Huntingtin’s Function in Axonal Transport Is Conserved in Drosophila melanogaster

Abstract

Huntington’s disease (HD) is a devastating dominantly inherited neurodegenerative disorder caused by an abnormal polyglutamine expansion in the N-terminal part of the huntingtin (HTT) protein. HTT is a large scaffold protein that interacts with more than a hundred proteins and is probably involved in several cellular functions. The mutation is dominant, and is thought to confer new and toxic functions to the protein. However, there is emerging evidence that the mutation also alters HTT’s normal functions. Therefore, HD models need to recapitulate this duality if they are to be relevant. Drosophila melanogaster is a useful in vivo model, widely used to study HD through the overexpression of full-length or N-terminal fragments of mutant human HTT. However, it is unclear whether Drosophila huntingtin (DmHTT) shares functions similar to the mammalian HTT. Here, we used various complementary approaches to analyze the function of DmHTT in fast axonal transport. We show that DmHTT interacts with the molecular motor dynein, associates with vesicles and co-sediments with microtubules. DmHTT co-localizes with Brain-derived neurotrophic factor (BDNF)-containing vesicles in rat cortical neurons and partially replaces mammalian HTT in a fast axonal transport assay. DmHTT-KO flies show a reduced fast axonal transport of synaptotagmin vesicles in motoneurons in vivo. These results suggest that the function of HTT in axonal transport is conserved between flies and mammals. Our study therefore validates Drosophila melanogaster as a model to study HTT function, and its dysfunction associated with HD.

Self-propelling vesicles define glycolysis as the minimal energy machinery for neuronal transport

“…We next immunostained all the glycolytic enzymes on rat primary cortical neurons that were grown in microfluidic devices, to separate axons from cell bodies and dendrites3132, allowing us to specifically assess their localization in axons (Fig. 2c).…”

Potential function for the Huntingtin protein as a scaffold for selective autophagy

“…A defect in selective autophagic clearance of the ironbinding protein ferritin (ferritinophagy) with loss of HTT function in HD patients and mice may therefore reflect an impairment in ferritinophagy (13,44,45). Both mutant HTT expression and HTT knockdown are found to impair axonal trafficking of vesicles, mitochondria, and autophagosomes in neurons in vitro and in vivo (7)(8)(9)12), consistent with a loss of Atg11/Atg23-like membrane trafficking.…”

“…Loss of HTT function leads to defects in mobility and ultimately survival as the adult flies age, and to a mitotic spindle misorientation, which is also observed in the conditional mouse neuronal HTT knockout. Drosophila HTT expression in mouse cells treated with mouse Htt siRNA can compensate for the loss of mouse HTT, demonstrating functional conservation of the mitotic spindle and axonal transport roles of HTT from these widely divergent species (11,12). In zebrafish, depletion of HTT produces symptoms of cellular iron deficiency, showing a requirement for HTT in cellular iron utilization (13), whereas knockout of HTT in Dictyostelium discoideum demonstrates it is needed for survival when nutrients are limiting (14).…”

Using Drosophila models of Huntington's disease as a translatable tool

“…Bi-directional transport of YFP::Rab11 associated vesicles was significantly slowed in larval neurons, while the transport rate of YFP::Rab5 associated vesicles was unaffected. Similar experiments in larval axons have indicated that DmHtt is required for maintaining the transport rate of vesicles containing amyloid precursor protein and synaptotagmin (Gunawardena et al, 2003;Zala et al, 2013). Several methods and tools have been established in Drosophila that will aid in dissecting the role of Htt in vesicle trafficking.…”

“…Rather, their development is grossly normal. However, closer analysis shows that the transport rate of synaptic vesicles is significantly slower in DmHtt KO larval neurons (Zala et al, 2013).…”