The endocycle promotes aneuploidy at both ends of the spindle. Donald T. Fox^1,2, Ruth Montague¹, Kevin Schoenfelder¹, Benjamin Stormo², Sarah Paramore¹. 1) Department of Pharmacology & Cancer Biology, Duke University Medical Center, Durham, NC; 2) Department of Cell Biology, Duke University Medical Center, Durham, NC.

   Polyploidy (extra chromosome sets) is a common property of both normal and cancerous animal cells. In cancer cells, polyploidy is linked to increased cell division errors, or chromosomal instability (CIN). Similarly, our study of Drosophila hindgut papillar formation found links between polyploid cell division and CIN. During hindgut metamorphosis, polyploid cells formed via the endocycle re-enter mitosis. As in mitosis of polyploid cancer cells, these papillar precursor divisions also exhibit CIN, leading to aneuploidy (cells with unbalanced chromosome content). Thus, CIN is a common property of polyploid cells, but mechanisms connecting polyploidy to CIN remain unknown. Through further characterization of papillar formation in flies, we now report two distinct mechanisms linking polyploidy via endocycles to CIN. First, we find the endocycle impairs localization of the spindle checkpoint regulator Mad2 to chromosomes. Using time-lapse imaging, we find Mad2 mis-localization coincides with chromosome mis-alignment defects and aneuploidy during papillar mitosis, indicative of failure in the Mad2-dependent checkpoint in anaphase delay. Second, we find polyploid papillar cells accumulate extra spindle poles, which fail to cluster into a bipolar spindle during anaphase. As a result, papillar development proceeds with frequent multipolar division. Contrary to multipolar division in diploid cells, we find multipolar divisions in polyploid cells frequently yield viable aneuploid daughters that contribute to organogenesis. Taken together, our study of papillar mitosis has identified two primary mechanisms by which endocycles disrupt faithful cell division: one at the chromosome end of the spindle (Mad2 mislocalization) and one at the spindle pole (failed pole clustering). Given recent findings that the endocycle induces aneuploidy and tumorigenesis in mammalian cells, our findings have potential implications for how the endocycle contributes to aneuploidy in cancer.