The Role of Basal Autophagy in Regulating Class Specific Dendrite Morphogenesis in Drosophila Sensory Neurons
Abstract
Dendrites serve as the primary sites for synaptic and sensory neuronal input and impairments in dendrite morphogenesis can result in various neurological diseases such as Rett, Down, and Fragile X Syndromes (Kaufmann & Moser, 2000). Thus, uncovering the cellular and molecular mechanisms that govern dendritogenesis in various neurons is important for advancing our understanding of these diseases. Macroautophagy (hereafter referred to as ‘autophagy’) has emerged as a topic of interest in this area, as postmitotic neurons are known to require high basal autophagy in terms of clearing misfolded and/or damaged organelles. Moreover, defects in basal autophagy have been directly linked to neurodegeneration, including neuronal cell death, axon degeneration, and aberrant synapase development which suggests that autophagy has a neuroprotective function (Shen & Ganetzky, 2009; Wong & Cuervo, 2010; Yang, Coleman, Zhang, Zheng, & Yue, 2013). However, despite a well documented role for autophagy in neuronal survival and axonal development, little is known regarding the role of basal autophagy in mediating dendritic arborization or the mechanisms by which the autophagy pathway may be transcriptionally regulated to mediate differential dendritic homeostasis. Thus, we have investigated these mechanisms by utilizing dendritic arborization (da) neurons in the Drosophila melanogaster peripheral nervous system (PNS), which serve as a powerful model for investigating class specific molecular mechanisms of dendrite development and maintenance (Corty, Matthews, & Grueber, 2009; Jan & Jan, 2010). Preliminary microarray analyses demonstrate that the homeodomain transcription factor Cut positively regulates expression of autophagy related genes (Atg) in da neurons. Due to the native differential expression of Cut across the four da neuron subclasses correlating with distinct levels of dendritic complexity (CI-IV) (Grueber, Jan, & Jan, 2002), the central hypothesis of the studies described herein is that Cut differentially regulates the autophagy pathway as a key mechanism in mediating class specific dendritic homeostasis. By utilizing a combination of in vivo genetics, live confocal microscopy, gene expression analysis, immunohistochemistry, and quantitative neuromorphometry, we investigated the role of basal autophagy in regulating class specific dendrite morphogenesis. Here we establish that basal autophagy is required to promote normal class specific dendrite morphogenesis and that it exerts differential effects across da neuron subclasses. We also demonstrate that key autophagy genes (Atg) are positively regulated, in part, by the transcription factor Cut, and that basal autophagy plays a neuroprotective role in regulating higher order dendritic complexity in neurons expressing Cut, namely CIII and CIV da neurons. Cut-independent effects were also observed in the relatively simple CI neurons, where basal autophagy functions to constrain dendritic arbors. Autophagic dysregulation through loss-of-function (LOF) and gain-of-function (GOF) studies reveals that proper regulation of autophagy is also required for mediating dendritic homeostasis in all classes. In addition, we establish that autophagy regulates the dual leucine zipper kinase (DLK) pathway in CIV da neurons. Collectively, these studies provide invaluable insight into how autophagic regulatory mechanisms are involved in dendritic development and homeostasis, demonstrating that the basal autophagy pathway functions as a downstream effector of Cut-mediated dendritogenesis, but also regulates CI dendritic morphologies independent of Cut. Furthermore, these studies shed novel insight on how defects in autophagy may be implicated in neurodegenerative pathologies.
Description
Keywords
Dendrite, Neuron, Autophagy, Morphogenesis, Drosophila