Alzheimer’s disease (AD) remains a complex and enigmatic neurological disorder characterized by progressive neurodegeneration. While extensive research has shed light on various aspects of AD pathology, the precise events triggering neuronal dysfunction and death have remained elusive. In this comprehensive article, we delve into the intricate molecular mechanisms underlying AD, focusing on recent discoveries related to RNA interference (RNAi) and its potential implications in combating this devastating disease.
At the heart of AD pathology lies the accumulation of plaques composed of amyloid-beta (Aβ) and hyper-phosphorylated tau protein (p-tau), which eventually form neurofibrillary tangles (NFTs). The end-stage of AD is marked by extensive neuronal loss, implicating multiple cell death pathways, including apoptosis, oxidative stress, mitochondria-mediated cell death, necrosis, necroptosis, pyroptosis, ferroptosis, parthanatos, and autophagy. Additionally, it is well-established that Aβ, particularly Aβ42, plays a toxic role, with a strong genetic association between familial AD (FAD) and Aβ42.
Recent research has shed light on the role of DNA damage accelerated by aging in AD, leading to somatic DNA alterations. However, until now, an RNA component had not been identified as a contributor to AD etiology.
RNA interference (RNAi) is a vital post-transcriptional regulation mechanism mediated primarily by microRNAs (miRNAs). MiRNAs, typically 19-25 nucleotides long, exert their influence by negatively regulating gene expression at the mRNA level. These small RNA molecules operate through complementary base pairing, predominantly targeting the 3′ untranslated region (3’UTR) of mRNAs, resulting in cleavage-independent translational silencing. The biogenesis of miRNAs involves a series of processing steps, starting with their transcription in the nucleus as primary miRNA precursors (pri-miRNA), followed by processing into pre-miRNAs and further maturation in the cytoplasm. Dicer and Argonaute (Ago) proteins are crucial players in this process.
A groundbreaking discovery in RNAi research was the identification of RISC-bound short RNAs (R-sRNAs) carrying G-rich 6mer seeds that can trigger cell death by targeting essential survival genes in a RISC- and Ago2-dependent manner, a phenomenon referred to as Death Induced by Survival gene Elimination (DISE). This mechanism has been found to be evolutionarily conserved and operates in humans and rodents. The ratio of miRNAs with toxic versus non-toxic 6mer seeds determines their sensitivity to DISE. While most miRNAs lack G-rich 6mer seeds, certain tumor suppressive miRNAs harness this mechanism to eliminate cancer cells. This led researchers to explore whether DISE might be excessively active in certain diseases, potentially resulting in reduced cancer incidence but excessive tissue loss due to heightened cell death.
Focusing on Alzheimer’s disease, studies have revealed a correlation between DISE and DNA damage, as well as neuronal cell death. Neurons from AD mouse models and induced pluripotent stem cell (iPSC)-derived neurons from AD patients exhibited reduced viability of 6mer seed-carrying R-sRNAs. Furthermore, the aging brain experiences a decline in the expression of non-toxic miRNAs. Interestingly, R-sRNAs from “SuperAgers,” individuals over 80 years old with memory capacities equivalent to those in their 50s to 60s, demonstrated a higher 6mer seed viability compared to control individuals.
Exposing differentiated SH-SY5Y (SH) cells to Aβ42 oligomers was found to shift RISC-bound sRNAs (R-sRNAs) towards more toxic 6mer seeds. Inhibiting RISC function or deleting Ago2 reduced the toxicity of Aβ42 and blocked Aβ42-induced DNA damage. Cells lacking nontoxic sRNAs (e.g., those lacking Drosha expression) exhibited hypersensitivity to Aβ42-induced death, which could be mitigated by reintroducing miRNAs with nontoxic 6mer seeds.
Discussion
The discovery of Death Induced by Survival gene Elimination (DISE) as a powerful cell death mechanism has opened up new avenues for understanding the intricate relationship between RNA interference (RNAi) and neurodegeneration, particularly in Alzheimer’s disease (AD). Our analysis of RISC-bound short RNAs (R-sRNAs) in various AD models provides compelling evidence supporting the hypothesis that DISE may play a crucial role in the survival of neurons.
Notably, while our analysis encompassed all types of sRNAs that can enter the RISC, it’s essential to recognize that in most cells, including neurons, over 95% of the RISC content consists of microRNAs (miRNAs). The role of miRNAs in AD has been extensively investigated, with numerous studies associating specific miRNAs with the disease. However, a critical factor often overlooked is that miRNAs differ significantly in their ability to be incorporated into the RISC, thereby influencing their functional impact.
To shed light on the potential role of DISE in AD and aging, we adopted an Ago-RP-Seq method, focusing on R-sRNAs rather than identifying miRNA targets. Our data strongly suggest a correlation between DISE and the observed neurotoxicity in AD.
In vitro experiments provided valuable insights into the involvement of the RISC and RNAi in the toxicity of Aβ42. Furthermore, we established a significant association between the level of neurotoxicity and the 6mer seed viability of R-sRNAs in various AD models. Lower 6mer seed viability was linked to increased cell death, while a higher overall 6mer seed viability in the RISC content appeared to protect neurons from demise. Interestingly, this viability was reduced in older mouse brains and in vitro aged iPSC-derived neurons, implying a loss of protective miRNAs with age. It is important to note that further confirmation of these findings will be necessary with larger participant cohorts.
Extensive literature supports the neuroprotective role of miRNAs in various disease models, emphasizing their significance in preserving neuronal function. For example, studies in Drosophila have shown that impairing miRNA processing exacerbates neurodegeneration induced by certain genetic mutations. In parallel, research has demonstrated how toxic sRNAs derived from CAG repeats can lead to cell death, underscoring the importance of miRNAs in neuronal survival. Additionally, experiments involving Dicer knockout mice have highlighted the protective role of miRNAs in adult dopamine neurons, and treatment with Enoxacin has shown promise in reducing disease severity in Amyotrophic Lateral Sclerosis (ALS) mice.
Based on our findings, we propose a model in which the RISC functions as a central regulator, influencing neuronal cell death and DNA damage observed during aging and in AD, which may have broader implications for other neurodegenerative diseases:
- Youthful Neurons: In young, healthy individuals or those without AD symptoms, the RISC is populated with abundant miRNAs, most of which carry non-toxic 6mer seeds. These miRNAs serve a dual purpose: regulating gene expression and acting as protective shields against toxic endogenous sRNAs with G and C-rich regions. This protective mechanism prevents the entry of harmful sRNAs into the RISC, thereby maintaining neuronal health.
- Aging and AD: As individuals age or develop AD, Aβ oligomers and Tau aggregation trigger a cellular response leading to the upregulation of endogenous toxic sRNAs, including miRNAs and other sRNAs such as tRNA and rRNA fragments. This response may occur directly due to AβOs or Tau aggregation, or indirectly through cumulative processes. Our data suggest that DISE contributes moderately but significantly to cell death induced by toxic Aβ42. Furthermore, Tau appears to be essential for Aβ42 to engage the RISC and mediate toxicity. Importantly, the DNA damage caused by Aβ42 treatment relies on RISC engagement, indicating its pivotal role in the observed DNA damage.
- Age-Related Shift: With advancing age, there is a shift in the balance of R-sRNAs with toxic versus non-toxic 6mer seeds towards lower 6mer seed viability. This renders neurons susceptible to DISE-inducing sRNAs. The age-dependent loss of non-toxic miRNAs may result from reduced miRNA production capacity in aging neurons. Factors such as increased oxidative stress, interferons, and the decline in Dicer stability could contribute to this phenomenon. Recent research has also shown that Drosha, a critical player in miRNA biogenesis, undergoes translocation from the nucleus to the cytosol under AD-related stress conditions, further supporting our observations.
Our data suggest that RISC activity plays a dual role in cellular responses to Aβ42: cell death through DISE and DNA damage. DISE, a combination of multiple cell death pathways, is dependent on the transcriptome of the affected cell, aligning with the varied cell death pathways implicated in AD. While cell death is considered a late event in AD pathogenesis, neuronal dysfunction can lead to cell death, particularly in post-mitotic neurons, eventually resulting in substantial brain cell loss in AD patients.
Moreover, our findings raise the possibility that the increase in toxic sRNAs coupled with the gradual loss of non-toxic miRNAs during aging may also be central to other degenerative brain diseases such as Parkinson’s disease (PD), Huntington’s disease (HD), and Amyotrophic Lateral Sclerosis (ALS). The presence of hexameric repeat sequences, similar to those found in the C9orf72 gene associated with ALS, can generate highly toxic sRNAs, adding to the potential relevance of DISE in various neurodegenerative diseases. The delayed onset of these diseases in adulthood, despite decades of symptom-free life, may be attributed to the gradual loss of protective miRNAs.
In conclusion, while extensive research in Alzheimer’s disease has predominantly focused on reducing amyloid plaque load and preventing tau phosphorylation, our data introduce a new perspective. We propose that the high expression of non-toxic miRNAs may serve as a protective mechanism against neurodegeneration. Increasing miRNA biogenesis or blocking toxic R-sRNAs may hold promise as a novel treatment strategy for various neurodegenerative diseases, including AD.
reference link : https://www.nature.com/articles/s41467-023-44465-8
