter) centrioles is not limited to their ability to organize microtubules. An important step in the maturation process is to gain the ability to give birth to a new daughter centriole (for conceptual review see Mazia, 1987). Thus, if the de novo–assembled centrioles become mature during the second cell cycle, then they should begin to replicate in a normal fashion. We tested this prediction by following the progeny of cells born without centrioles with continuous time lapse microscopy over three consecutive cell cycles. We were able to obtain full three–cell cycle-long pedigrees for two cells born without centrioles. Analyses of these pedigrees revealed several important features of the centriole cycle. First, we found that centrioles formed de novo replicate in the second cell cycle (Fig. 5, cell a, which fortuitously inherited two de novo–formed centrioles, and its progeny aa and ab). Second, we found that the de novo pathway becomes active whenever the resident centrioles disappear from the cell. This was evident from those cases in which all the de novo–formed centrioles were distributed to only one of the progeny (Fig. 5, cell a; and Figs. S1 and S2). As a result, the sister cell was born without a centrosome (Fig. 5, cell b), not because of laser ablation, but because of centriole misdistribution. This cell exhibited de novo assembly of eight centrioles that were later distributed in a four-and-four fashion between the two progeny in the next mitosis (Fig. 5, cell b and its progeny ba and bb). It is noteworthy that this mitosis was multipolar; however, because of retraction of one of the cytokinesis furrows, it resulted in the formation of two cells—one mononucleated and one binucleated (not depicted). Thus, de novo–assembled centrioles become completely mature in the next cell cycle after their assembly.


The de novo centriole assembly pathway in HeLa cells: cell cycle progression and centriole assembly/maturation.

La Terra S, English CN, Hergert P, McEwen BF, Sluder G, Khodjakov A - J. Cell Biol. (2005)

Pedigree of a cell born without centrosome. In this particular experiment, the acentrosomal cell formed two centrioles during the first cell cycle (compare 22:00 with 29:00). When this cell underwent mitosis, both de novo–formed centrioles were distributed to one of the two progeny (a), leaving the other one acentrosomal (b). This cell activated the de novo pathway, which now resulted in the formation of eight new centrioles (b, compare 46:00 with 53:45). During third mitosis these 8 centrioles were equally distributed between the two progeny (ba and bb). Cell a, which inherited two de novo–formed centrioles as the result of the second mitosis, replicated these centrioles during the second cell cycle so that both progeny of this cell (aa and ab) inherited two centrioles (one mother and one daughter). Control cell (sister of the cell born without a centrosome) and its progeny exhibited the expected orderly replication of centrioles in both first and second cell cycles. Time is shown in hours:minutes. Bar, 5 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171814&req=5

fig5: Pedigree of a cell born without centrosome. In this particular experiment, the acentrosomal cell formed two centrioles during the first cell cycle (compare 22:00 with 29:00). When this cell underwent mitosis, both de novo–formed centrioles were distributed to one of the two progeny (a), leaving the other one acentrosomal (b). This cell activated the de novo pathway, which now resulted in the formation of eight new centrioles (b, compare 46:00 with 53:45). During third mitosis these 8 centrioles were equally distributed between the two progeny (ba and bb). Cell a, which inherited two de novo–formed centrioles as the result of the second mitosis, replicated these centrioles during the second cell cycle so that both progeny of this cell (aa and ab) inherited two centrioles (one mother and one daughter). Control cell (sister of the cell born without a centrosome) and its progeny exhibited the expected orderly replication of centrioles in both first and second cell cycles. Time is shown in hours:minutes. Bar, 5 μm.
Mentions: The difference between mature (mother) and immature (daughter) centrioles is not limited to their ability to organize microtubules. An important step in the maturation process is to gain the ability to give birth to a new daughter centriole (for conceptual review see Mazia, 1987). Thus, if the de novo–assembled centrioles become mature during the second cell cycle, then they should begin to replicate in a normal fashion. We tested this prediction by following the progeny of cells born without centrioles with continuous time lapse microscopy over three consecutive cell cycles. We were able to obtain full three–cell cycle-long pedigrees for two cells born without centrioles. Analyses of these pedigrees revealed several important features of the centriole cycle. First, we found that centrioles formed de novo replicate in the second cell cycle (Fig. 5, cell a, which fortuitously inherited two de novo–formed centrioles, and its progeny aa and ab). Second, we found that the de novo pathway becomes active whenever the resident centrioles disappear from the cell. This was evident from those cases in which all the de novo–formed centrioles were distributed to only one of the progeny (Fig. 5, cell a; and Figs. S1 and S2). As a result, the sister cell was born without a centrosome (Fig. 5, cell b), not because of laser ablation, but because of centriole misdistribution. This cell exhibited de novo assembly of eight centrioles that were later distributed in a four-and-four fashion between the two progeny in the next mitosis (Fig. 5, cell b and its progeny ba and bb). It is noteworthy that this mitosis was multipolar; however, because of retraction of one of the cytokinesis furrows, it resulted in the formation of two cells—one mononucleated and one binucleated (not depicted). Thus, de novo–assembled centrioles become completely mature in the next cell cycle after their assembly.

Bottom Line: Here, we show that removal of resident centrioles (by laser ablation or needle microsurgery) does not impede cell cycle progression in HeLa cells.This maturation is not simply a time-dependent phenomenon, because de novo-formed centrioles do not mature if they are assembled in S phase-arrested cells.By selectively ablating only one centriole at a time, we find that the presence of a single centriole inhibits the assembly of additional centrioles, indicating that centrioles have an activity that suppresses the de novo pathway.

View Article: PubMed Central - PubMed

Affiliation: Wadsworth Center, New York State Department of Health, Albany, NY 12201, USA.

ABSTRACT
It has been reported that nontransformed mammalian cells become arrested during G1 in the absence of centrioles (Hinchcliffe, E., F. Miller, M. Cham, A. Khodjakov, and G. Sluder. 2001. Science. 291:1547-1550). Here, we show that removal of resident centrioles (by laser ablation or needle microsurgery) does not impede cell cycle progression in HeLa cells. HeLa cells born without centrosomes, later, assemble a variable number of centrioles de novo. Centriole assembly begins with the formation of small centrin aggregates that appear during the S phase. These, initially amorphous "precentrioles" become morphologically recognizable centrioles before mitosis. De novo-assembled centrioles mature (i.e., gain abilities to organize microtubules and replicate) in the next cell cycle. This maturation is not simply a time-dependent phenomenon, because de novo-formed centrioles do not mature if they are assembled in S phase-arrested cells. By selectively ablating only one centriole at a time, we find that the presence of a single centriole inhibits the assembly of additional centrioles, indicating that centrioles have an activity that suppresses the de novo pathway.

Show MeSH
Related in: MedlinePlus