Biology subjects

Vanyai, H.

Publications and source records attributed to Vanyai, H..

5 recordsLinked to original sources

bioRxiv2024

Transcriptomic alterations including p53 pathway dysregulation prime DNMT3A mutant cells for transformation

DNMT3A mutations are prevalent in haematologic malignancies. Our mouse model introduced the murine homologue (R878H) of the human hotspot R882H mutation into the mouse Dnmt3a locus, resulting in globally reduced DNA methylation in all tissues. Mice with heterozygous R878H mutations developed {gamma}-radiation induced thymic lymphoma more rapidly than control mice, suggesting a vulnerability to stress stimuli in Dnmt3aR878H/+ cells. In competitive transplantations, Dnmt3aR878H/+ Lin-Sca-1+Kit+ (LSK) cells had a competitive advantage over wt cells, indicating a self-renewal phenotype at the expense of differentiation. RNA-sequencing of Dnmt3aR878H/+ LSKs exposed to low dose {gamma}-radiation showed downregulation of the p53 pathway. Accordingly, reduced PUMA expression was observed by flow cytometry in the bone marrow of {gamma}-irradiated Dnmt3aR878H/+ mice due to altered p53 signalling. These findings provide new insights as to how DNMT3A mutations cause subtle changes in the transcriptome of LSK cells which contribute to their increased self-renewal and propensity for malignant transformation. SIGNIFICANCEHotspot DNMT3A R882H mutations are overrepresented in leukaemia suggesting that they confer a susceptibility to disease if further mutations are acquired. To advance therapies for DNMT3A mutant disease, it is essential to understand the mechanisms by which this mutation primes cells for malignant transformation.

cancer biology↗

bioRxiv2024

Slc35a2 mosaic knockout impacts cortical development, dendritic arborisation, and neuronal firing in the developing brain

ObjectiveMild malformation of cortical development with oligodendroglial hyperplasia in epilepsy (MOGHE) is an important cause of drug-resistant epilepsy. A significant subset of individuals diagnosed with MOGHE display somatic mosaicism for loss-of-function variants in SLC35A2, which encodes the UDP-galactose transporter. We developed a mouse model to investigate the mechanism by which disruption of this transporter leads to a malformation of cortical development. MethodsWe used in utero electroporation and CRISPR/Cas9 to knockout Slc35a2 in a subset of layer 2/3 cortical neuronal progenitors in the developing brains of fetal mice to model mosaic expression. ResultsHistology of brain tissue in the mosaic Slc35a2 knockout mice revealed the presence of upper layer-derived cortical neurons in the white matter. In contrast, oligodendrocyte patterning was unchanged. Reconstruction of single filled neurons identified altered dendritic arborisation with Slc35a2 knockout neurons having increased complexity. Whole-cell electrophysiological recordings revealed that Slc35a2 knockout neurons display reduced action potential firing and increased afterhyperpolarisation duration compared with control neurons. Mosaic Slc35a2 knockout mice also exhibited significantly increased epileptiform spiking and increased locomotion. InterpretationWe successfully generated a mouse model of mosaic Slc35a2 deficiency, which recapitulates features of the human phenotype, including impaired neuronal migration. We show that knockout in layer 2/3 cortical neuron progenitors is sufficient to disrupt neuronal excitability and increase epileptiform activity and hyperactivity in mosaic mice. Our mouse model provides a unique opportunity to investigate the disease mechanism(s) that underpin MOGHE and facilitate the development of precision therapies.

neuroscience↗

bioRxiv2022

PKN2 deficiency leads both to prenatal congenital cardiomyopathy and defective angiotensin II stress responses.

BackgroundThe protein kinase PKN2 is required for embryonic development, and PKN2 knockout mice die as a result of failure in expansion of mesoderm tissues, cardiac development and neural tube closure. In the adult, cardiomyocyte PKN2 and PKN1 (in combination) are required for cardiac adaptation to pressure-overload. The role of PKN2 in contractile cardiomyocytes during development and its role in the adult heart remain to be fully established. MethodsWe used mice with cardiomyocyte-directed knockout of PKN2 or global PKN2 haploinsufficiency. Cardiac function and dimensions were assessed with high resolution episcopic microscopy, MRI, micro-CT and echocardiography. Biochemical and histological changes were assessed. ResultsCardiomyocyte-directed PKN2 knockout embryos displayed striking abnormalities in the compact myocardium, with frequent myocardial clefts and diverticula, ventricular septal defects and abnormal heart shape. The sub-Mendelian homozygous knockout survivors developed cardiac failure. RNASeq data showed upregulation of PKN2 in patients with dilated cardiomyopathy, suggesting an involvement in adult heart disease. Given the rarity of homozygous survivors with cardiomyocyte-specific deletion of PKN2, this was explored using mice with constitutive heterozygous PKN2 knockout. Cardiac hypertrophy resulting from hypertension induced by angiotensin II was reduced in haploinsufficient PKN2 mice relative to wild-type littermates, with suppression of cardiomyocyte hypertrophy and cardiac fibrosis. ConclusionsCardiomyocyte PKN2 is essential for heart development and formation of compact myocardium, and is also required for cardiac hypertrophy in hypertension. Thus, PKN signalling may offer therapeutic options for managing congenital and adult heart diseases.

developmental biology↗

bioRxiv2021

SMCHD1 has separable roles in chromatin architecture and gene silencing that could be targeted in disease

The interplay between 3D chromatin architecture and gene silencing is incompletely understood. Here, we report a novel point mutation in the non-canonical SMC protein SMCHD1 that enhances its silencing capacity at endogenous developmental targets and at the facioscapulohumeral muscular dystrophy associated macro-array, D4Z4. Heightened SMCHD1 silencing perturbs developmental Hox gene activation, causing a homeotic transformation in mice. Paradoxically, the mutant SMCHD1 appears to enhance insulation against another epigenetic regulator complex, PRC2, while depleting long range chromatin interactions akin to what is observed in the absence of SMCHD1. These data suggest that SMCHD1s role in long range chromatin interactions is not directly linked to gene silencing or insulating the chromatin, refining the model for how the different levels of SMCHD1-mediated chromatin regulation interact to bring about gene silencing in normal development and disease.

molecular biology↗

bioRxiv2019

Mask family proteins ANKHD1 and ANKRD17 regulate YAP nuclear import, stability and phase separation.

The Mask family of multiple ankyrin repeat and KH domain proteins were discovered in Drosophila to promote the activity of the transcriptional coactivator Yorkie (Yki), the sole fly homolog of mammalian YAP (YAP1) and TAZ (WWTR1). The molecular function of Mask, or its mammalian homologs Mask1 (ANKHD1) and Mask2 (ANKRD17), remains unclear. Mask family proteins contain two Ankyrin repeat domains that bind Yki/YAP as well as a conserved nuclear localisation sequence (NLS) and nuclear export sequence (NES), suggesting a role in nucleo-cytoplasmic transport. Here we show that Mask acts to promote nuclear import of Yki, and that addition of an ectopic NLS to Yki is sufficient to bypass the requirement for Mask in Yki-driven tissue growth. Mammalian Mask1/2 proteins also promote nuclear import of YAP, as well as stabilising YAP and driving colloidal phase separation into large liquid droplets. Mask1/2 and YAP normally colocalise in a granular fashion in both nucleus and cytoplasm, and are co-regulated during mechanotransduction. Our results suggest that Mask family proteins promote YAP nuclear import and phase separation to regulate YAP stability and transcriptional activity.

developmental biology↗