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NEN_68_6_Gentil_200506_SDC3

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NEN_68_6_Gentil_200506_SDC3

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  1. Supplementary Figure 2. Unlike motor neurons, aggregation of mutant NFL in SW13vim- cells co-expressing NFH to enable NF assembly is not consistently prevented by HSPB1 and HSPA8. Only HSPB1 prevented mutant NFL aggregation and only of the P8R mutant. SW13vim cells were transiently transfected with pRc-CMV-NFM and one of pCDNA4-NFL (A, B, C), pCDNA4-NFLQ333P (D–F) orpCDNA4-NFLP8R (G–I), plus either pCDNA-HSPB1 (B, E, H) or pRc-CMV-HSPA1 (C, F, I). Cultures were fixed 24 hours following transfection and double-labelled with anti-myc (to visualize plasmid-derived NFL) and either goat anti-HSPB1 or mouse anti-HSPA1. Micrographs are representative of quantitative data presented in Supplementary Table 2 and summarized in Table 3. Scale bar: 10 mm. Note that endogenous HSPB1 aggregates with Q333P mutant and that HSPA1 aggregates with both NFL mutants (F, I). Scale bar: 10 mm. HSP = heat shock protein; NFL = neurofilament light chain; NFM = medium molecular weight neurofilament subunit, SW13vim- = Human adrenal carcinoma cell line that lackscytoplasmic intermediate filaments. Expression vectors are as described in (31).

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