Role of minor splicing in brain development – U4ATAC-BRAIN

In seminal papers published in 2011, our group and others identified mutations in a non-coding gene, RNU4ATAC, in an autosomal recessive disorder named microcephalic osteodysplastic primordial dwarfism type 1 (MOPD1, OMIM 210710) or Taybi-Linder syndrome (TALS). This disorder is characterized by multiple malformations including severe microcephaly, severe cortical brain malformations (neuronal migration defects), corpus callosum agenesis/dysgenesis, cerebellar vermis hypoplasia, mental retardation, severe ante and postnatal growth retardation and early unexplained death occurring within the first two years of life in more than 70% of the cases.
U4atac is a non-coding small nuclear (sn) RNA of 131 nucleotides; snRNAs are RNA components of the spliceosome, a ribonucleoprotein complex catalysing the removal of introns from pre-mRNAs to produce mature mRNAs. Two highly homologous spliceosomes exist in vertebrates and other species, the U2-dependent (major) spliceosome which splices the vast majority of introns, and the U12-dependent (minor) spliceosome, which removes ~850 introns located in 700 genes (that also contain U2 introns). Spliceosomes are formed by the sequential interactions of >100 proteins and five snRNAs (U1, U2, U4, U5 and U6 for the major spliceosome, U11, U12, U4atac, U5 and U6atac for the minor one) that interact with one another and with the pre-mRNA substrate. Interestingly, other very rare congenital disorders named Roifman syndrome (RFMN, OMIM 616651) and Lowry Wood syndrome (LWS, OMIM 226960) have also been recently associated with biallelic RNU4ATAC mutations. While both RFMN and LWS have features overlapping with TALS (i.e. microcephaly, growth retardation, skeletal dysplasia, intellectual disability), it is worth noting that RFMN cases have a specific antibody deficiency that is the hallmark of this rare immunodeficiency syndrome.
Our preliminary unpublished results provide direct evidence for minor intron retentions in TALS patient-derived fibroblast and lymphoblast cells. How these minor splicing defects mediate brain developmental anomalies remains unknown. In this work we will combine multiple approaches including clinical genetics, genomics, transcriptomics, bioinformatics, biochemistry, molecular biology, cell biology and zebrafish models to understand the cascade of events linking RNU4ATAC mutations to brain developmental anomalies. Towards this aim, we will search for a possible defect in U4atac-U6atac.U5 trisnRNP assembly in RNU4ATAC mutant cells, which could explain the minor spliceosome dysfunction. We will also study the expression and splicing pattern of U12 genes in control individuals. Furthermore, we will improve the knowledge on the spectrum of RNU4ATAC variants and their resulting phenotypes, which will help providing an accurate medical management of the patients affected with a RNU4ATAC-associated phenotype and an informed genetic counselling of their family.