In this study we provide evidence that the spindle matrix protein Skeletor in Drosophila interacts with the human ASCIZ (also known as ATMIN and ZNF822) ortholog, Digitor/dASCIZ. This interaction was first detected in a yeast two-hybrid screen and subsequently confirmed by pull-down assays. We also confirm a previously documented function of Digitor/dASCIZ as a regulator of Dynein light chain/Cut up expression. Using transgenic expression of a mCitrine-labeled Digitor construct, we show that Digitor/dASCIZ is a nuclear protein that is localized to interband and developmental puff chromosomal regions during interphase but redistributes to the spindle region during mitosis. Its mitotic localization and physical interaction with Skeletor suggest the possibility that Digitor/dASCIZ plays a direct role in mitotic progression as a member of the spindle matrix complex. Furthermore, we have characterized a P-element insertion that is likely to be a true null Digitor/dASCIZ allele resulting in complete pupal lethality when homozygous, indicating that Digitor/dASCIZ is an essential gene. Phenotypic analysis of the mutant provided evidence that Digitor/dASCIZ plays critical roles in regulation of metamorphosis and organogenesis as well as in the DNA damage response. In the Digitor/dASCIZ null mutant larvae there was greatly elevated levels of γH2Av, indicating accumulation of DNA double-strand breaks. Furthermore, reduced levels of Digitor/dASCIZ decreased the resistance to paraquat-induced oxidative stress resulting in increased mortality in a stress test paradigm. We show that an early developmental consequence of the absence of Digitor/dASCIZ is reduced third instar larval brain size although overall larval development appeared otherwise normal at this stage. While Digitor/dASCIZ mutant larvae initiate pupation, all mutant pupae failed to eclose and exhibited various defects in metamorphosis such as impaired differentiation, incomplete disc eversion, and faulty apoptosis. Altogether we provide evidence that Digitor/dASCIZ is a nuclear protein that performs multiple roles in Drosophila larval and pupal development.
Pubmed ID: 27861562 RIS Download
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View all literature mentionsCell line ND12577 is a Transformed cell line with a species of origin Homo sapiens
View all literature mentionsDrosophila melanogaster with name y[1] w[*]; P{w[+mC]=Act5C-GAL4}25FO1/CyO, y[+] from BDSC.
View all literature mentionsDrosophila melanogaster with name Canton-S from BDSC.
View all literature mentionsThis monoclonal targets histone 2A gamma variant, phosphorylated
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View all literature mentionsThis polyclonal targets Phospho-Histone H3 (Ser10)
View all literature mentionsThis monoclonal targets HA
View all literature mentionsThis monoclonal targets RNA Polymerase II H14
View all literature mentionsThis unknown targets eGFP
View all literature mentionsThis monoclonal targets α-Tubulin
View all literature mentionsDrosophila melanogaster with name w[1118]; P{w[+mC]=Sgs3-GAL4.PD}TP1 from BDSC.
View all literature mentionsDrosophila melanogaster with name y[1] w[*]; P{w[+mC]=Act5C-GAL4}17bFO1/TM6B, Tb[1] from BDSC.
View all literature mentionsDrosophila melanogaster with name w[*]; P{w[+mC]=matalpha4-GAL-VP16}V2H from BDSC.
View all literature mentionsDrosophila melanogaster with name w[*]; P{w[+mC]=UAS-ChRFP-Tub}2 from BDSC.
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