Familial dysautonomia (FD), a hereditary sensory and autonomic neuropathy, is caused by missplicing of exon 20, resulting from an intronic mutation in the inhibitor of kappa light polypeptide gene enhancer in B cells, kinase complex-associated protein (IKBKAP) gene encoding IKK complex-associated protein (IKAP)/elongator protein 1 (ELP1). A newly established splicing reporter assay allowed us to visualize pathogenic splicing in cells and to screen small chemicals for the ability to correct the aberrant splicing of IKBKAP. Using this splicing reporter, we screened our chemical libraries and identified a compound, rectifier of aberrant splicing (RECTAS), that rectifies the aberrant IKBKAP splicing in cells from patients with FD. Here, we found that the levels of modified uridine at the wobble position in cytoplasmic tRNAs are reduced in cells from patients with FD and that treatment with RECTAS increases the expression of IKAP and recovers the tRNA modifications. These findings suggest that the missplicing of IKBKAP results in reduced tRNA modifications in patients with FD and that RECTAS is a promising therapeutic drug candidate for FD. IKAP is currently known as elongator protein 1 (ELP1), an integral component of the human Elongator complex, which was originally identified in Saccharomyces cerevisiae and shown to be well conserved among species (1). Although multiple functions of IKAP/ELP1 in JNK signaling, neuronal development during embryogenesis, exocytosis, and actin cytoskeleton regulation have been reported (reviewed in refs. 2, 3), yeast genetic analyses have shown that the Elongator complex is also required for the formation of the C5-substituent of 5-carbamoylmethyl (ncm 5 ), 5-methoxycarbonylmethyl (mcm 5 ), and its derivatives at the wobble uridine in tRNAs recognizing purine-ending codons (4, 5). Most recently, it was demonstrated that conditional IKAP/Elp1 KO in mouse testes results in male infertility by disrupting meiotic progression, along with the reduction of modified nucleosides [5-methoxycarbonylmethyl uridine (mcm 5 U), 5-carbamoylmethyl uridine (ncm 5 U), and 5-methoxycarbonylmethyl-2-thiouridine (mcm 5 s 2 U)] of total tRNAs in the testes (6). These modifications are highly likely to play critical roles in the maintenance of translational fidelity, suggesting that the defects in these modifications lead to the mistranslation of various proteins.Familial dysautonomia (FD; Riley-Day syndrome), an autosomal recessive neurodegenerative disease, is characterized by impaired development and progressive degeneration of the sensory and autonomic nerves. Patients who have FD exhibit various symptoms, including cardiovascular instability, recurrent pneumonia, vomiting/dysautonomic crisis, gastrointestinal dysfunction, decreased sensitivity to pain and temperature, and defective lacrimation. FD is a very common disorder in the Ashkenazi Jewish population, with a carrier frequency of 1 in 27. More than 99% of patients who have FD harbor a homozygous mutation in intron 20 (IVS20 + 6T > C: FD mutation)...

The 5' splice site mutation (IVS20+6T>C) of the () gene in familial dysautonomia (FD) is at the sixth intronic nucleotide of the 5' splice site. It is known to weaken U1 snRNP recognition and result in an aberrantly spliced mRNA product in neuronal tissue, but normally spliced mRNA in other tissues. Aberrantly spliced mRNA abrogates IKK complex-associated protein (IKAP)/elongator protein 1 (ELP1) expression and results in a defect of neuronal cell development in FD. To elucidate the tissue-dependent regulatory mechanism, we screened an expression library of major RNA-binding proteins (RBPs) with our mammalian dual-color splicing reporter system and identified RBM24 as a regulator. RBM24 functioned as a cryptic intronic splicing enhancer binding to an element (IVS20+13-29) downstream from the intronic 5' splice site mutation in the gene and promoted U1 snRNP recognition only to the mutated 5' splice site (and not the wild-type 5' splice site). Our results show that tissue-specific expression of RBM24 can explain the neuron-specific aberrant splicing of exon 20 in familial dysautonomia, and that ectopic expression of RBM24 in neuronal tissue could be a novel therapeutic target of the disease.

Three major organic matrix components, nacrein, MSI60 and N16 have been reported from the nacreous layer of Japanese pearl oyster, Pinctada fucata. Though several in vitro experiments have been carried out to elucidate the functions of these molecules details have not yet been clarified. In this report, we tempt to clarify the gene expression levels encoding the above three proteins between samples of 1) summer and winter seasons and 2) ocean and aquarium environments by using realtime polymerase chain reaction (PCR).It was confirmed that the biomineralization process of P. fucata is mainly influenced by the circatidal rhythm of the ocean environment. The gene expressions coding for N16 and MSI60 increased at the time of high tide, while that of nacrein increased at the time of low tide. The similar tendency observed in N16 and MSI60 showed the possibility that both components are secreted simultaneously, supporting a hypothesis that N16 forms crosslinkage with MSI60 to form the membrane. The expressions of MSI60, N16 and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes were remarkable in winter season, while no variation was found in the expression level of the nacrein gene in summer and winter season.The study is the first attempt regarding the seasonal and circadian rhythms observed on gene expressions incorporated into molluscan shell formation. The results will give a new insight into the relationship between molluscan physiology and the mechanism of shell formation.