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Interphase centrosome organization by the PLP-Cnn scaffold is required for centrosome function.

Lerit DA, Jordan HA, Poulton JS, Fagerstrom CJ, Galletta BJ, Peifer M, Rusan NM - J. Cell Biol. (2015)

Bottom Line: Pericentriolar material (PCM) mediates the microtubule (MT) nucleation and anchoring activity of centrosomes.A scaffold organized by Centrosomin (Cnn) serves to ensure proper PCM architecture and functional changes in centrosome activity with each cell cycle.Focusing on the mitotic-to-interphase transition in Drosophila melanogaster embryos, we show that the elaboration of the interphase Cnn scaffold defines a major structural rearrangement of the centrosome.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cell Biology and Physiology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892.

ABSTRACT
Pericentriolar material (PCM) mediates the microtubule (MT) nucleation and anchoring activity of centrosomes. A scaffold organized by Centrosomin (Cnn) serves to ensure proper PCM architecture and functional changes in centrosome activity with each cell cycle. Here, we investigate the mechanisms that spatially restrict and temporally coordinate centrosome scaffold formation. Focusing on the mitotic-to-interphase transition in Drosophila melanogaster embryos, we show that the elaboration of the interphase Cnn scaffold defines a major structural rearrangement of the centrosome. We identify an unprecedented role for Pericentrin-like protein (PLP), which localizes to the tips of extended Cnn flares, to maintain robust interphase centrosome activity and promote the formation of interphase MT asters required for normal nuclear spacing, centrosome segregation, and compartmentalization of the syncytial embryo. Our data reveal that Cnn and PLP directly interact at two defined sites to coordinate the cell cycle-dependent rearrangement and scaffolding activity of the centrosome to permit normal centrosome organization, cell division, and embryonic viability.

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PLP maintains genome stability. (A) Live H2A-RFP in embryos. Broken circles show mitotic asynchrony. Arrowheads show lagging chromosomes (9:00) followed by NUF (16:00). (B) NUF (broken circle) detected by DAPI. (C and D) The frequency (C) and amount (D) of NUF is quantified. (E) γH2Av (red) labels nuclei (DAPI, blue) ejected from the cortex. Arrowheads show nuclei that have undergone nuclear fallout and stain positive for γ-H2A.The negative sign indicates distance below the embryo surface. (F) Embryos stained with DAPI (red; all nuclei) and pH3 (green; mitotic nuclei) to detect mitotic asynchrony. Results are quantified in F′. Data are mean ± SEM (error bars) for C, all other data are mean ± SD. Time is given in minutes:seconds. *, P < 0.01; ***, P < 0.0001; n.s., not significant. Data shown are from a single representative experiment out of two or more repeats. Bars: (A) 10 µm; (B, E, and F) 20 µm.
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fig7: PLP maintains genome stability. (A) Live H2A-RFP in embryos. Broken circles show mitotic asynchrony. Arrowheads show lagging chromosomes (9:00) followed by NUF (16:00). (B) NUF (broken circle) detected by DAPI. (C and D) The frequency (C) and amount (D) of NUF is quantified. (E) γH2Av (red) labels nuclei (DAPI, blue) ejected from the cortex. Arrowheads show nuclei that have undergone nuclear fallout and stain positive for γ-H2A.The negative sign indicates distance below the embryo surface. (F) Embryos stained with DAPI (red; all nuclei) and pH3 (green; mitotic nuclei) to detect mitotic asynchrony. Results are quantified in F′. Data are mean ± SEM (error bars) for C, all other data are mean ± SD. Time is given in minutes:seconds. *, P < 0.01; ***, P < 0.0001; n.s., not significant. Data shown are from a single representative experiment out of two or more repeats. Bars: (A) 10 µm; (B, E, and F) 20 µm.

Mentions: Defects in centrosome positioning and interphase MT arrangement may contribute to aberrant nuclear spacing and cell cycle length. Indeed, polyploid pseudo-cells were observed in ∼20% of plp− embryos (Fig. 6, G and H). Additional analysis shows that cell cycle progression is altered in some cells, as nuclei in plp− mutants lose the stereotyped mitotic synchrony characteristic of WT syncytial divisions (Fig. 7, F and F′). Together, these data support a role for PLP in organizing the symmetric, compact PCM necessary for proper MT organization, centrosome separation, and efficient cell cycle progression. Furthermore, the extensive similarities observed in plp− and cnn− loss-of-function studies further support a model where PLP and Cnn function in a common pathway to modulate centrosome size and activity.


Interphase centrosome organization by the PLP-Cnn scaffold is required for centrosome function.

Lerit DA, Jordan HA, Poulton JS, Fagerstrom CJ, Galletta BJ, Peifer M, Rusan NM - J. Cell Biol. (2015)

PLP maintains genome stability. (A) Live H2A-RFP in embryos. Broken circles show mitotic asynchrony. Arrowheads show lagging chromosomes (9:00) followed by NUF (16:00). (B) NUF (broken circle) detected by DAPI. (C and D) The frequency (C) and amount (D) of NUF is quantified. (E) γH2Av (red) labels nuclei (DAPI, blue) ejected from the cortex. Arrowheads show nuclei that have undergone nuclear fallout and stain positive for γ-H2A.The negative sign indicates distance below the embryo surface. (F) Embryos stained with DAPI (red; all nuclei) and pH3 (green; mitotic nuclei) to detect mitotic asynchrony. Results are quantified in F′. Data are mean ± SEM (error bars) for C, all other data are mean ± SD. Time is given in minutes:seconds. *, P < 0.01; ***, P < 0.0001; n.s., not significant. Data shown are from a single representative experiment out of two or more repeats. Bars: (A) 10 µm; (B, E, and F) 20 µm.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4494003&req=5

fig7: PLP maintains genome stability. (A) Live H2A-RFP in embryos. Broken circles show mitotic asynchrony. Arrowheads show lagging chromosomes (9:00) followed by NUF (16:00). (B) NUF (broken circle) detected by DAPI. (C and D) The frequency (C) and amount (D) of NUF is quantified. (E) γH2Av (red) labels nuclei (DAPI, blue) ejected from the cortex. Arrowheads show nuclei that have undergone nuclear fallout and stain positive for γ-H2A.The negative sign indicates distance below the embryo surface. (F) Embryos stained with DAPI (red; all nuclei) and pH3 (green; mitotic nuclei) to detect mitotic asynchrony. Results are quantified in F′. Data are mean ± SEM (error bars) for C, all other data are mean ± SD. Time is given in minutes:seconds. *, P < 0.01; ***, P < 0.0001; n.s., not significant. Data shown are from a single representative experiment out of two or more repeats. Bars: (A) 10 µm; (B, E, and F) 20 µm.
Mentions: Defects in centrosome positioning and interphase MT arrangement may contribute to aberrant nuclear spacing and cell cycle length. Indeed, polyploid pseudo-cells were observed in ∼20% of plp− embryos (Fig. 6, G and H). Additional analysis shows that cell cycle progression is altered in some cells, as nuclei in plp− mutants lose the stereotyped mitotic synchrony characteristic of WT syncytial divisions (Fig. 7, F and F′). Together, these data support a role for PLP in organizing the symmetric, compact PCM necessary for proper MT organization, centrosome separation, and efficient cell cycle progression. Furthermore, the extensive similarities observed in plp− and cnn− loss-of-function studies further support a model where PLP and Cnn function in a common pathway to modulate centrosome size and activity.

Bottom Line: Pericentriolar material (PCM) mediates the microtubule (MT) nucleation and anchoring activity of centrosomes.A scaffold organized by Centrosomin (Cnn) serves to ensure proper PCM architecture and functional changes in centrosome activity with each cell cycle.Focusing on the mitotic-to-interphase transition in Drosophila melanogaster embryos, we show that the elaboration of the interphase Cnn scaffold defines a major structural rearrangement of the centrosome.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cell Biology and Physiology Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892.

ABSTRACT
Pericentriolar material (PCM) mediates the microtubule (MT) nucleation and anchoring activity of centrosomes. A scaffold organized by Centrosomin (Cnn) serves to ensure proper PCM architecture and functional changes in centrosome activity with each cell cycle. Here, we investigate the mechanisms that spatially restrict and temporally coordinate centrosome scaffold formation. Focusing on the mitotic-to-interphase transition in Drosophila melanogaster embryos, we show that the elaboration of the interphase Cnn scaffold defines a major structural rearrangement of the centrosome. We identify an unprecedented role for Pericentrin-like protein (PLP), which localizes to the tips of extended Cnn flares, to maintain robust interphase centrosome activity and promote the formation of interphase MT asters required for normal nuclear spacing, centrosome segregation, and compartmentalization of the syncytial embryo. Our data reveal that Cnn and PLP directly interact at two defined sites to coordinate the cell cycle-dependent rearrangement and scaffolding activity of the centrosome to permit normal centrosome organization, cell division, and embryonic viability.

Show MeSH
Related in: MedlinePlus