Septo-optic dysplasia caused by a novel FLNA splice site mutation: a case report.

BACKGROUND: Septo-optic dysplasia (SOD), also known as de-Morsier syndrome, is a rare disorder characterized by any combination of optic nerve hypoplasia, pituitary gland hypoplasia, and midline abnormalities of the brain including absence of the septum pellucidum and corpus callosum dysgenesis. The variable presentation of SOD includes visual, neurologic, and/or hypothalamic-pituitary endocrine defects. The unclear aetiology of a large proportion of SOD cases underscores the importance of identifying novel SOD-associated genes. CASE PRESENTATION: To identify the disease-causing gene in a male infant with neonatal hypoglycaemia, dysmorphic features, and hypoplasia of the optic nerve and corpus callosum, we designed a targeted next-generation sequencing panel for brain morphogenesis defects. We identified a novel hemizygous deletion, c.6355 + 4_6355 + 5delAG, in intron 38 of the FLNA gene that the patient had inherited from his mother. cDNA studies showed that this variant results in the production of 3 aberrant FLNA transcripts, the most abundant of which results in retention of intron 38 of FLNA. CONCLUSIONS: We report for the first time a case of early-onset SOD associated with a mutation in the FLNA gene. This finding broadens the spectrum of genetic causes of this rare disorder and expands the phenotypic spectrum of the FLNA gene.

Severe course of community-acquired pneumonia in an adult patient who is heterozygous for Q481P in the perforin gene: are carriers of the mutation free of risk?

Most cases of autosomal recessive hemophagocytic lymphohistiocytosis (HLH) are associated with over 50 mutations in the perforin gene. Some of these mutations have no clear functional association. Only homozygous patients display a full-blown syndrome, whereas no severe disease has been described in heterozygous carriers of these mutations despite the presence of functional and phenotypic alterations in cytotoxic cells. We study the family of a child who died from HLH at 6 months of age due to a Q481P mutation in the perforin gene. The study is particularly interesting because the patient's heterozygous father experienced severe community-acquired pneumonia that could be attributed to deficient in vitro NK cell activity despite normal perforin expression. This case report suggests that impaired NK cell activity in a heterozygote can result in poorer initial control of infections with severe clinical expression.

A novel mutation in the E-cadherin gene in the first family with hereditary diffuse gastric cancer reported in Spain.

AIMS: Mutations of the E-cadherin gene (CDH1) result in dominantly inherited hereditary diffuse gastric cancer (HDGC). We report a study in the first family diagnosed with HDGC in Spain, examining the presence of mutations in the CDH1 gene. METHODS: The presence of mutations was studied by direct sequencing of all CDH1 exons. Immunohistochemical analysis with specific antibodies was used to detect the expression of E-cadherin in normal and tumour tissue. RESULTS: A novel 1610delC mutation in exon 11 has been found in a Spanish family diagnosed with HDGC. This mutation generates a premature stop codon at position 1667 giving rise to a truncated protein that lacks the transmembrane and beta-catenin-binding domains. The presence of a 1610delC germline mutation was confirmed in three family members diagnosed with diffuse gastric cancer, and also in six asymptomatic members. Of note, the diffuse gastric cancer coexisted with a gastric lymphoma in the proband. Furthermore, immunohistochemical analyses of tumour tissue showed the complete absence of E-cadherin in the proband, revealing a second genetic hit at the CDH1 locus. CONCLUSIONS: We have identified a HDGC family in Spain that carries a novel germline truncating mutation in the CDH1 gene.

Case-control study and meta-analysis of low density lipoprotein receptor-related protein gene exon 3 polymorphism in Alzheimer's disease.

The low density lipoprotein receptor-related protein (LRP) may influence both the clearance and the production of beta-amyloid peptide and thus plays a role in Alzheimer's disease (AD) pathogenesis. Previous studies, although inconsistent, have suggested that the LRP exon 3 CC genotype contributes to the risk of AD. A case-control study utilizing a clinically well-defined group of 305 sporadic AD patients and 304 control subjects was performed to test this association in an ethnically homogeneous population from Spain. In the current study, the LRP CC genotype was not over-represented in AD patients compared to non-demented controls. A meta-analysis of previous studies revealed a weak correlation of LRP CC genotype with AD (odds ratio of 1.35, P=0.01).

Tubulin folding is altered by mutations in a putative GTP binding motif.

Tubulins contain a glycine-rich loop, that has been implicated in microtubule dynamics by means of an intramolecular interaction with the carboxy-terminal region. As a further extension of the analysis of the role of the carboxy-terminal region in tubulin folding we have mutated the glycine-rich loop of tubulin subunits. An alpha-tubulin point mutant with a T150-->G substitution (the corresponding residue present in beta-tubulin) was able to incorporate into dimers and microtubules. On the other hand, four beta-tubulin point mutants, including the G148-->T substitution, did not incorporate into dimers, did not release monomers, but were able to form C900 and C300 complexes (intermediates in the process of tubulin folding). Three other mutants within this region (which approximately encompasses residues 137-152) were incapable of forming dimers and C300 complexes but gave rise to the formation of C900 complexes. These results suggest that tubulin goes through two sequential folding states during the folding process, first in association with TCP1-complexes (C900) prior to the transfer to C300 complexes. It is this second step that implies binding/hydrolysis of GTP, reinforcing our previous proposed model for tubulin folding and assembly.

Beta-tubulin folding is modulated by the isotype-specific carboxy-terminal domain.

To investigate the contribution of the carboxy-terminal domain in the process of tubulin folding and dimer formation, we constructed a beta 1-beta 3 tubulin chimaera and two truncated carboxy-terminal beta 3-tubulins. The capacity of these altered polypeptides to incorporate into dimers and into microtubules was tested by non-denaturing electrophoresis and co-assembly experiments. The chimaera and the truncated protein with a deletion encompassing the last 12 amino acid residues (beta 3 delta C12) were incorporated into dimers and microtubules, though the level of incorporation was diminished compared to wild-type beta 3-tubulin. However, the level of incorporation of beta 3 delta C12 into subtilisin-digested dimers was similar to the incorporation of wild-type beta 3-tubulin. Since subtilisin deletes the carboxy-terminal region, these results suggest a regulatory role of the carboxy-terminal region in the folding process itself and not in the formation of the dimer.

A 14 kDa release factor is involved in GTP-dependent beta-tubulin folding.

The tubulin folding pathway is a model system to understand protein folding in the cell. It involves the interaction of several chaperones, including TCP-1 and other as yet uncharacterized factors. Release of tubulin monomers from folding intermediates (C900 and C300) and their incorporation into tubulin dimers is dependent on GTP hydrolysis, magnesium ions and release factors. In this work, we have purified to homogeneity the protein factor responsible for the release of beta-tubulin monomers from C300 complexes. It has an apparent molecular mass of 14 kDa (p14) as judged by SDS electrophoresis. The protein behaved as a dimer of about 28 kDa when analyzed by gel filtration chromatography. Furthermore, the p14-dependent release of beta-tubulin monomers from C300 complexes takes place in the presence of GTP. These results suggest that p14 is a new chaperone that assists in tubulin folding by facilitating the acquisition of the native conformation.

Incorporation of tubulin subunits into dimers requires GTP hydrolysis.

A toroid multisubunit complex of 800-900 kDa has been implicated in assisting protein folding of at least two cytoplasmic proteins, actin and tubulin. This process is dependent on the presence of magnesium ions and ATP hydrolysis. In vitro translation of cDNAs encoding different alpha- and beta-tubulin isotypes also gives rise to the formation of complexes of about 300 kDa. These complexes have been functionally implicated in the incorporation of tubulin monomers within the tubulin heterodimer. This work shows that, in addition to ATP hydrolysis, the incorporation of newly synthesized tubulin subunits into functional heterodimers requires GTP hydrolysis in the presence of magnesium ions. A two-step process is suggested, a first ATP-dependent step in which the 900 kDa complexes are implicated in a similar way to the step taking place in actin folding, and a second GTP-dependent step in which the 300 kDa complexes are involved in the assembly of the heterodimer.