Biology subjects

Misztal, K.

Publications and source records attributed to Misztal, K..

5 recordsLinked to original sources

bioRxiv2026

Enhancer-mediated metabolic pre-patterning defines trabecular cardiomyocyte identity prior to morphogenesis

Cardiac trabeculae are essential structures of the vertebrate heart critical for ventricular wall integrity and contractility. Despite their importance, the transcriptional regulatory program defining trabecular cardiomyocyte identity and driving their development remains poorly understood. We identify a novel enhancer, -6.8got2b, that drives gene expression specifically in the trabecular myocardium of the zebrafish heart. Functional analyses revealed that - 6.8got2b is required for trabeculae formation, likely acting through its target gene, got2b, which encodes a mitochondrial enzyme. We delineated a 166bp core region responsible for restricting gene expression to trabecular cardiomyocytes, harboring binding motifs for multiple transcription factors (TFs), including Tbx20, whose motif deletion abolished trabeculae-specific expression. A human genomic region sharing partial sequence similarity and TF motifs with -6.8got2b was found near GOT2P4 - a pseudogene related to the human GOT2 gene. Strikingly, the human sequence drove expression in the zebrafish trabeculae and ventricular myocardium, indicating partial but functionally relevant conservation. Using - 6.8got2b as a molecular marker, we transcriptionally profiled trabecular cardiomyocytes and show that, compared to compact layer cardiomyocytes, they exhibit distinct gene signatures indicative of transcriptional priming for elevated energy metabolism prior to the formation of overt trabecular structure. Our findings reveal a conserved -6.8got2b-got2b regulatory axis driving trabecular cardiomyocyte development downstream of Tbx20, establishing a new layer of transcriptional regulation in heart morphogenesis.

developmental biology↗

bioRxiv2025

p70S6 kinase-dependent phosphorylation of the μ2 subunit of the AP2 adaptor complex is needed for clathrin-mediated endocytosis.

Clathrin-mediated endocytosis (CME) internalizes cell-surface receptors via clathrin-coated invaginations of the plasma membrane. Both clathrin and endocytic cargo are recruited to these sites by the adaptor protein complex AP2. AP2 cycles between a closed cytoplasmic conformation and an open membrane-bound state, and efficient CME requires both conformations and their dynamic interconversion. The mechanisms regulating these conformational changes, which include post-translational modifications of the AP2, remain incompletely understood. Here, we report that p70S6 kinase phosphorylates the {micro}2 subunit of the AP2 and that the phosphorylation of serine 45 (S45) depends on p70S6K activity. Loss of S45-{micro}2 phosphorylation results in decreased internalization of canonical CME cargo such as transferrin and PDGF receptors. In Caenorhabditis elegans, lack of S45-{micro}2 phosphorylation produces directionally similar but markedly weaker phenotypes than AP2 loss of function. Live imaging and in silico dynamic modelling suggest that S45-2 phosphorylation has impact on the conformational changes of the AP2 complex. These findings identify a p70S6K-dependent mechanism that modulates AP2 function and further strengthen the importance of post-translational regulation in controlling CME. Summary statementClathrin-mediated endocytosis is essential for receptor internalization and cell signaling. This study identifies a novel regulatory mechanism in which p70S6K-mediated phosphorylation of the AP2 subunit 2 at Ser45 modulates AP2 conformational dynamics and facilitates receptor endocytosis.

cell biology↗

bioRxiv2021

Pronounced sequence specificity of the TET enzyme catalytic domain guides its cellular function

TET (ten-eleven translocation) enzymes catalyze the oxidation of 5-methylcytosine bases in DNA, thus driving active and passive DNA demethylation. Here, we report that the catalytic cores of mammalian TET enzymes favor CpGs embedded within bHLH and bZIP transcription factor binding sites, with 250-fold preference in vitro. Crystal structures and molecular dynamics calculations show that sequence preference is caused by intra-substrate interactions and CpG flanking sequence indirectly affecting enzyme conformation. TET sequence preferences are physiologically relevant as they explain the rates of DNA demethylation in TET-rescue experiments in culture and in vivo within the zygote and germline. Most and least favorable TET motifs represent DNA sites that are bound by methylation-sensitive immediate-early transcription factors and OCT4, respectively, illuminating TET function in transcriptional responses and pluripotency support. One-Sentence SummaryThe catalytic domains of the enzymes that facilitate passive and drive active DNA demethylation have intrinsic sequence preferences that target DNA demethylation to bHLH and bZIP transcription factor binding sites.

molecular biology↗

bioRxiv2021

Ectosome effect on endothelial monolayers in hyperglycemic and normoglycemic conditions

Extracellular vesicles, namely those larger ones - Ectosomes (Ect), are thought to be important cell-to-cell communication medium. Ect are considered as a potential therapeutic for type-1 and type-2 diabetes mellitus. Ect can be internalized by endothelial cells and, owing to their cargo, they modulate targeted cell behavior. Under hyperglycemic conditions (HGC), endothelial cells changed their properties and became stiffer and less mobile which causes endothelial dysfunction and abnormalities in micro- and macrovascular systems. The aim of this study was to find whether Ect restore mobility and motility of macrovascular endothelial cells under HGC. Uptake of Ect, cell morphology, cytoskeleton organization and membrane stiffness (by atomic force microscopy) were analyzed after the exposure to isolated Ect. To find which cellular pathways were deregulated by HGC and whether Ect could potentially restore gene expression profile, transcriptome analysis was done. We observed that endothelial cells internalized more Ect under normoglycemic conditions (NGC) then HGC. Hyperglycemic cells (HG) were bigger and showed the stiffer surface with denser actin cytoskeleton in comparison to normoglycemic cells. Number of metabolic pathways was influenced under HGC, especially those related to intracellular transport, metabolism and cellular component organization and Ect did not restore HGC impaired cell signaling. Ectosomes cannot reverse this harmful effect of hyperglycemia in endothelial cells, which can have clinical implication in use Ect as therapeutic target in diabetes treatment.

cell biology↗

bioRxiv2021

Adar-mediated A-to-I editing is required for establishment of embryonic body axes in zebrafish

Adenosine deaminases (ADARs) catalyze the deamination of adenosine to inosine, also known as A-to-I editing, in RNA. Although A-to-I editing occurs widely across animals, and is well studied, new biological roles are still being discovered. Here, we study the role of A-to-I editing in early zebrafish development. We demonstrate that Adar, the zebrafish orthologue of mammalian ADAR1, is essential for establishing the antero-posterior and dorso-ventral axes and patterning. Genome-wide editing discovery revealed pervasive editing in maternal and the earliest zygotic transcripts, the majority of which occurred in the 3-UTR. Interestingly, transcripts implicated in gastrulation as well as dorso-ventral and antero-posterior patterning were found to contain multiple editing sites. Adar knockdown or overexpression affected gene expression and global editing patterns at 12 hpf, but not earlier. Our study established that RNA editing by Adar is necessary for the earliest steps of embryonic patterning along the zebrafish antero-posterior and dorso-ventral axes.

developmental biology↗