Biology subjects

Zarybnicky, T.

Publications and source records attributed to Zarybnicky, T..

3 recordsLinked to original sources

bioRxiv2024

GLE1 dysfunction compromises cellular homeostasis, spatial organization, and peripheral axon branching

The GLE1 protein is an enigmatic factor of RNA processing, associated with multiple developmental disorders including lethal congenital contracture syndrome 1 (LCCS1). Using in vivo genetic engineering to study disturbed GLE1 functions under physiological conditions we demonstrate that inactivation of Gle1 impedes cellular function and organization and causes pre-gastrulation lethality due to defects in adhesion and lineage specification. In contrast, the knock-in mice genocopying LCCS1-associated GLE1FinMajor variant (Gle1PFQ/PFQ) survive prenatal period but die suddenly at mid-adulthood. Gle1PFQ/PFQ mice present irregular count and distribution of spinal motor neurons and impaired development of neural crest-derived tissues as demonstrated by defects in their sympathetic innervation of heart ventricles, paravertebral sympathetic ganglia volume, and adrenal chromaffin cell counts. Unlike previously reported for yeast and HeLa cells, analysis of molecular consequences of GLE1FinMajor variant identified normal poly(A)+ RNA distribution in Gle1PFQ/PFQ cells, which however were impaired in RNA and protein synthesis and simultaneously showed typical signs of cellular senescence. Gle1PFQ/PFQ also induced disturbed stress responses with significant changes in G3BP1-positive stress granule count. Our results show necessity of GLE1 functions for life and indicate that LCCS1 etiology is resultant of pathogenic GLE1FinMajor variant impinging differentiation of neural crest derivatives and leading to complex multiorgan defects. HighlightsO_LITotal inactivation of GLE1 results in disorganization of blastocyst inner cell mass and early embryonic lethality. C_LIO_LIThe Gle1 knock-in (KI) mice, which genocopy the human GLE1FinMajor variant causative for lethal congenital contracture syndrome 1 (LCCS1), die suddenly in mid-adulthood. C_LIO_LINormal poly(A)+ RNA distribution was observed in Gle1 KI cells, but decreased number of G3BP1-positive stress granules were detected in response to stress. C_LIO_LIAbnormal sympathetic innervation of heart ventricles was detected in Gle1 KI mice. C_LIO_LINeural crest-derived tissues represent a new target of GLE1FinMajor and GLE1-related disorders. C_LI

developmental biology↗

bioRxiv2024

Restoration of mitochondrial complex III function in hepatocytes highlights the liver as a key thermogenic organ independent of brown adipocyte activation

Liver is the key hub of systemic energy metabolism and growth regulation, yet its roles in mitochondrial disease pathophysiology remain relatively understudied. Bcs1lp.S78G knock-in mice, carrying a patient mutation causing respiratory complex III (CIII)-deficiency, present juvenile-onset liver and kidney disease, growth restriction, lipodystrophy, and early death. We restored CIII function in the hepatocytes of these mice using recombinant adeno-associated viral vectors (rAAVs) expressing wild-type Bcs1l. A single intraperitoneal injection of rAAVs into presymptomatic juvenile mice prevented liver disease, improved hypoglycemia and growth, normalized hepatic fuel utilization, and doubled the survival. The mutant mice showed hypothermia and brown adipose tissue (BAT) inflammation, and lacked BAT activation basally and upon acute cold challenge. Disrupted foot pad innervation suggested sensory neuropathy and impaired thermosensation as a contributor to the BAT inactivity. Surprisingly, the rAAV-treated mice maintained near-normal body temperature without significant effect on BAT. Increasing cellular respiration via transgenic alternative oxidase (AOX) was sufficient to prevent the hypothermia. The CIII-deficient mice did not reach euthermia until at an ambient temperature of 35{degrees}C, housing at which relieved metabolic stress and ameliorated hepatocyte senescence. We conclude that mitochondrial respiration in hepatocytes is essential for euthermia in mice. Our findings highlight the crucial role of the liver in thermoregulation, hypothermia as a consequence of mitochondrial dysfunction, and the therapeutic potential of rAAV-based gene delivery in a preclinical model of a multiorgan mitochondrial disease. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=150 HEIGHT=200 SRC="FIGDIR/small/612616v2_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@1cc59ecorg.highwire.dtl.DTLVardef@1239b01org.highwire.dtl.DTLVardef@95697eorg.highwire.dtl.DTLVardef@140fa94_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2024

Biomechanical regulation of cell shapes promotes branching morphogenesis of the ureteric bud epithelium

BackgroundBranching morphogenesis orchestrates organogenesis in many tissues including kidney, where ureteric bud branching determines kidney size and nephron number. Defects in branching morphogenesis result in congenital renal anomalies which manifest as deviations in size, function, and nephron number thus critically compromising the lifelong renal functional capacity established during development. Advances in the genetic and molecular understanding of ureteric bud branching regulation have proved insufficient to improve prognosis of congenital renal defects. Thus, we addressed mechanisms regulating three-dimensional (3D) ureteric bud epithelial cell morphology and cell shape changes during novel branch initiation to uncover the contributions of cellular mechanics on cellular functions and tissue organization in normal and branching-compromised bud tips. MethodsWe explored epithelial cell behavior at all scales by utilizing a combination of mouse genetics and a custom machine-learning segmentation pipeline in MATLAB. Ureteric bud epithelial cell shapes and sizes were quantified in 3D wholemount kidneys. A combination with live imaging of fluorescently labelled UB cells, traction force microscopy, and primary UB cells were used to determine how basic cellular features and niche biomechanics contribute to complex novel branch point determination in the process that aims at gaining optimal growth and epithelial density in a limited space. ResultsMachine learning-based segmentation of tip epithelia identified geometrical round-to-elliptical transformation as a key cell shape change facilitating shifts in growth direction that enable propitious branching complexity. Cell shape and molecular analyses in branching-compromised epithelia demonstrated a failure to condense cell size and conformation. Analysis of branching-compromised ureteric bud derived epithelial cells demonstrated disrupted E-CADHERIN and PAXILLIN mediated adhesive forces and defective cytoskeletal dynamics as detected by fluorescent labelling of actin in primary ureteric bud epithelial cells. Branching-compromised ureteric bud epithelial cells showed wrinkled nuclear shapes and alterations in MYH9-based microtubule organization, which suggest a stiff cellular niche with disturbed sensing of and response to biomechanical cues. ConclusionsOur results indicate that the adhesive forces within the epithelium and towards the niche composed of nephron progenitors must dynamically fluctuate to allow complexity in arborization during new branch formation. The data collectively propose a model where epithelial cell crowding in tandem with stretching transforms individual cells into elliptical and elongated shapes. This creates local curvatures that drive new branch formation during the ampulla-to-asymmetric ampulla transition of ureteric bud.

developmental biology↗