a: Immature female

a: Immature female. divide during the sequence, although a slight slowing and desynchronization of the regular division cycles in observed. The right embryo shows a stronger effect with some cells showing a marked division delay and detaching from the embryo.(2.83 MB MOV) pone.0013994.s004.mov (2.6M) GUID:?50DEF148-8D4C-44A4-9178-CC85692ED89F Movie S2: Comparison of gastrulation in an uninjected embryo (left) and one developed from an egg injected with 2mM Poc1-MO (right) filmed in parallel. The sequence covers 19 hours with images acquired using DIC optics every 4 minutes. The embryos were filmed in agarose wells to retain them in the observation field, but continue to swim and rotate around the embryonic polarity axis by ciliary beating. Gastrulation proceeds by cell ingression from the oral pole (top) in both embryos, but is much delayed and more irregular in the Poc1-MO embryo. Note that some undivided cells are already present in the blastocoel of the Poc1-MO embryo prior to gastrulation, and further accumulate during the period of the film, likely contributing to the gastrulation troubles.(17.24 MB MOV) pone.0013994.s005.mov (16M) GUID:?ADE31A5D-83BE-4CCE-B417-81A9131C2FA6 Abstract Poc1 (Protein of Centriole 1) proteins are highly conserved WD40 domain-containing centriole components, well characterized in the alga the ciliated protazoan the insect and in vertebrate cells including and zebrafish embryos. Functions and localizations related to the centriole and ciliary axoneme have been exhibited for Poc1 in a TAK-981 range of species. The vertebrate Poc1 protein has also been reported to show an additional association with mitochondria, including enrichment in the specialized germ plasm region of oocytes. We have identified and characterized a highly conserved Poc1 protein in the cnidarian Poc1 mRNA was found to be strongly expressed in eggs and early embryos, showing a punctate perinuclear localization in young oocytes. Fluorescence-tagged Poc1 proteins expressed in developing embryos showed strong localization to centrioles, including basal bodies. Anti-human Poc1 antibodies TAK-981 decorated mitochondria in Poc1. Injection of specific morpholino oligonucleotides into eggs prior to fertilization to repress Poc1 mRNA translation interfered with cell division from the blastula stage, likely corresponding to when neosynthesis normally takes over from maternally supplied protein. Cell cycle lengthening and arrest were observed, phenotypes consistent with an impaired centriolar biogenesis or function. The specificity of the defects could be exhibited by injection of synthetic Poc1 mRNA, which restored normal development. We conclude that in embryos, Poc1 has an essentially centriolar localization and function. Introduction IFI30 Centrioles are intruiging cellular organelles that have fascinated biologists for over a century. They are present in a wide range of eukaryotes including the vast majority of animal cells, and appear to act as organizers not only for the mitotic spindle but also for the interphase microtubule cytoskeleton, TAK-981 thereby playing key role in the positioning and traffic of intracellular organelles. There is thus intense interest in characteristing the structure and composition of the centriole, and in understanding how it interacts with other cellular components. The WD40 repeat-containing protein Poc1 is an integral component of the centriole and is required for basal body stability and cilia formation. It has been exceptionally well conserved through eukaryotic evolution, being identified in most species in a wide genome survey [1], and has been consistently identified in centriole proteomics studies, for instance basal bodies of the unicellular alga Poc1 proteins has suggested that they may have additional localizations outside the centriole/axoneme. In human cells Poc1a and Poc1b (?=? Pix2 and Pix1 in eggs Poc1 proteins were detected in a particular region in the oocyte rich in mitochondria known as germ plasm [8]. Germ plasm is usually a characteristic component of many animal oocytes, often.