CTNS mRNA molecular analysis revealed a novel mutation in a child with infantile nephropathic cystinosis: a case report.

BACKGROUND: Cystinosis is an autosomal recessive lysosomal storage disorder characterized by accumulation of cystine in lysosomes throughout the body. Cystinosis is caused by mutations in the CTNS gene that encodes the lysosomal cystine carrier protein cystinosin. CTNS mutations result in either complete absence or reduced cystine transporting function of the protein. The diagnosis of nephropathic cystinosis is generally based on measuring leukocyte cystine level, demonstration of corneal cystine crystals by the slit lamp examination and confirmed by genetic analysis of the CTNS gene. CASE PRESENTATION: A boy born to consanguineous Caucasian parents had the characteristic clinical features of the infantile nephropathic cystinosis including renal Fanconi syndrome (polydipsia/polyuria, metabolic acidosis, hypokalemia, hypophosphatemia, low molecular weight proteinuria, glycosuria, cystine crystals in the cornea) and elevated WBC cystine levels. Initially we performed RFLP analysis of the common in the Northern European population 57-kb deletion of proband's DNA, then a direct Sanger sequencing which revealed no mutations in the coding part of the CTNS gene. To confirm the diagnosis we performed RT-PCR analysis of total RNA obtained from patient-derived fibroblasts in combination with cDNA sequencing. This revealed the skipping of exon 4 and exon 5 in the CTNS in our patient. Therefore, we detected a novel 9-kb homozygous deletion in the CTNS gene at genomic DNA level, spanning region from intron 3 to intron 5. In order to identify the inheritance pattern of the deletion we analyzed DNA of proband's mother and father. Both parents were found to be heterozygous carriers of the CTNS mutation. CONCLUSIONS: Analysis of CTNS gene transcript allowed to identify a large homozygous deletion in the patient with infantile nephropathic cystinosis. Mutational detection at RNA level may be an efficient tool to establish the genetic defect in some cystinosis patients.

Nedd9 restrains renal cystogenesis in Pkd1-/- mice.

Mutations inactivating the cilia-localized Pkd1 protein result in autosomal dominant polycystic kidney disease (ADPKD), a serious inherited syndrome affecting ∼ 1 in 500 people, in which accumulation of renal cysts eventually destroys kidney function. Severity of ADPKD varies throughout the population, for reasons thought to involve differences both in intragenic Pkd1 mutations and in modifier alleles. The scaffolding protein NEDD9, commonly dysregulated during cancer progression, interacts with Aurora-A (AURKA) kinase to control ciliary resorption, and with Src and other partners to influence proliferative signaling pathways often activated in ADPKD. We here demonstrate Nedd9 expression is deregulated in human ADPKD and a mouse ADPKD model. Although genetic ablation of Nedd9 does not independently influence cystogenesis, constitutive absence of Nedd9 strongly promotes cyst formation in the tamoxifen-inducible Pkd1fl/fl;Cre/Esr1(+) mouse model of ADPKD. This cystogenic effect is associated with striking morphological defects in the cilia of Pkd1(-/-);Nedd9(-/-) mice, associated with specific loss of ciliary localization of adenylase cyclase III in the doubly mutant genotype. Ciliary phenotypes imply a failure of Aurora-A activation: Compatible with this idea, Pkd1(-/-);Nedd9(-/-) mice had ciliary resorption defects, and treatment of Pkd1(-/-) mice with a clinical Aurora-A kinase inhibitor exacerbated cystogenesis. In addition, activation of the ADPKD-associated signaling effectors Src, Erk, and the mTOR effector S6 was enhanced, and Ca(2+) response to external stimuli was reduced, in Pkd1(-/-);Nedd9(-/-) versus Pkd1(-/-) mice. Together, these results indicated an important modifier action of Nedd9 on ADPKD pathogenesis involving failure to activate Aurora-A.

Cilia and cilia-associated proteins in cancer.

The primary cilium is a well-established target in the pathogenesis of numerous developmental and chronic disorders, and more recently is attracting interest as a structure relevant to cancer. Here we discuss mechanisms by which changes in cilia can contribute to the formation and growth of tumors. We emphasize the cancer-relevance of cilia-dependent signaling pathways and proteins including mTOR, VHL, TSC, WNT, Aurora-A, NEDD9, and Hedgehog, and highlight the emerging role of ciliary dysfunction in renal cell carcinoma, medulloblastoma, and breast cancer.

mRNA analysis revealed a novel pathogenic EIF2S3 variant causing MEHMO syndrome.

EIF2S3 pathogenic variants have been shown to cause MEHMO syndrome - a rare X-linked intellectual disability syndrome. In most cases, DNA diagnostics of MEHMO syndrome is performed using exome sequencing. We describe two cousins with profound intellectual disability, severe microcephaly, microgenitalism, hypoglycemia, epileptic seizures, and hypertrichosis, whose clinical symptoms allowed us to suspect MEHMO syndrome. To confirm this diagnosis, we designed an mRNA analysis for the EIF2S3 gene. It is a cost-effective method to detect coding sequence variants in multi-exonic genes, as well as splicing defects and allelic imbalance. Our mRNA sequence analysis revealed a novel EIF2S3 variant c.820C>G in both cousins. We also found the same variant in female family members in the heterozygous state. To investigate the pathogenicity of the c.820C>G variant, we performed expression analysis, which showed that the DDIT3 transcript level was significantly increased in the patient relative to the controls. We, thus, demonstrate that mRNA analysis is an efficient tool for performing genetic testing in patients with distinct phenotypic features.