Sexual Dimorphism in Age-Dependent Neurodegeneration After Mild Head Trauma in Drosophila: Unveiling the Adverse Impact of Female Reproductive Signaling.

Environmental insults, including mild head trauma, significantly increase the risk of neurodegeneration. However, it remains challenging to establish a causative connection between early-life exposure to mild head trauma and late-life emergence of neurodegenerative deficits, nor do we know how sex and age compound the outcome. Using a Drosophila model, we demonstrate that exposure to mild head trauma causes neurodegenerative conditions that emerge late in life and disproportionately affect females. Age-at-injury further exacerbates this effect in a sexually dimorphic manner. We further identify Sex Peptide (SP) signaling as a key factor in female susceptibility to post-injury brain deficits. RNA sequencing highlights changes in innate immune defense transcripts specifically in mated females during late life. Our findings establish a causal relationship between early head trauma and late-life neurodegeneration, emphasizing sex differences in injury response and the impact of age-at-injury. Finally, our findings reveal that reproductive signaling adversely impacts female response to mild head insults and elevates vulnerability to late-life neurodegeneration.

Drosophila melanogaster as a model to study age and sex differences in brain injury and neurodegeneration after mild head trauma.

Repetitive physical insults to the head, including those that elicit mild traumatic brain injury (mTBI), are a known risk factor for a variety of neurodegenerative conditions including Alzheimer's disease (AD), Parkinson's disease (PD), and chronic traumatic encephalopathy (CTE). Although most individuals who sustain mTBI typically achieve a seemingly full recovery within a few weeks, a subset experience delayed-onset symptoms later in life. As most mTBI research has focused on the acute phase of injury, there is an incomplete understanding of mechanisms related to the late-life emergence of neurodegeneration after early exposure to mild head trauma. The recent adoption of Drosophila-based brain injury models provides several unique advantages over existing preclinical animal models, including a tractable framework amenable to high-throughput assays and short relative lifespan conducive to lifelong mechanistic investigation. The use of flies also provides an opportunity to investigate important risk factors associated with neurodegenerative conditions, specifically age and sex. In this review, we survey current literature that examines age and sex as contributing factors to head trauma-mediated neurodegeneration in humans and preclinical models, including mammalian and Drosophila models. We discuss similarities and disparities between human and fly in aging, sex differences, and pathophysiology. Finally, we highlight Drosophila as an effective tool for investigating mechanisms underlying head trauma-induced neurodegeneration and for identifying therapeutic targets for treatment and recovery.

The disease-associated proteins Drosophila Nab2 and Ataxin-2 interact with shared RNAs and coregulate neuronal morphology.

Nab2 encodes the Drosophila melanogaster member of a conserved family of zinc finger polyadenosine RNA-binding proteins (RBPs) linked to multiple steps in post-transcriptional regulation. Mutation of the Nab2 human ortholog ZC3H14 gives rise to an autosomal recessive intellectual disability but understanding of Nab2/ZC3H14 function in metazoan nervous systems is limited, in part because no comprehensive identification of metazoan Nab2/ZC3H14-associated RNA transcripts has yet been conducted. Moreover, many Nab2/ZC3H14 functional protein partnerships remain unidentified. Here, we present evidence that Nab2 genetically interacts with Ataxin-2 (Atx2), which encodes a neuronal translational regulator, and that these factors coordinately regulate neuronal morphology, circadian behavior, and adult viability. We then present the first high-throughput identifications of Nab2- and Atx2-associated RNAs in Drosophila brain neurons using RNA immunoprecipitation-sequencing (RIP-Seq). Critically, the RNA interactomes of each RBP overlap, and Nab2 exhibits high specificity in its RNA associations in neurons in vivo, associating with a small fraction of all polyadenylated RNAs. The identities of shared associated transcripts (e.g., drk, me31B, stai) and of transcripts specific to Nab2 or Atx2 (e.g., Arpc2 and tea) promise insight into neuronal functions of, and genetic interactions between, each RBP. Consistent with prior biochemical studies, Nab2-associated neuronal RNAs are overrepresented for internal A-rich motifs, suggesting these sequences may partially mediate Nab2 target selection. These data support a model where Nab2 functionally opposes Atx2 in neurons, demonstrate Nab2 shares associated neuronal RNAs with Atx2, and reveal Drosophila Nab2 associates with a more specific subset of polyadenylated mRNAs than its polyadenosine affinity alone may suggest.

A protocol to detect neurodegeneration in Drosophila melanogaster whole-brain mounts using advanced microscopy.

Drosophila melanogaster is an excellent model organism to study neurodegeneration. Assessing evident neurodegeneration within the fly brain involves the laborious preparation of thin-sectioned H&E-stained heads to visualize brain vacuole degeneration. Here, we present an advanced microscopy-based protocol, without the need for sectioning, to detect vacuole degeneration within whole fly brains by applying commonly used stains to reveal the brain parenchyma. This approach preserves the whole-brain architecture and enables rapid, reproducible, and quantitative analyses of vacuole-like degeneration associated with specific brain regions. For complete details on the use and execution of this protocol, please refer to Behnke et al. (2021).

Repetitive mild head trauma induces activity mediated lifelong brain deficits in a novel Drosophila model.

Mild head trauma, including concussion, can lead to chronic brain dysfunction and degeneration but the underlying mechanisms remain poorly understood. Here, we developed a novel head impact system to investigate the long-term effects of mild head trauma on brain structure and function, as well as the underlying mechanisms in Drosophila melanogaster. We find that Drosophila subjected to repetitive head impacts develop long-term deficits, including impaired startle-induced climbing, progressive brain degeneration, and shortened lifespan, all of which are substantially exacerbated in female flies. Interestingly, head impacts elicit an elevation in neuronal activity and its acute suppression abrogates the detrimental effects in female flies. Together, our findings validate Drosophila as a suitable model system for investigating the long-term effects of mild head trauma, suggest an increased vulnerability to brain injury in female flies, and indicate that early altered neuronal excitability may be a key mechanism linking mild brain trauma to chronic degeneration.