Activation of polo-like kinase 1 correlates with selective motor neuron vulnerability in familial ALS

Mutations in the Fused in Sarcoma (FUS) gene cause familial amyotrophic lateral sclerosis (ALS), characterized by selective degeneration of spinal motor neurons (sMNs) with relative sparing of cortical neurons (CNs). The mechanisms underlying this cell-type vulnerability remain unclear. Here, we compare CNs and sMNs derived from FUS-ALS models to assess differential responses to FUS mutations. We find that CNs are less affected than sMNs in DNA damage repair, axonal organelle trafficking, and stress granule dynamics. RNA sequencing (RNA-seq) reveals distinct transcriptomic signatures, with sMNs uniquely activating DNA damage responses involving cell cycle regulators, particularly polo-like kinase 1 (PLK1). PLK1 is highly expressed in sMNs but not CNs, correlating with greater nuclear FUS loss and splicing defects in sMNs. Cross-comparison with other familial ALS RNA-seq datasets highlights PLK1 upregulation as a shared molecular feature. These findings identify intrinsic differences between CNs and sMNs in FUS-ALS and suggest PLK1 as a potential driver of sMN vulnerability. Patient-derived isogenic spinal motoneurons (MN) or cortical neurons (CN) expressing either normal (FUS WT) or mutant P525L FUS-eGFP (FUS P525L) were matured for 21 DIV. The main cell model used in this study were neurons derived from FUS-WT-EGFP and FUS-P525L-EGFP that were generated as a part of previous study (Naumann, Pal et al. 2018). Briefly, fibroblasts carrying R521C FUS mutation obtained from a 58-year-old female were reprogrammed into iPSC using cDNA of OCT4, SOX2, KLF4 and cMYC delivered via retroviral vectors. Next, CRISPR/Cas9 genome editing was used to i) correct the R521C mutation into WT FUS and to tag it with EGFP to generate the FUS WT EGFP line or ii) replace the R521C mutation into P525L FUS mutation and to tag it with EGFP to generate FUS P525L EGFP line. Differentiation of hiPSC into spinal motor neurons (MNs) was carried out based on the protocol from Reinhardt et al. 2013. Differentiation of hiPSC into cortical neurons (CN) was based on the protocol published by Burkhardt et al.(Burkhardt, Martinez et al. 2013, Japtok, Lojewski et al. 2015).