Antonio Velayos-Baeza

LRRTM1 on chromosome 2p12 is a maternally suppressed gene that is associated paternally with handedness and schizophrenia

Left–right asymmetrical brain function underlies much of human cognition, behavior and emotion. Abnormalities of cerebral asymmetry are associated with schizophrenia and other neuropsychiatric disorders. The molecular, developmental and evolutionary origins of human brain asymmetry are unknown. We found significant association of a haplotype upstream of the gene LRRTM1 (Leucine-rich repeat transmembrane neuronal 1) with a quantitative measure of human handedness in a set of dyslexic siblings, when the haplotype was inherited paternally (P=0.00002). While we were unable to find this effect in an epidemiological set of twin-based sibships, we did find that the same haplotype is overtransmitted paternally to individuals with schizophrenia/schizoaffective disorder in a study of 1002 affected families (P=0.0014). We then found direct confirmatory evidence that LRRTM1 is an imprinted gene in humans that shows a variable pattern of maternal downregulation. We also showed that LRRTM1 is expressed during the development of specific forebrain structures, and thus could influence neuronal differentiation and connectivity. This is the first potential genetic influence on human handedness to be identified, and the first putative genetic effect on variability in human brain asymmetry. LRRTM1 is a candidate gene for involvement in several common neurodevelopmental disorders, and may have played a role in human cognitive and behavioral evolution.

Chorein Detection for the Diagnosis of Chorea-Acanthocytosis

Chorea‐acanthocytosis (ChAc) is a severe, neurodegenerative disorder that shares clinical features with Huntingtons disease and McLeod syndrome. It is caused by mutations in VPS13A, which encodes a large protein called chorein. Using antichorein antisera, we found expression of chorein in all human cells analyzed. However, chorein expression was absent or noticeably reduced in ChAc patient cells, but not McLeod syndrome and Huntingtons disease cells. This suggests that loss of chorein expression is a diagnostic feature of ChAc. Ann Neurol 2004;56:299–302

Alternative splicing in the dyslexia-associated gene KIAA0319

The KIAA0319 gene in chromosome 6p22 has been strongly associated with developmental dyslexia. In this article we show a wide expression pattern of this gene in human adult brain by Northern blot analysis. We also performed RT-PCR analysis to detect alternative splicing variants in human brain. Most of the detected variants involve alternative splicing of the exons at the 5′ and the 3′ ends. Two main forms differing in the length of the 5′ UTR are detected at approximately the same rate. Two variants (B and C) lacking exon 19, which encodes the transmembrane domain, are the main alternative forms detected among those predicted to encode protein. These two variants could be secreted and might be involved in signaling functions. A similar RT-PCR analysis performed in mouse and rat adult brains showed that only some of the alternative splicing variants are equivalent to those found in the human gene.

The dyslexia-associated KIAA0319 protein undergoes proteolytic processing with {gamma}-secretase-independent intramembrane cleavage.

The KIAA0319 gene has been associated with reading disability in several studies. It encodes a plasma membrane protein with a large, highly glycosylated, extracellular domain. This protein is proposed to function in adhesion and attachment and thought to play an important role during neuronal migration in the developing brain. We have previously proposed that endocytosis of this protein could constitute an important mechanism to regulate its function. Here we show that KIAA0319 undergoes ectodomain shedding and intramembrane cleavage. At least five different cleavage events occur, four in the extracellular domain and one within the transmembrane domain. The ectodomain shedding processing cleaves the extracellular domain, generating several small fragments, including the N-terminal region with the Cys-rich MANEC domain. It is possible that these fragments are released to the extracellular medium and trigger cellular responses. The intramembrane cleavage releases the intracellular domain from its membrane attachment. Our results suggest that this cleavage event is not carried out by γ-secretase, the enzyme complex involved in similar processing in many other type I proteins. The soluble cytoplasmic domain of KIAA0319 is able to translocate to the nucleus, accumulating in nucleoli after overexpression. This fragment has an unknown role, although it could be involved in regulation of gene expression. The absence of DNA-interacting motifs indicates that such a function would most probably be mediated through interaction with other proteins, not by direct DNA binding. These results suggest that KIAA0319 not only has a direct role in neuronal migration but may also have additional signaling functions.

Trafficking of the Menkes copper transporter ATP7A is regulated by clathrin-, AP-2–, AP-1–, and Rab22-dependent steps

ATP7A mediates copper absorption and feeds cuproenzymes in the trans-Golgi network. To regulate copper homeostasis, ATP7A cycles between the TGN and plasma membrane. The roles of clathrin, adaptor complexes, lipid rafts, and Rab22a are assessed in an attempt to decipher the regulatory proteins involved in ATP7A cycling.

The dyslexia-associated protein KIAA0319 interacts with adaptor protein 2 and follows the classical clathrin-mediated endocytosis pathway

Recently, genetic studies have implicated KIAA0319 in developmental dyslexia, the most common of the childhood learning disorders. The first functional data indicated that the KIAA0319 protein is expressed on the plasma membrane and may be involved in neuronal migration. Further analysis of the subcellular distribution of the overexpressed protein in mammalian cells indicates that KIAA0319 can colocalize with the early endosomal marker early endosome antigen 1 (EEA1) in large intracellular vesicles, suggesting that it is endocytosed. Antibody internalization assays with full-length KIAA0319 and deletion constructs confirmed that KIAA0319 is internalized and showed the importance of the cytoplasmic juxtamembranal region in this process. The present study has identified the medium subunit (μ2) of adaptor protein 2 (AP-2) as a binding partner of KIAA0319 in a yeast two-hybrid screen. Using Rab5 mutants or depletion of the μ-subunit of AP-2 or clathrin heavy chain by RNA interference, we demonstrate that KIAA0319 follows a clathrin-mediated endocytic pathway. We also identify tyrosine-995 of KIAA0319 as a critical amino acid required for the interaction with AP-2 and subsequent internalization. These results suggest the surface expression of KIAA0319 is regulated by endocytosis, supporting the idea that the internalization and recycling of the protein may be involved in fine tuning its role in neuronal migration.

Identification of a VPS13A founder mutation in French Canadian families with chorea-acanthocytosis

Mutations in VPS13A cause chorea-acanthocytosis (ChAc), an autosomal recessive neurodegenerative disorder. VPS13A is located in a tail-to-tail arrangement with GNA14 on chromosome 9q21. ChAc shows substantial allelic heterogeneity, with no single VPS13A mutation causing the majority of cases. We examined 11 patients in four French Canadian ChAc pedigrees for mutations in VPS13A. Affected members of three families were homozygous for a 37-kb deletion of the four terminal exons of VPS13A (EX70_EX73del). This deletion also encompasses the two terminal exons of GNA14. Two affected females in family 4 were homozygous for the splicing mutation 4242+1G>T. Remarkably, the affected males in this highly consanguineous pedigree were compound heterozygotes for EX70_EX73del and 4242+1G>T. PCR analysis of the deletion breakpoint junction revealed that an additional patient with French Canadian ancestry was heterozygous for the EX70_EX73del allele. The identification of a common 9q21 haplotype associated with EX70_EX73del in at least four apparently unrelated ChAc families implies that ChAc shows a founder effect in French Canadians, and that routine testing for EX70_EX73del in suspected ChAc cases may therefore be worthwhile in this population. The deletion breakpoint PCR described here will enable rapid identification of both homozygous and heterozygous carriers of EX70_EX73del.

Identification of VPS13C as a Galectin-12-Binding Protein That Regulates Galectin-12 Protein Stability and Adipogenesis.

Galectin-12, a member of the galectin family of β-galactoside-binding animal lectins, is preferentially expressed in adipocytes and required for adipocyte differentiation in vitro. This protein was recently found to regulate lipolysis, whole body adiposity, and glucose homeostasis in vivo. Here we identify VPS13C, a member of the VPS13 family of vacuolar protein sorting-associated proteins highly conserved throughout eukaryotic evolution, as a major galectin-12-binding protein. VPS13C is upregulated during adipocyte differentiation, and is required for galectin-12 protein stability. Knockdown of Vps13c markedly reduces the steady-state levels of galectin-12 by promoting its degradation through primarily the lysosomal pathway, and impairs adipocyte differentiation. Our studies also suggest that VPS13C may have a broader role in protein quality control. The regulation of galectin-12 stability by VPS13C could potentially be exploited for therapeutic intervention of obesity and related metabolic diseases.

The Function of Chorein

Chorein is the protein encoded by gene VPS13A which is altered in chorea-acanthocytosis (ChAc). It belongs to the VPS13 protein family which, in mammals, has three other members: VPS13B, VPS13C and VPS13D. These pro- teins are similar to Vps13p, a yeast protein shown to be involved in intra-cellular trafficking of a number of transmembrane proteins. Chorein and its homologous human proteins lack domains or motifs of known function. This, together with their large size, makes the functional characterisation of these proteins a difficult task. Nevertheless, we have undertaken this task following a molecular and cellular biology approach. We have cloned the cDNA for the human VPS13 genes and used them for transfection of mammalian cell lines. We present here an overview of the

Dominant transmission of chorea-acanthocytosis with VPS13A mutations remains speculative

Ishida et al. [1] report the neuropathological study of a patient with chorea-acanthocytosis (ChAc) from the family published earlier by Saiki et al. [2–4]. Using detailed neuropathological and immunohistochemical studies, Ishida and colleagues found signiWcant diVerences as well as similarities between ChAc and Huntington’s disease. While comparing their observations with those in ChAc patients reported elsewhere, the authors comment on the similarities between their case, with supposedly autosomal dominant transmission, and the typical autosomal recessive cases. However, Ishida et al.’s point may not be valid as it is unclear whether their cases truly show autosomal dominant inheritance. In the two aVected siblings, only a single heterozygous G > A transversion at the last nucleotide of exon 57 on one allele of VPS13A was found [2]. Their unaVected mother, who was not consanguineous with the father of the siblings, did not possess this mutation. No other family members were studied by molecular genetic methods. Father and paternal grandfather were both deceased and were reported as being “presumptively aVected”, while two paternal aunts and a son of one of these were reported to have also had hyperkinetic movements, hyporeXexia, and acanthocytosis. Thus, it was thought that the mutation had been introduced from the paternal side, and was transmitted in an autosomal dominant manner. Although the symptoms described for other family members are consistent with ChAc, other conditions were not excluded, and Wndings more strongly suggestive of ChAc are not mentioned, e.g., feeding-induced tongue dystonia, lipor tongue-biting or other self-mutilation, seizures, psychiatric abnormalities, or cognitive impairment. Ishida et al. postulate autosomal dominant transmission of a condition which is typically autosomal recessive. However, the possibility of autosomal recessive inheritance should not be excluded in their family. Mutations such as heterozygous whole exon deletions or rearrangements cannot be detected using the usual mutation screening methods [5]. Thus, it is possible that the second mutation of the index siblings was missed due to technical reasons, as was reported in other cases [6]. The lack of a complete genetic analysis of the VPS13A gene in the proband’s father, grandfather and other family members as well as the high ChAc incidence in Japan [7] weaken the claim of an autosomal dominant trait. It is possible that a mutated allele transmitted by the mother was missed in both her and her two children, and that the mutation discovered in the siblings came from their father, whose clinical status does not permit a diagnosis. ChAc cannot clearly be excluded on the paternal side, yet such features could be explained by complex patterns of intermarriage such as seen in French-Canadian sibships in whom an initial claim of dominant inheritance was dropped after further analyses [6, 8]. A major additional issue is the absence of information about the presence of chorein, the VPS13A product. Detection of this protein by Western blot (WB) has been proven B. Bader (&) · A. Danek Neurologische Klinik und Poliklinik, Ludwig-Maximilians-Universität München, Munich, Germany e-mail: benedikt.bader@med.uni-muenchen.de