The nuclear import receptor Kapβ2 modifies neurotoxicity mediated by poly(GR) in C9orf72-linked ALS/FTD

Expanded intronic G4C2 repeats in the C9ORF72 gene cause amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). These intronic repeats are translated through a non-AUG-dependent mechanism into five different dipeptide repeat proteins (DPRs), including poly-glycine-arginine (GR), which is aggregation-prone and neurotoxic. Here, we report that Kapβ2 and GR interact, co-aggregating, in cultured neurons in vitro and CNS tissue in vivo. Importantly, this interaction significantly decreased the risk of death of cultured GR-expressing neurons. Downregulation of Kapβ2 is detrimental to their survival, whereas increased Kapβ2 levels mitigated GR-mediated neurotoxicity. As expected, GR-expressing neurons displayed TDP-43 nuclear loss. Raising Kapβ2 levels did not restore TDP-43 into the nucleus, nor did alter the dynamic properties of GR aggregates. Overall, our findings support the design of therapeutic strategies aimed at up-regulating Kapβ2 expression levels as a potential new ..., Human tissues: Human post-mortem CNS tissue from healthy controls and patients carrying the C9ORF72 intronic nucleotide repeat expansion was obtained from the Target ALS and the Jefferson Weinberg ALS center biobanks. Information and demographics of patients and controls are presented in Table 4. Human post-mortem tissues (fresh frozen) were stored at -80 C.
Cell cultures: HEK293 cells were cultured in DMEM medium (Cytiva Cat.# SH30243.LS ) supplemented with 10% FBS (Cytiva Cat.# SH30071.01HI), penicillin, and streptomycin (Thermo Fisher Scientific Cat.# SV30010). Cells were passaged every 3-4 days using 0.05% trypsin (Corning Cat.# 25-051-C).
Primary cortical neurons: After meninges removal, brains were dissected from embryonic day 16 (E16) rat embryos. Cortices and midbrain regions were cut into small pieces and incubated on a shaker at 80 rcf for 45 min at 37°C in 0.2% trypsin in HBSS without Ca2+ and Mg2+ (Cytiva Cat.# SH30588.01). FBS (Cytiva Cat.# SH30071.01HI) was added, and the ..., , # The nuclear import receptor Kapβ2 modifies neurotoxicity mediated by poly(GR) in C9orf72-linked ALS/FTD

[https://doi.org/10.5061/dryad.v41ns1s4f](https://doi.org/10.5061/dryad.v41ns1s4f)

These datasets contain all the source files for the graphs in the associated paper.

## Description of the data and file structure

Source data for each graph labeled as panel yx (y=number of figure, x=letter of the panel)

### Figure 1. GR does not affect Kapβ2 levels in neurons.

* a) qPCR analysis of Kapβ2 levels in cortical neurons transduced with GFP or GR50. Data are represented as mean ± S.E.M. (n=6 biological replicates, Student t-test, n.s. = not significant)
* c) The graph bar shows the quantification of the WB of cortical neuron extracts for Kapβ2 protein. Total protein staining was used to normalize. Data are represented as mean ± S.E.M. (n=3 biological replicates, Student t-test, n.s. = not significant).
* d) qPCR analysis of Kapβ2 transcript levels in the cortex and spinal cord o...