Faster regeneration associated to high expression of Fam65b and Hdac6 in dysferlin-deficient mouse.

Dysferlin is a sarcolemmal muscle protein associated with the processes of membrane repair, trafficking, and fusion of intracellular vesicles and muscle regeneration. Mutations in the DYSF gene cause clinically distinct forms of muscular dystrophies. The dysferlin-deficient SJL/J mouse model presents a reduction of 85% of the protein but shows mild weakness and discrete histopathological alterations. To study the effect of dysferlin deficiency in the muscle regenerative process, we used a model of electrical injury by electroporation to induce muscle degeneration/regeneration in the SJL/J mouse. The relative expression of the genes Pax7, MyoD, Myf5, and Myog was accompanied by the histopathological evaluation during muscle recovery at different time points after injury. We also investigated the effects of dysferlin deficiency in the expression of genes encoding FAM65B and HDAC6 proteins, recently described as forming a tricomplex with dysferlin at the beginning of myoblast differentiation. We observed an altered time course through the process of degeneration and regeneration in dysferlin-deficient mice, with remarkable regenerative capacity characterized by a faster and effective response in the first days after injury, as compared to the WT mice. Also, dysferlin deficiency seems to significantly alter the gene expression of Fam65b and Hdac6 during regeneration, since higher levels of expression of both genes were observed in dysferlin-deficient mice. These results need further attention to define their relevance in the disease mechanism.

10q23.31 microduplication encompassing PTEN decreases mTOR signalling activity and is associated with autosomal dominant primary microcephaly.

BACKGROUND: Hereditary primary microcephaly (MCPH) is mainly characterised by decreased occipitofrontal circumference and variable degree of intellectual disability. MCPH with a dominant pattern of inheritance is a rare condition, so far causally linked to pathogenic variants in the ALFY, DPP6, KIF11 and DYRK1A genes. OBJECTIVE: This study aimed at identifying the causative variant of the autosomal dominant form of MCPH in a Brazilian family with three affected members. METHODS: Following clinical evaluation of two sibs and their mother presenting with autosomal dominant MCPH, array comparative genome hybridisation was performed using genomic DNA from peripheral blood of the family members. Gene and protein expression studies were carried out in cultured skin fibroblasts. RESULTS: A 382 kb microduplication at 10q23.31 was detected, encompassing the entire PTEN, KLLN and ATAD1 genes. PTEN haploinsufficiency has been causally associated with macrocephaly and autism spectrum disorder and, therefore, was considered the most likely candidate gene to be involved in this autosomal dominant form of MCPH. In the patients' fibroblasts, PTEN mRNA and protein were found to be overexpressed, and the phosphorylation patterns of upstream and downstream components of the mammalian target of rapamycin (mTOR) signalling pathway were dysregulated. CONCLUSIONS: Taken together, our results demonstrate that the identified submicroscopic 10q23.31 duplication in a family with MCPH leads to markedly increased expression of PTEN and reduced activity of the mTOR signalling pathway. These results suggest that the most probable pathomechanism underlying the microcephaly phenotype in this family involves downregulation of the mTOR pathway through overexpression of PTEN.

EIF4A3 deficient human iPSCs and mouse models demonstrate neural crest defects that underlie Richieri-Costa-Pereira syndrome.

Biallelic loss-of-function mutations in the RNA-binding protein EIF4A3 cause Richieri-Costa-Pereira syndrome (RCPS), an autosomal recessive condition mainly characterized by craniofacial and limb malformations. However, the pathogenic cellular mechanisms responsible for this syndrome are entirely unknown. Here, we used two complementary approaches, patient-derived induced pluripotent stem cells (iPSCs) and conditional Eif4a3 mouse models, to demonstrate that defective neural crest cell (NCC) development explains RCPS craniofacial abnormalities. RCPS iNCCs have decreased migratory capacity, a distinct phenotype relative to other craniofacial disorders. Eif4a3 haploinsufficient embryos presented altered mandibular process fusion and micrognathia, thus recapitulating the most penetrant phenotypes of the syndrome. These defects were evident in either ubiquitous or NCC-specific Eif4a3 haploinsufficient animals, demonstrating an autonomous requirement of Eif4a3 in NCCs. Notably, RCPS NCC-derived mesenchymal stem-like cells (nMSCs) showed premature bone differentiation, a phenotype paralleled by premature clavicle ossification in Eif4a3 haploinsufficient embryos. Likewise, nMSCs presented compromised in vitro chondrogenesis, and Meckel's cartilage was underdeveloped in vivo. These findings indicate novel and essential requirements of EIF4A3 for NCC migration and osteochondrogenic differentiation during craniofacial development. Altogether, complementary use of iPSCs and mouse models pinpoint unique cellular mechanisms by which EIF4A3 mutation causes RCPS, and provide a paradigm to study craniofacial disorders.