RESUMO
Autism spectrum disorder (ASD) is a group of neurological and developmental disabilities characterised by clinical and genetic heterogeneity. The current study aimed to expand ASD genotyping by investigating potential associations with SYNE2 mutations. Specifically, the disease-causing variants of SYNE2 in 410 trios manifesting neurodevelopmental disorders using whole-exome sequencing were explored. The consequences of the identified variants were studied at the transcript level using quantitative polymerase chain reaction (qPCR). For validation, immunofluorescence and immunoblotting were performed to analyse mutational effects at the protein level. The compound heterozygous variants of SYNE2 (NM_182914.3:c.2483T>G; p.(Val828Gly) and NM_182914.3:c.2362G>A; p.(Glu788Lys)) were identified in a 4.5-year-old male, clinically diagnosed with autism spectrum disorder, developmental delay and intellectual disability. Both variants reside within the nesprin-2 giant spectrin repeat (SR5) domain and are predicted to be highly damaging using in silico tools. Specifically, a significant reduction of nesprin-2 giant protein levels is revealed in patient cells. SYNE2 transcription and the nuclear envelope localisation of the mutant proteins was however unaffected as compared to parental control cells. Collectively, these data provide novel insights into the cardinal role of the nesprin-2 giant in neurodevelopment and suggest that the biallelic hypomorphic SYNE2 mutations may be a new cause of intellectual disability and ASD.
Assuntos
Transtorno do Espectro Autista/genética , Deficiência Intelectual/genética , Proteínas dos Microfilamentos/genética , Proteínas do Tecido Nervoso/genética , Transtorno do Espectro Autista/metabolismo , Transtorno do Espectro Autista/patologia , Células Cultivadas , Criança , Heterozigoto , Humanos , Deficiência Intelectual/metabolismo , Deficiência Intelectual/patologia , Masculino , Proteínas dos Microfilamentos/química , Proteínas dos Microfilamentos/metabolismo , Mutação de Sentido Incorreto , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/metabolismo , Domínios Proteicos , Transporte ProteicoRESUMO
Mechanotransduction is defined as the ability of cells to sense mechanical stimuli from their surroundings and translate them into biochemical signals. Epidermal keratinocytes respond to mechanical cues by altering their proliferation, migration, and differentiation. In vitro cell culture, however, utilises tissue culture plastic, which is significantly stiffer than the in vivo environment. Current epidermal models fail to consider the effects of culturing keratinocytes on plastic prior to setting up three-dimensional cultures, so the impact of this non-physiological exposure on epidermal assembly is largely overlooked. In this study, primary keratinocytes cultured on plastic were compared with those grown on 4, 8, and 50 kPa stiff biomimetic hydrogels that have similar mechanical properties to skin. Our data show that keratinocytes cultured on biomimetic hydrogels exhibited major changes in cellular architecture, cell density, nuclear biomechanics, and mechanoprotein expression, such as specific Linker of Nucleoskeleton and Cytoskeleton (LINC) complex constituents. Mechanical conditioning of keratinocytes on 50 kPa biomimetic hydrogels improved the thickness and organisation of 3D epidermal models. In summary, the current study demonstrates that the effects of extracellular mechanics on keratinocyte cell biology are significant and therefore should be harnessed in skin research to ensure the successful production of physiologically relevant skin models.
