An optimized temporally controlled Gal4 system in Drosophila reveals degeneration caused by adult-onset neuronal Vps13D knockdown.

Mutations in the human gene VPS13D cause the adult-onset neurodegenerative disease ataxia. Our previous work showed that disruptions in the Vps13D gene in Drosophila neurons causes mitochondrial defects. However, developmental lethality caused by Vps13D loss limited our understanding of the long-term physiological effects of Vps13D perturbation in neurons. Here, we optimized a previously generated system to temporally knock down Vps13D expression precisely in adult Drosophila neurons using a modification to the Gal4/UAS system. Adult-onset activation of Gal4 was enacted using the chemically-inducible tool which fuses a destabilization-domain to the Gal4 repressor Gal80 (Gal80-DD). Optimization of the Gal80-DD tool shows that feeding animals the DD-stabilizing drug trimethoprim (TMP) during development and rearing at a reduced temperature maximally represses Gal4 activity. Temperature shift and removal of TMP from the food after eclosion robustly activates Gal4 expression in adult neurons. Using the optimized Gal80-DD system, we find that adult-onset Vps13D RNAi expression in neurons causes the accumulation of mitophagy intermediates, progressive deficits in locomotor activity, early lethality, and brain vacuolization characteristic of neurodegeneration. The development of this optimized system allows us to more precisely examine the degenerative phenotypes caused by Vps13D disruption, and can likely be utilized in the future for other genes associated with neurological diseases whose manipulation causes developmental lethality in Drosophila.

Misregulation of Nucleoporins 98 and 96 leads to defects in protein synthesis that promote hallmarks of tumorigenesis.

Nucleoporin 98KD (Nup98) is a promiscuous translocation partner in hematological malignancies. Most disease models of Nup98 translocations involve ectopic expression of the fusion protein under study, leaving the endogenous Nup98 loci unperturbed. Overlooked in these approaches is the loss of one copy of normal Nup98 in addition to the loss of Nup96 - a second Nucleoporin encoded within the same mRNA and reading frame as Nup98 - in translocations. Nup98 and Nup96 are also mutated in a number of other cancers, suggesting that their disruption is not limited to blood cancers. We found that reducing Nup98-96 function in Drosophila melanogaster (in which the Nup98-96 shared mRNA and reading frame is conserved) de-regulates the cell cycle. We found evidence of overproliferation in tissues with reduced Nup98-96, counteracted by elevated apoptosis and aberrant signaling associated with chronic wounding. Reducing Nup98-96 function led to defects in protein synthesis that triggered JNK signaling and contributed to hallmarks of tumorigenesis when apoptosis was inhibited. We suggest that partial loss of Nup98-96 function in translocations could de-regulate protein synthesis, leading to signaling that cooperates with other mutations to promote tumorigenesis.

Cell Cycle Re-entry in the Nervous System: From Polyploidy to Neurodegeneration.

Terminally differentiated cells of the nervous system have long been considered to be in a stable non-cycling state and are often considered to be permanently in G0. Exit from the cell cycle during development is often coincident with the differentiation of neurons, and is critical for neuronal function. But what happens in long lived postmitotic tissues that accumulate cell damage or suffer cell loss during aging? In other contexts, cells that are normally non-dividing or postmitotic can or re-enter the cell cycle and begin replicating their DNA to facilitate cellular growth in response to cell loss. This leads to a state called polyploidy, where cells contain multiple copies of the genome. A growing body of literature from several vertebrate and invertebrate model organisms has shown that polyploidy in the nervous system may be more common than previously appreciated and occurs under normal physiological conditions. Moreover, it has been found that neuronal polyploidization can play a protective role when cells are challenged with DNA damage or oxidative stress. By contrast, work over the last two and a half decades has discovered a link between cell-cycle reentry in neurons and several neurodegenerative conditions. In this context, neuronal cell cycle re-entry is widely considered to be aberrant and deleterious to neuronal health. In this review, we highlight historical and emerging reports of polyploidy in the nervous systems of various vertebrate and invertebrate organisms. We discuss the potential functions of polyploidization in the nervous system, particularly in the context of long-lived cells and age-associated polyploidization. Finally, we attempt to reconcile the seemingly disparate associations of neuronal polyploidy with both neurodegeneration and neuroprotection.