Biology subjects

Santangelo, S.

Publications and source records attributed to Santangelo, S..

3 recordsLinked to original sources

bioRxiv2024

NEK1 haploinsufficiency worsens DNA damage but not defective ciliogenesis in C9ORF72 patient-derived iPSC-motoneurons

A hexanucleotide G4C2 repeat expansion (HRE) in C9ORF72 gene is the major cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), leading to both loss- and gain-of-function pathomechanisms. The wide clinical heterogeneity among C9ORF72 patients suggests potential modifying genetic factors. Notably, C9ORF72 mutations often co-occur with other variants in ALS/FTD-associated genes, such as NEK1, which encodes for a kinase involved in multiple pathways including DNA damage response and ciliogenesis. In this study, we generated induced pluripotent stem cells (iPSCs) and differentiated motoneurons (iPSC-MNs) derived from an ALS patient carrying both C9ORF72 HRE and a NEK1 loss-of-function mutation to study the effect of the NEK1 haploinsufficiency on C9ORF72 pathology. Double mutant C9ORF72/NEK1 cells showed increased pathological C9ORF72 RNA foci in iPSCs and higher DNA damage levels in iPSC-MNs compared to single mutant C9ORF72 cells. In contrast, ciliogenesis was similarly impaired in both C9ORF72 and C9ORF72/NEK1 iPSC-MNs showing shorter cilia. Altogether, our study supports the use of patient-derived iPSCs to functionally explore the contribution of genetic modifiers in C9ORF72-associated pathology.

neuroscience↗

bioRxiv2024

NEK1 haploinsufficiency impairs ciliogenesis in human iPSC-derived motoneurons and brain organoids

Primary cilia are microtubule-based organelles acting as specialized signalling antennae that respond to specific stimuli to maintain cellular integrity and homeostasis. Recent studies indicate defective primary cilia in post-mortem human brains and animal models of neurodegenerative conditions, including Amyotrophic Lateral Sclerosis (ALS). Heterozygous loss-of-function mutations (LOF) in NEK1 gene are present in about 1% of familial and sporadic ALS cases. The protein kinase NEK1 regulates various cellular processes, including ciliogenesis, but a clear link between NEK1 LOF mutation in ALS and primary cilia is unknown. In this study we generated a human iPSC line carrying a NEK1 LOF mutation by gene editing, leading to NEK1 protein haploinsufficiency. In differentiated iPSC-motoneurons (MNs) we observed that primary cilia were significantly shorter in NEK1-LOF iPSC-MNs compared to wild-type (WT) iPSC-MNs and that also the percentage of ciliated iPSC-MNs was significantly decreased in NEK1-LOF cells. We also investigated ciliogenesis in NEK1-LOF iPSC-brain organoids confirming that primary cilia were thinner with no apparent alteration in the ultrastructure by transmission electron microscopy. Our data suggest that NEK1 protein plays a role in regulating ciliogenesis in both 2D and 3D human iPSC-derived neuronal models and that NEK1 LOF mutations associated to ALS, leading to NEK1 haploinsufficiency and likely to reduced kinase activity, impair primary cilium formation. The involvement of ciliogenesis dysfunction in ALS deserves further investigation providing novel therapeutic targets and strategies to be addressed for this incurable disease.

neuroscience↗

bioRxiv2024

Altered nuclear envelope homeostasis is a key pathogenic event in C9ORF72-linked ALS/FTD

ALS and FTD are complex neurodegenerative disorders that primarily affects motor neurons in the brain and spinal cord, and cortical neurons in the frontal lobe. Although the pathogenesis of ALS/FTD is unclear, recent research spotlights nucleocytoplasmic transport impairment, DNA damage, and nuclear abnormalities as drivers of neuronal death. In this study, we show that loss of nuclear envelope (NE) integrity is a key pathology associated with nuclear pore complex (NPC) injury in C9ORF72 mutant neurons. Importantly, we show that mechanical stresses generated by cytoskeletal forces on the NE can lead to NPC injury, loss of nuclear integrity, and accumulation of DNA damage. Importantly, we demonstrate that restoring NE tensional homeostasis, by disconnecting the nucleus from the cytoskeleton, can rescue NPC injury and reduce DNA damage in C9ORF72 mutant cells. Together, our data suggest that modulation of NE homeostasis and repair may represent a novel and promising therapeutic target for ALS/FTD.

neuroscience↗