Cryptic splicing events in the iron transporter ABCB7 and other key target genes in SF3B1-mutant myelodysplastic syndromes.

The splicing factor SF3B1 is the most frequently mutated gene in myelodysplastic syndromes (MDS), and is strongly associated with the presence of ring sideroblasts (RS). We have performed a systematic analysis of cryptic splicing abnormalities from RNA sequencing data on hematopoietic stem cells (HSCs) of SF3B1-mutant MDS cases with RS. Aberrant splicing events in many downstream target genes were identified and cryptic 3' splice site usage was a frequent event in SF3B1-mutant MDS. The iron transporter ABCB7 is a well-recognized candidate gene showing marked downregulation in MDS with RS. Our analysis unveiled aberrant ABCB7 splicing, due to usage of an alternative 3' splice site in MDS patient samples, giving rise to a premature termination codon in the ABCB7 mRNA. Treatment of cultured SF3B1-mutant MDS erythroblasts and a CRISPR/Cas9-generated SF3B1-mutant cell line with the nonsense-mediated decay (NMD) inhibitor cycloheximide showed that the aberrantly spliced ABCB7 transcript is targeted by NMD. We describe cryptic splicing events in the HSCs of SF3B1-mutant MDS, and our data support a model in which NMD-induced downregulation of the iron exporter ABCB7 mRNA transcript resulting from aberrant splicing caused by mutant SF3B1 underlies the increased mitochondrial iron accumulation found in MDS patients with RS.

Disruption of SF3B1 results in deregulated expression and splicing of key genes and pathways in myelodysplastic syndrome hematopoietic stem and progenitor cells.

The splicing factor SF3B1 is the most commonly mutated gene in the myelodysplastic syndrome (MDS), particularly in patients with refractory anemia with ring sideroblasts (RARS). We investigated the functional effects of SF3B1 disruption in myeloid cell lines: SF3B1 knockdown resulted in growth inhibition, cell cycle arrest and impaired erythroid differentiation and deregulation of many genes and pathways, including cell cycle regulation and RNA processing. MDS is a disorder of the hematopoietic stem cell and we thus studied the transcriptome of CD34(+) cells from MDS patients with SF3B1 mutations using RNA sequencing. Genes significantly differentially expressed at the transcript and/or exon level in SF3B1 mutant compared with wild-type cases include genes that are involved in MDS pathogenesis (ASXL1 and CBL), iron homeostasis and mitochondrial metabolism (ALAS2, ABCB7 and SLC25A37) and RNA splicing/processing (PRPF8 and HNRNPD). Many genes regulated by a DNA damage-induced BRCA1-BCLAF1-SF3B1 protein complex showed differential expression/splicing in SF3B1 mutant cases. This is the first study to determine the target genes of SF3B1 mutation in MDS CD34(+) cells. Our data indicate that SF3B1 has a critical role in MDS by affecting the expression and splicing of genes involved in specific cellular processes/pathways, many of which are relevant to the known RARS pathophysiology, suggesting a causal link.

Referential communication skills in children with Down Syndrome.

AIM: The aim of this study was to investigate referential communication skills in children with Down Syndrome (DS). We studied the development of pragmatic language in relation to nonverbal skills by analyzing referential communication skills in children with DS, in both production and comprehension of referential messages: oriented and/or ambiguous, adequate (appropriate) and inadequate (inappropriate). METHODS: The study population was 24 children: 12 with DS (mean age, 7.8 years) and 12 with normal development (control group) (mean age, 7.4 years). In the first phase (F1), all 24 children were administered a test to evaluate their nonverbal skills, followed in a second phase (F2) by a test to specifically evaluate their referential communication skills. RESULTS: A strong relationship between nonverbal and referential communication skills was found, in which the nonverbal skills contributed to variations in referential performance, especially among the children with DS. Furthermore, the children with DS showed better performance in the "talking" than the "listener" condition. Within the listener condition, a significant correlation emerged between adequate and inadequate messages. CONCLUSION: Referential communication skills are influenced by several verbal and nonverbal factors.

Scanning tunneling spectroscopy simulations of poly(3-dodecylthiophene) chains adsorbed on highly oriented pyrolytic graphite.

We report on a hybrid scheme to perform efficient and accurate simulations of scanning tunneling spectroscopy (STS) of molecules weakly bonded to surfaces. Calculations are based on a tight binding (TB) technique, including a self-consistent calculation of the electronic structure of the molecule, to predict STS conductance spectra. The use of a local basis makes our model easily applicable to systems with several hundreds of atoms. We performed first-principles density-functional calculations to extract the geometrical and electronic properties of the system. In this way, we can include, in the TB scheme, the effects of structural relaxation upon adsorption on the electronic structure of the molecule. This approach is applied to the study of regioregular poly(3-dodecylthiophene) polymer chains adsorbed on highly oriented pyrolytic graphite. Results of spectroscopic calculations are discussed and compared with recently obtained experimental data.