Abstract
The centrosome is the primary microtubule organising centre of the cell. It is composed of many proteins, some of which make up the core of the centrosome, whereas others are used for specific functions. Although the cellular roles of many centrosomal proteins are well defined, much less is known about their functions and the role of the centrosome in development. In this study we investigated the function of NEDD1, a critical component of the centrosome essential for microtubule nucleation, in zebrafish (Danio rerio) development. The zebrafish homologue of NEDD1 (zNEDD1) was cloned and found to have a similar localisation and function to mammalian NEDD1. We show that zNEDD1 is essential for survival, as a high level of knockdown was embryonic lethal. Partial knockdown of zNEDD1 caused abnormalities including an increase in mitotic and apoptotic cells. Pronounced phenotypic defects were seen in the brain, with a lack of defined brain structures, incomplete neural tube formation and a disorganisation of neurons. In addition, we show that a reduction in zNEDD1 resulted in the loss of γ-tubulin at the centrosome. Our data thus demonstrate that zNEDD1 is critical for the recruitment of γ-tubulin to the centrosome, and is essential for the proper development of zebrafish.
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Accession codes
Abbreviations
- γ-TuRC:
-
γ-tubulin ring complex
- AP:
-
alkaline phosphatase
- DIC:
-
differential interference contrast
- DIG:
-
digoxigenin
- DTT:
-
1, 4-dithiothreitol
- EDTA:
-
ethylenediaminetetra-acetic acid
- GST:
-
glutathione-S-transferase
- hpf:
-
hours post fertilisation
- HRP:
-
horseradish peroxidase
- MO:
-
morpholino
- MTOC:
-
microtubule organising centre
- ORF:
-
open reading frame
- PBS:
-
phosphate-buffered saline
- PBST:
-
PBS-Tween20
- PFA:
-
paraformaldehyde
- pH3:
-
phosphorylated histone H3
- PVDF:
-
polyvinylidene difluoride
- RB:
-
Rohon–Beard
- RT:
-
room temperature
- SDS:
-
sodium dodecyl sulphate
- SDS-PAGE:
-
SDS-polyacrylamide gel electrophoresis
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Acknowledgements
This work was supported in part by an Australian Postgraduate Award to JAM and a NHMRC Senior Principal Research Fellowship to SK.
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Manning, J., Lewis, M., Koblar, S. et al. An essential function for the centrosomal protein NEDD1 in zebrafish development. Cell Death Differ 17, 1302–1314 (2010). https://doi.org/10.1038/cdd.2010.12
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DOI: https://doi.org/10.1038/cdd.2010.12
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