scholarly journals PCMD-1 bridges the centrioles and the PCM scaffold in C. elegans

Development ◽  
2021 ◽  
Author(s):  
Lisa Stenzel ◽  
Alina Schreiner ◽  
Elisa Zuccoli ◽  
Sim Üstüner ◽  
Judith Mehler ◽  
...  
Keyword(s):  
Coiled Coil ◽  
Scaffold Proteins ◽  
Animal Cells ◽  
C Elegans ◽  
Mitotic Kinase ◽  
Spindle Poles ◽  
Coiled Coil Domain ◽  
Pericentriolar Material ◽  
Translocation Assay ◽  
Assembly Proteins

Correct cell division relies on the formation of a bipolar spindle. In animal cells, microtubule nucleation at the spindle poles is facilitated by the pericentriolar material (PCM), which assembles around a pair of centrioles. Although centrioles are essential for PCM assembly, proteins that anchor the PCM to the centrioles are less known. Here we investigate the molecular function of PCMD-1 in bridging the PCM and the centrioles in Caenorhabditis elegans. We demonstrate that the centrosomal recruitment of PCMD-1 is dependent on the outer centriolar protein SAS-7. While the most C-terminal part of PCMD-1 is sufficient to target it to the centrosome, the coiled-coil domain promotes its accumulation by facilitating self-interaction. We reveal that PCMD-1 is interacting with the PCM scaffold protein SPD-5, the mitotic kinase PLK-1 and the centriolar protein SAS-4. Using an ectopic translocation assay, we show that PCMD-1 can selectively recruit downstream PCM scaffold components to an ectopic location in the cell, indicating that PCMD-1 is able to anchor the PCM scaffold proteins at the centrioles. Our work suggests that PCMD-1 is an essential functional bridge between the centrioles and the PCM.

scholarly journals PCMD-1 bridges the centrioles and the PCM scaffold in C. elegans

2020 ◽  
Author(s):  
Lisa Stenzel ◽  
Judith Mehler ◽  
Alina Schreiner ◽  
Sim Üstüner ◽  
Elisa Zuccoli ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Coiled Coil ◽  
Protein Protein Interactions ◽  
Animal Cells ◽  
C Elegans ◽  
Mitotic Kinase ◽  
Spindle Poles ◽  
Pericentriolar Material ◽  
Translocation Assay ◽  
Assembly Proteins

ABSTRACTCorrect cell division relies on the formation of a bipolar spindle. In animal cells, microtubule nucleation at the spindle poles is facilitated by the pericentriolar material (PCM), which assembles around a pair of centrioles. Although centrioles are essential for PCM assembly, proteins that anchor the PCM to the centrioles are less known. Here we investigate the molecular function of PCMD-1 in bridging the PCM and the centrioles in Caenorhabditis elegans.We demonstrate that centrosomal recruitment of PCMD-1 is dependent on the outer centriolar protein SAS-7. While the most C-terminal part of PCMD-1 is sufficient to target it to the centrosome, the coiled-coil domain promotes its accumulation by facilitating self-interaction. We reveal that PCMD-1 is bridging the centrioles and PCM scaffold through protein-protein interactions with the PCM scaffold protein SPD-5, the mitotic kinase PLK-1 and the centriolar protein SAS-4. Using an ectopic translocation assay, we show that PCMD-1 is able to selectively recruit downstream PCM scaffold components to an ectopic location in the cell, indicating that PCMD-1 is sufficient to anchor the PCM scaffold proteins to the centrioles. Our work suggests that PCMD-1 is an essential functional bridge between the centrioles and the PCM.

scholarly journals α-/γ-Taxilin are required for centriolar subdistal appendage assembly and microtubule organization

2021 ◽  
Author(s):  
Dandan Ma ◽  
Rongyi Wang ◽  
Fulin Wang ◽  
Zhiquan Chen ◽  
Ning Huang ◽  
...  
Keyword(s):  
Coiled Coil ◽  
Animal Cells ◽  
Microtubule Organization ◽  
Hierarchical Assembly ◽  
Mother And Daughter ◽  
Mother Centriole ◽  
Functional Understanding ◽  
Pericentriolar Material ◽  
Functional Analyses ◽  
Molecular Components

AbstractThe centrosome, composed of a pair of centrioles (mother and daughter centrioles) and pericentriolar material, is mainly responsible for microtubule nucleation and anchorage in animal cells. The subdistal appendage (SDA) is a centriolar structure located at the subdistal region on the mother centriole, and it functions in microtubule anchorage. However, the molecular composition and detailed structure of SDA remain largely unknown. Here, we identified a-taxilin and r-taxilin as new SDA components, which form a complex via their coiled-coil domains and serve as a new subgroup during SDA hierarchical assembly. Their SDA localization is dependent on ODF2, and α-taxilin recruits CEP170 to the SDA. Functional analyses suggest that α-taxilin and γ-taxilin are responsible for centrosomal microtubule anchorage during interphase, as well as for proper spindle orientation during metaphase. Altogether, our results shed light on the molecular components and functional understanding of the SDA hierarchical assembly and microtubule organization.

scholarly journals Centriole and PCM cooperatively recruit CEP192 to spindle poles to promote bipolar spindle assembly

2021 ◽  
Vol 220 (2) ◽  
Author(s):  
Takumi Chinen ◽  
Kaho Yamazaki ◽  
Kaho Hashimoto ◽  
Ken Fujii ◽  
Koki Watanabe ◽  
...  
Keyword(s):  
A549 Cells ◽  
Spindle Assembly ◽  
Spindle Pole ◽  
Scaffold Proteins ◽  
Spindle Formation ◽  
Bipolar Spindle ◽  
Spindle Poles ◽  
Pericentriolar Material ◽  
Mitotic Cells

The pericentriolar material (PCM) that accumulates around the centriole expands during mitosis and nucleates microtubules. Here, we show the cooperative roles of the centriole and PCM scaffold proteins, pericentrin and CDK5RAP2, in the recruitment of CEP192 to spindle poles during mitosis. Systematic depletion of PCM proteins revealed that CEP192, but not pericentrin and/or CDK5RAP2, was crucial for bipolar spindle assembly in HeLa, RPE1, and A549 cells with centrioles. Upon double depletion of pericentrin and CDK5RAP2, CEP192 that remained at centriole walls was sufficient for bipolar spindle formation. In contrast, through centriole removal, we found that pericentrin and CDK5RAP2 recruited CEP192 at the acentriolar spindle pole and facilitated bipolar spindle formation in mitotic cells with one centrosome. Furthermore, the perturbation of PLK1, a critical kinase for PCM assembly, efficiently suppressed bipolar spindle formation in mitotic cells with one centrosome. Overall, these data suggest that the centriole and PCM scaffold proteins cooperatively recruit CEP192 to spindle poles and facilitate bipolar spindle formation.

scholarly journals The Caenorhabditis elegans pericentriolar material components SPD-2 and SPD-5 are monomeric in the cytoplasm before incorporation into the PCM matrix

2014 ◽  
Vol 25 (19) ◽  
pp. 2984-2992 ◽  
Author(s):  
Oliver Wueseke ◽  
Jakob Bunkenborg ◽  
Marco Y. Hein ◽  
Andrea Zinke ◽  
Valeria Viscardi ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Fluorescence Correlation Spectroscopy ◽  
Coiled Coil ◽  
Correlation Spectroscopy ◽  
Regulatory Proteins ◽  
Native State ◽  
Animal Cells ◽  
Microtubule Organization ◽  
Fluorescence Correlation ◽  
Pericentriolar Material

Centrosomes are the main microtubule-organizing centers in animal cells. Centrosomes consist of a pair of centrioles surrounded by a matrix of pericentriolar material (PCM) that assembles from cytoplasmic components. In Caenorhabditis elegans embryos, interactions between the coiled-coil proteins SPD-5 and SPD-2 and the kinase PLK-1 are critical for PCM assembly. However, it is not known whether these interactions promote the formation of cytoplasmic complexes that are added to the PCM or whether the components interact only during incorporation into the PCM matrix. Here we address this problem by using a combination of live-cell fluorescence correlation spectroscopy, mass spectrometry, and hydrodynamic techniques to investigate the native state of PCM components in the cytoplasm. We show that SPD-2 is monomeric, and neither SPD-2 nor SPD-5 exists in complex with PLK-1. SPD-5 exists mostly as a monomer but also forms complexes with the PP2A-regulatory proteins RSA-1 and RSA-2, which are required for microtubule organization at centrosomes. These results suggest that the interactions between SPD-2, SPD-5, and PLK-1 do not result in formation of cytoplasmic complexes, but instead occur in the context of PCM assembly.

scholarly journals Co-translational protein targeting facilitates centrosomal recruitment of PCNT during centrosome maturation in vertebrates

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Guadalupe Sepulveda ◽  
Mark Antkowiak ◽  
Ingrid Brust-Mascher ◽  
Karan Mahe ◽  
Tingyoung Ou ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Protein Targeting ◽  
Mrna Localization ◽  
Cytoplasmic Dynein ◽  
Animal Cells ◽  
Bipolar Spindle ◽  
Spindle Poles ◽  
Cytoplasmic Proteins ◽  
Pericentriolar Material ◽  
Early Mitosis

As microtubule-organizing centers of animal cells, centrosomes guide the formation of the bipolar spindle that segregates chromosomes during mitosis. At mitosis onset, centrosomes maximize microtubule-organizing activity by rapidly expanding the pericentriolar material (PCM). This process is in part driven by the large PCM protein pericentrin (PCNT), as its level increases at the PCM and helps recruit additional PCM components. However, the mechanism underlying the timely centrosomal enrichment of PCNT remains unclear. Here, we show that PCNT is delivered co-translationally to centrosomes during early mitosis by cytoplasmic dynein, as evidenced by centrosomal enrichment of PCNT mRNA, its translation near centrosomes, and requirement of intact polysomes for PCNT mRNA localization. Additionally, the microtubule minus-end regulator, ASPM, is also targeted co-translationally to mitotic spindle poles. Together, these findings suggest that co-translational targeting of cytoplasmic proteins to specific subcellular destinations may be a generalized protein targeting mechanism.

scholarly journals Cell polarity–dependent centrosome separation in the C. elegans embryo

2019 ◽  
Vol 218 (12) ◽  
pp. 4112-4126
Author(s):  
Alexandra Bondaz ◽  
Luca Cirillo ◽  
Patrick Meraldi ◽  
Monica Gotta
Keyword(s):  
Cell Polarity ◽  
Cell Biology ◽  
Dependent Manner ◽  
Animal Cells ◽  
Cell Stage ◽  
C Elegans ◽  
Mitotic Kinase ◽  
Centrosome Separation ◽  
A Cell ◽  
Cortical Localization

In animal cells, faithful chromosome segregation depends on the assembly of a bipolar spindle driven by the timely separation of the two centrosomes. Here we took advantage of the highly stereotypical cell divisions in Caenorhabditis elegans embryos to identify new regulators of centrosome separation. We find that at the two-cell stage, the somatic AB cell initiates centrosome separation later than the germline P1 cell. This difference is strongly exacerbated by the depletion of the kinesin-13 KLP-7/MCAK, resulting in incomplete centrosome separation at NEBD in AB but not P1. Our genetic and cell biology data indicate that this phenotype depends on cell polarity via the enrichment in AB of the mitotic kinase PLK-1, which itself limits the cortical localization of the dynein-binding NuMA orthologue LIN-5. We postulate that the timely separation of centrosomes is regulated in a cell type–dependent manner.

scholarly journals Centriolar SAS-7 acts upstream of SPD-2 to regulate centriole assembly and pericentriolar material formation

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Kenji Sugioka ◽  
Danielle R Hamill ◽  
Joshua B Lowry ◽  
Marie E McNeely ◽  
Molly Enrick ◽  
...  
Keyword(s):  
Cell Polarity ◽  
Molecular Basis ◽  
Coiled Coil ◽  
Genetic Data ◽  
C Elegans ◽  
Centriole Duplication ◽  
Pericentriolar Material ◽  
Daughter Centriole ◽  
Material Formation ◽  
Anterior Posterior

The centriole/basal body is a eukaryotic organelle that plays essential roles in cell division and signaling. Among five known core centriole proteins, SPD-2/Cep192 is the first recruited to the site of daughter centriole formation and regulates the centriolar localization of the other components in C. elegans and in humans. However, the molecular basis for SPD-2 centriolar localization remains unknown. Here, we describe a new centriole component, the coiled-coil protein SAS-7, as a regulator of centriole duplication, assembly and elongation. Intriguingly, our genetic data suggest that SAS-7 is required for daughter centrioles to become competent for duplication, and for mother centrioles to maintain this competence. We also show that SAS-7 binds SPD-2 and regulates SPD-2 centriolar recruitment, while SAS-7 centriolar localization is SPD-2-independent. Furthermore, pericentriolar material (PCM) formation is abnormal in sas-7 mutants, and the PCM-dependent induction of cell polarity that defines the anterior-posterior body axis frequently fails. We conclude that SAS-7 functions at the earliest step in centriole duplication yet identified and plays important roles in the orchestration of centriole and PCM assembly.

scholarly journals Co-translational protein targeting facilitates centrosomal recruitment of PCNT during centrosome maturation

2017 ◽  
Author(s):  
Guadalupe Sepulveda ◽  
Mark Antkowiak ◽  
Ingrid Brust-Mascher ◽  
Karan Mahe ◽  
Tingyoung Ou ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Protein Targeting ◽  
Mrna Localization ◽  
Cytoplasmic Dynein ◽  
Animal Cells ◽  
Bipolar Spindle ◽  
Spindle Poles ◽  
Cytoplasmic Proteins ◽  
Pericentriolar Material ◽  
Early Mitosis

AbstractAs microtubule-organizing centers of animal cells, centrosomes guide the formation of the bipolar spindle that segregates chromosomes during mitosis. At mitosis onset, centrosomes maximize microtubule-organizing activity by rapidly expanding the pericentriolar material (PCM). This process is in part driven by the large PCM protein pericentrin (PCNT), as its level increases at the PCM and helps recruit additional PCM components. However, the mechanism underlying the timely centrosomal enrichment of PCNT remains unclear. Here we show that PCNT is delivered co-translationally to centrosomes during early mitosis by cytoplasmic dynein, as evidenced by centrosomal enrichment of PCNT mRNA, its translation near the centrosome, and requirement of intact polysomes for PCNT mRNA localization. Additionally, the microtubule minus-end regulator, ASPM, is also targeted co-translationally to mitotic spindle poles. Together, these findings suggest that co-translational targeting of cytoplasmic proteins to specific subcellular destinations may be a generalized protein targeting mechanism.

scholarly journals Centriole-less pericentriolar material serves as a microtubule organizing center at the base of C. elegans sensory cilia

2020 ◽  
Author(s):  
Jérémy Magescas ◽  
Sani Eskinazi ◽  
Michael V. Tran ◽  
Jessica L. Feldman
Keyword(s):  
Sensory Neurons ◽  
Super Resolution ◽  
Spatial Separation ◽  
Animal Cells ◽  
Microtubule Organizing Center ◽  
Tissue Specific ◽  
C Elegans ◽  
Organizing Center ◽  
Pericentriolar Material ◽  
Specific Degradation

SummaryDuring mitosis in animal cells, the centrosome acts as a microtubule organizing center (MTOC) to assemble the mitotic spindle. MTOC function at the centrosome is driven by proteins within the pericentriolar material (PCM), however the molecular complexity of the PCM makes it difficult to differentiate the proteins required for MTOC activity from other centrosomal functions. We used the natural spatial separation of PCM proteins during mitotic exit to identify a minimal module of proteins required for centrosomal MTOC function in C. elegans. Using tissue specific degradation, we show that SPD-5, the functional homolog of CDK5RAP2, is essential for embryonic mitosis while SPD-2/CEP192 and PCMD-1, which are essential in the zygote, are dispensable. Surprisingly, although the centriole is known to be degraded in the ciliated sensory neurons in C. elegans [1-3], we find evidence for “centriole-less PCM” at the base of cilia and use this structure as a minimal testbed to dissect centrosomal MTOC function. Super-resolution imaging revealed that this PCM inserts inside the lumen of the ciliary axoneme and directly nucleates the assembly of dendritic microtubules towards the cell body. Tissue-specific degradation in ciliated sensory neurons revealed a role for SPD-5 and the conserved microtubule nucleator γ-TuRC, but not SPD-2 or PCMD-1, in MTOC function at centriole-less PCM. This MTOC function was in the absence of regulation by mitotic kinases, highlighting the intrinsic ability of these proteins to drive microtubule growth and organization and further supporting a model that SPD-5 is the primary driver of MTOC function at the PCM.

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