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A complex containing the Sm protein CAR-1 and the RNA helicase CGH-1 is required for embryonic cytokinesis in Caenorhabditis elegans.

Audhya A, Hyndman F, McLeod IX, Maddox AS, Yates JR, Desai A, Oegema K - J. Cell Biol. (2005)

Bottom Line: Inhibition of CAR-1 by RNA-mediated depletion or mutation results in a specific defect in embryonic cytokinesis.This cytokinesis failure likely results from an anaphase spindle defect in which interzonal microtubule bundles that recruit Aurora B kinase and the kinesin, ZEN-4, fail to form between the separating chromosomes.Cumulatively, our results suggest that CAR-1 functions with CGH-1 to regulate a specific set of maternally loaded RNAs that is required for anaphase spindle structure and cytokinesis.

View Article: PubMed Central - PubMed

Affiliation: Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA. aaudhya@ucsd.edu

ABSTRACT
Cytokinesis completes cell division and partitions the contents of one cell to the two daughter cells. Here we characterize CAR-1, a predicted RNA binding protein that is implicated in cytokinesis. CAR-1 localizes to germline-specific RNA-containing particles and copurifies with the essential RNA helicase, CGH-1, in an RNA-dependent fashion. The atypical Sm domain of CAR-1, which directly binds RNA, is dispensable for CAR-1 localization, but is critical for its function. Inhibition of CAR-1 by RNA-mediated depletion or mutation results in a specific defect in embryonic cytokinesis. This cytokinesis failure likely results from an anaphase spindle defect in which interzonal microtubule bundles that recruit Aurora B kinase and the kinesin, ZEN-4, fail to form between the separating chromosomes. Depletion of CGH-1 results in sterility, but partially depleted worms produce embryos that exhibit the CAR-1-depletion phenotype. Cumulatively, our results suggest that CAR-1 functions with CGH-1 to regulate a specific set of maternally loaded RNAs that is required for anaphase spindle structure and cytokinesis.

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Schematic illustrating the asymmetric furrow ingression in wild-type and car-1(RNAi) embryos. End-on and side views are shown. (A) The contractile ring is positioned asymmetrically within the circumference of the cell equator. In a side view, a primary furrow is observed coming in from one side of the embryo. Contact between the primary furrow and the interzonal microtubule bundles between the separated chromosomes triggers the other side of the contractile ring (the secondary furrow) to pull away from the edge of the embryo, and the contractile ring closes down around the compacted interzonal microtubule bundles (the midbody). (B) In car-1(RNAi) embryos, the primary furrow ingresses, but fails to come into contact with interzonal microtubule bundles. This prevents the structural transition that allows the contractile ring to pull away from the side of the embryo opposite the primary furrow. The furrow fails to close completely and cytokinesis fails.
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fig9: Schematic illustrating the asymmetric furrow ingression in wild-type and car-1(RNAi) embryos. End-on and side views are shown. (A) The contractile ring is positioned asymmetrically within the circumference of the cell equator. In a side view, a primary furrow is observed coming in from one side of the embryo. Contact between the primary furrow and the interzonal microtubule bundles between the separated chromosomes triggers the other side of the contractile ring (the secondary furrow) to pull away from the edge of the embryo, and the contractile ring closes down around the compacted interzonal microtubule bundles (the midbody). (B) In car-1(RNAi) embryos, the primary furrow ingresses, but fails to come into contact with interzonal microtubule bundles. This prevents the structural transition that allows the contractile ring to pull away from the side of the embryo opposite the primary furrow. The furrow fails to close completely and cytokinesis fails.

Mentions: To analyze the cytokinesis defect in CAR-1–depleted embryos, we constructed a C. elegans strain expressing a fusion of GFP with a pleckstrin homology (PH) domain derived from mammalian PLC1δ1 to monitor cleavage furrow ingression (Fig. 5). The PLC1δ1 PH domain binds with high affinity to a phosphoinositide lipid (PI4,5P2) that is generated on the plasma membrane in most cell types (for review see Hurley and Meyer, 2001). Imaging of GFP:PHPLC1δ1 in wild-type embryos revealed a striking asymmetry in cleavage furrow ingression. A primary furrow was observed initially coming in from one side of the embryo (yellow line on kymograph in Fig. 5 C; Video 5). As the primary furrow ceased its ingression, a secondary furrow began to come in from the opposite side of the embryo (pink line on kymograph in Fig. 5 C). Simultaneous visualization of microtubules and the plasma membrane (Video 6) revealed that the secondary furrow began to ingress when the primary furrow came into contact with the interzonal microtubule bundles that form between the separating chromosomes (see Fig. 9 for a schematic of this transition). After ingression of the secondary furrow slows, a small gap remains that gradually closes (Fig. 5, B and C). This asymmetric ingression is not a consequence of embryo compression (unpublished data).


A complex containing the Sm protein CAR-1 and the RNA helicase CGH-1 is required for embryonic cytokinesis in Caenorhabditis elegans.

Audhya A, Hyndman F, McLeod IX, Maddox AS, Yates JR, Desai A, Oegema K - J. Cell Biol. (2005)

Schematic illustrating the asymmetric furrow ingression in wild-type and car-1(RNAi) embryos. End-on and side views are shown. (A) The contractile ring is positioned asymmetrically within the circumference of the cell equator. In a side view, a primary furrow is observed coming in from one side of the embryo. Contact between the primary furrow and the interzonal microtubule bundles between the separated chromosomes triggers the other side of the contractile ring (the secondary furrow) to pull away from the edge of the embryo, and the contractile ring closes down around the compacted interzonal microtubule bundles (the midbody). (B) In car-1(RNAi) embryos, the primary furrow ingresses, but fails to come into contact with interzonal microtubule bundles. This prevents the structural transition that allows the contractile ring to pull away from the side of the embryo opposite the primary furrow. The furrow fails to close completely and cytokinesis fails.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2171198&req=5

fig9: Schematic illustrating the asymmetric furrow ingression in wild-type and car-1(RNAi) embryos. End-on and side views are shown. (A) The contractile ring is positioned asymmetrically within the circumference of the cell equator. In a side view, a primary furrow is observed coming in from one side of the embryo. Contact between the primary furrow and the interzonal microtubule bundles between the separated chromosomes triggers the other side of the contractile ring (the secondary furrow) to pull away from the edge of the embryo, and the contractile ring closes down around the compacted interzonal microtubule bundles (the midbody). (B) In car-1(RNAi) embryos, the primary furrow ingresses, but fails to come into contact with interzonal microtubule bundles. This prevents the structural transition that allows the contractile ring to pull away from the side of the embryo opposite the primary furrow. The furrow fails to close completely and cytokinesis fails.
Mentions: To analyze the cytokinesis defect in CAR-1–depleted embryos, we constructed a C. elegans strain expressing a fusion of GFP with a pleckstrin homology (PH) domain derived from mammalian PLC1δ1 to monitor cleavage furrow ingression (Fig. 5). The PLC1δ1 PH domain binds with high affinity to a phosphoinositide lipid (PI4,5P2) that is generated on the plasma membrane in most cell types (for review see Hurley and Meyer, 2001). Imaging of GFP:PHPLC1δ1 in wild-type embryos revealed a striking asymmetry in cleavage furrow ingression. A primary furrow was observed initially coming in from one side of the embryo (yellow line on kymograph in Fig. 5 C; Video 5). As the primary furrow ceased its ingression, a secondary furrow began to come in from the opposite side of the embryo (pink line on kymograph in Fig. 5 C). Simultaneous visualization of microtubules and the plasma membrane (Video 6) revealed that the secondary furrow began to ingress when the primary furrow came into contact with the interzonal microtubule bundles that form between the separating chromosomes (see Fig. 9 for a schematic of this transition). After ingression of the secondary furrow slows, a small gap remains that gradually closes (Fig. 5, B and C). This asymmetric ingression is not a consequence of embryo compression (unpublished data).

Bottom Line: Inhibition of CAR-1 by RNA-mediated depletion or mutation results in a specific defect in embryonic cytokinesis.This cytokinesis failure likely results from an anaphase spindle defect in which interzonal microtubule bundles that recruit Aurora B kinase and the kinesin, ZEN-4, fail to form between the separating chromosomes.Cumulatively, our results suggest that CAR-1 functions with CGH-1 to regulate a specific set of maternally loaded RNAs that is required for anaphase spindle structure and cytokinesis.

View Article: PubMed Central - PubMed

Affiliation: Ludwig Institute for Cancer Research, Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA. aaudhya@ucsd.edu

ABSTRACT
Cytokinesis completes cell division and partitions the contents of one cell to the two daughter cells. Here we characterize CAR-1, a predicted RNA binding protein that is implicated in cytokinesis. CAR-1 localizes to germline-specific RNA-containing particles and copurifies with the essential RNA helicase, CGH-1, in an RNA-dependent fashion. The atypical Sm domain of CAR-1, which directly binds RNA, is dispensable for CAR-1 localization, but is critical for its function. Inhibition of CAR-1 by RNA-mediated depletion or mutation results in a specific defect in embryonic cytokinesis. This cytokinesis failure likely results from an anaphase spindle defect in which interzonal microtubule bundles that recruit Aurora B kinase and the kinesin, ZEN-4, fail to form between the separating chromosomes. Depletion of CGH-1 results in sterility, but partially depleted worms produce embryos that exhibit the CAR-1-depletion phenotype. Cumulatively, our results suggest that CAR-1 functions with CGH-1 to regulate a specific set of maternally loaded RNAs that is required for anaphase spindle structure and cytokinesis.

Show MeSH
Related in: MedlinePlus