Endocrine Abstracts

Mutations in the Seipin/BSCL2 gene cause either type 2 congenital generalized lipodystrophy (BSCL) or dominant motor neuron diseases. However, we recently discovered a c.985C>T mutation in the BSCL2 gene that results in a novel fatal neurodegenerative syndrome (celia encephalopathy). This mutation induces an alternative splicing which results in skipping of exon 7 and a reading frame shift (Guillen-Navarro et al. 2013 J Med Genet 50 401–409).

Homozygous patients suffer from progressive encephalopathy since ages 2–3 years, with fatal outcome at ages 6–8 years; however, carriers for the c.985C>T mutation are asymptomatic, conflicting with the gain of toxic function attributed to the mutation. Our previous studies showed a partial nuclear localization of wt and exon 7 skipped seipin. This location might be related to the presence of intranuclear ubiquitin-positive inclusions in brain tissue from a homozygous patient (index case), which are likely to consist of misfolded, aggregated mutant seipin. Besides, ER stress in cells expressing exon 7 skipped seipin was also found. Here we report further molecular characterization of exon 7 skipped seipin. Using density gradient ultracentrifugation, we found that wt seipin oligomerizes forming tetramers according to calibration with a set of proteins of known MW. We also found that exon 7 skipped seipin forms much larger aggregates under similar conditions, corresponding to dodecamers. We hypothesized that wt and exon 7 skipped seipin might interact and form mixedoligomers. Given that levels of wt seipin expressed by heterozigous individuals are substantially higher than those of exon 7 skipped seipin, we reasoned that wt seipin might rescue the phenotype by recruiting exon 7 skipped seipin into mixed normal size oligomers, thus impeding their aggregation into larger, presumably toxic, and oligomers. We therefore expressed wt and exon 7 skipped seipin at a 3:1 ratio in HeLa cells and performed density gradient ultracentrifugation of cell extracts. We found a change in the pattern of distribution of each of the seipin isoforms, with exon 7 skipped seipin emerging at earlier fractions. This result clearly indicates the interaction between both seipin isoforms through the formation of mixed oligomers.

Together, our findings provide a clue about the origin of the intranuclear aggregates observed in neurons from the index case, and offer a possible explanation of the absence of phenotype in heterozygous mutation carriers (funded by Consellería de Industria (Xunta de Galicia), 10PXIB208013PR and ISCIII-FEDER PI10/02873).