SPOCK3, a risk gene for adult ADHD and personality disorders.

Attention-deficit/hyperactivity disorder (ADHD) is the most frequent psychiatric disorder in children, where it displays a global prevalence of 5 %. In up to 50 % of the cases, ADHD may persist into adulthood (aADHD), where it is often comorbid with personality disorders. Due to a potentially heritable nature of this comorbidity, we hypothesized that their genetic framework may contain common risk-modifying genes. SPOCK3, a poorly characterized, putatively Ca(2+)-binding extracellular heparan/chondroitin sulfate proteoglycan gene encoded by the human chromosomal region 4q32.3, was found to be associated with polymorphisms among the top ranks in a genome-wide association study (GWAS) on ADHD and a pooled GWAS on personality disorder (PD). We therefore genotyped 48 single nucleotide polymorphisms (SNPs) representative of the SPOCK3 gene region in 1,790 individuals (n aADHD = 624, n PD = 630, n controls = 536). In this analysis, we found two SNPs to be nominally associated with aADHD (rs7689440, rs897511) and four PD-associated SNPs (rs7689440, rs897511, rs17052671 and rs1485318); the latter even reached marginal significance after rigorous Bonferroni correction. Bioinformatics tools predicted a possible influence of rs1485318 on transcription factor binding, whereas the other candidate SNPs may have effects on alternative splicing. Our results suggest that SPOCK3 may modify the genetic risk for ADHD and PD; further studies are, however, needed to identify the underlying mechanisms.

Quantification of nuclear protein transport using induced heterodimerization.

Trafficking of proteins between the cytoplasm and nucleus occurs exclusively across the nuclear pore complex of eucaryotic cells. Fundamental aspects of this process affect temporal and spatial parameters, the latter carried out by specific import [nuclear localization sequence (NLS)] and export [nuclear export sequence (NES)] sequences. In this study, we focused on the adaptation of a protein heterodimerization assay to kinetically measure Crm1-mediated nuclear export in living cells using the rapalog AP21967, a heterodimerizing agent and NLS- and NES-containing fusion proteins equipped with distinct AP21967-specific binding motifs. In HeLa cells, we observed rapid nuclear export of the NLS-containing fusion protein in the presence of AP21967, with the extent of this process being a function of the number of AP21967-binding motifs. AP21967-induced nuclear export was specifically inhibited by the Crm1-binding molecule leptomycin B. Half maximal export was achieved after approximately 10 min. We further applied protein heterodimerization in HeLa cells to study induced NLS-mediated nuclear import. Only in the presence of heterodimerizer AP21967 nuclear import of a cytoplasmically localizing fusion protein was observed. Induced protein heterodimerization is thus a valuable tool to quantitatively study nucleocytoplasmic protein trafficking in cultured cells, in a non-invasive, time-saving manner.

Nuclear protein import is reduced in cells expressing nuclear envelopathy-causing lamin A mutants.

Lamins, which form the nuclear lamina, not only constitute an important determinant of nuclear architecture, but additionally play essential roles in many nuclear functions. Mutations in A-type lamins cause a wide range of human genetic disorders (laminopathies). The importance of lamin A (LaA) in the spatial arrangement of nuclear pore complexes (NPCs) prompted us to study the role of LaA mutants in nuclear protein transport. Two mutants, causing prenatal skin disease restrictive dermopathy (RD) and the premature aging disease Hutchinson Gilford progeria syndrome, were used for expression in HeLa cells to investigate their impact on the subcellular localization of NPC-associated proteins and nuclear protein import. Furthermore, dynamics of the LaA mutants within the nuclear lamina were studied. We observed affected localization of NPC-associated proteins, diminished lamina dynamics for both LaA mutants and reduced nuclear import of representative cargo molecules. Intriguingly, both LaA mutants displayed similar effects on nuclear morphology and functions, despite their differences in disease severity. Reduced nuclear protein import was also seen in RD fibroblasts and impaired lamina dynamics for the nucleoporin Nup153. Our data thus represent the first study of a direct link between LaA mutant expression and reduced nuclear protein import.

Further evidence for plasma progranulin as a biomarker in bipolar disorder.

BACKGROUND: A recent study suggested that progranulin (encoded by the fronto-temporal dementia risk gene GRN) plasma levels are decreased in bipolar disorder (BD). Replication of this finding is however lacking. METHODS: Progranulin plasma levels of bipolar patients (n=104) and healthy controls (n=80) were measured by enzyme-linked immunosorbent assay (ELISA). Participants were also genotyped for three single nucleotide polymorphisms (SNPs) in the GRN gene (rs2879096, rs4792938 and rs5848), and the effect of genetic variation on progranulin levels was examined. RESULTS: Plasma progranulin levels were decreased in BD (ANCOVA, p=0.001). Furthermore, age was significantly and positively correlated with plasma progranulin (Pearson׳s correlation, r=0.269, p<0.001). Also, lithium treatment but no other medication had a significant effect on progranulin plasma levels (ANCOVA, p=0.007). Specifically in BD, the GRN SNP rs5848 was associated with progranulin plasma levels (Kruskal-Wallis test, p<0.005). LIMITATIONS: Subgroup analysis regarding bipolar I vs. bipolar II subtype and polarity of the episode at sampling (manic vs. depressed vs. mixed vs. rapid cycling vs. euthymic) could only be performed with limited validity due to the relatively small sample size. The suitability of peripheral progranulin as a biomarker for BD is limited due to the overlap between patients and controls. CONCLUSION: The findings strengthen the evidence for progranulin being involved in pathomechanisms of bipolar disorder, and suggest a genetic determinant of progranulin concentrations that is relevant specifically in bipolar patients.