Abstract
ABSTRACTThe centrosome-associated proteins Ninein (Nin) and Ninein-like protein (Nlp) play significant roles in microtubule stability, nucleation and anchoring at the centrosome in mammalian cells. Here, we investigate Blastoderm specific gene 25D (Bsg25D), which encodes the only Drosophila protein that is closely related to Nin and Nlp. In early embryos, we find that Bsg25D mRNA and Bsg25D protein are closely associated with centrosomes and astral microtubules. We show that sequences within the coding region and 3′UTR of Bsg25D mRNAs are important for proper localization of this transcript in oogenesis and embryogenesis. Ectopic expression of eGFP-Bsg25D from an unlocalized mRNA disrupts microtubule polarity in mid-oogenesis and compromises the distribution of the axis polarity determinant Gurken. Using total internal reflection fluorescence microscopy, we show that an N-terminal fragment of Bsg25D can bind microtubules in vitro and can move along them, predominantly toward minus-ends. While flies homozygous for a Bsg25D null mutation are viable and fertile, 70% of embryos lacking maternal and zygotic Bsg25D do not hatch and exhibit chromosome segregation defects, as well as detachment of centrosomes from mitotic spindles. We conclude that Bsg25D is a centrosomal protein that, while dispensable for viability, nevertheless helps ensure the integrity of mitotic divisions in Drosophila.
Highlights
Establishment of embryonic patterning in Drosophila melanogaster requires localized translation of numerous maternally deposited mRNAs in specific regions of the embryo during the initial nuclear divisions in the syncytial stage of embryogenesis (Lasko, 2012)
In situ hybridization experiments indicated that Blastoderm specific gene 25D (Bsg25D) mRNA is expressed throughout oogenesis
We found that purified BsgN alone in BRB80 buffer bound efficiently to microtubules, unlike the control protein Dynein light chain 90f (Dlc90f ), which requires Dynein intermediate chain (Dic) and Dynein heavy chain (Dhc) to bind to microtubules (Fig. 5A-D; Movies 1, 2) (Song et al, 2007)
Summary
Establishment of embryonic patterning in Drosophila melanogaster requires localized translation of numerous maternally deposited mRNAs in specific regions of the embryo during the initial nuclear divisions in the syncytial stage of embryogenesis (Lasko, 2012). Received 17 May 2016; Accepted 14 June 2016 transiently accumulate in a perinuclear pattern around the pole cell nuclei during nuclear division 9 in embryogenesis, namely germ cell-less (Jongens et al, 1992), polar granule component (HanyuNakamura et al, 2008), nanos (nos) (Wang and Lehmann, 1991), spire (Dahlgaard et al, 2007), Tao (Sato et al, 2007), arrest (Parisi et al, 2001), exuperantia (Winslow et al, 1988), oo RNA-Binding Protein (Lantz et al, 1994), tramtrack (Read et al, 1992), cyclin B (Kadyrova et al, 2007), and pumilio (Asaoka-Taguchi et al, 1999; Lécuyer et al, 2007) One of these mRNAs, nos, is first anchored to the posterior actin cytoskeleton, and transported to the migrating posterior nuclei by the motor protein Dynein along astral microtubules (Lerit and Gavis, 2011). It is assumed that other mRNAs with the same distribution pattern localize through a similar mechanism, this has not been directly investigated
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