Biocompatible fluorescent reporters with spectral properties spanning the entire visible spectrum are indispensable tools for imaging the biochemistry of living cells and organisms in real time. ...Here, we report the engineering of a fluorescent chemogenetic reporter with tunable optical and spectral properties. A collection of fluorogenic chromophores with various electronic properties enables to generate bimolecular fluorescent assemblies that cover the visible spectrum from blue to red using a single protein tag engineered and optimized by directed evolution and rational design. The ability to tune the fluorescence color and properties through simple molecular modulation provides a broad experimental versatility for imaging proteins in live cells, including neurons, and in multicellular organisms, and opens avenues for optimizing Förster resonance energy transfer (FRET) biosensors in live cells. The ability to tune the spectral properties and fluorescence performance enables furthermore to match the specifications and requirements of advanced super-resolution imaging techniques.
Tissue organization in Drosophila is regulated by the core planar cell polarity (PCP) proteins Frizzled, Dishevelled, Prickle, Van Gogh and Flamingo. Core PCP proteins are conserved in mammals and ...function in mammalian tissue organization. Recent studies have identified another group of Drosophila PCP proteins, consisting of the protocadherins Fat and Dachsous (Ds) and the transmembrane protein Four-jointed (Fj). In Drosophila, Fat represses fj transcription, and Ds represses Fat activity in PCP. Here we show that Fat4 is an essential gene that has a key role in vertebrate PCP. Loss of Fat4 disrupts oriented cell divisions and tubule elongation during kidney development, leading to cystic kidney disease. Fat4 genetically interacts with the PCP genes Vangl2 and Fjx1 in cyst formation. In addition, Fat4 represses Fjx1 expression, indicating that Fat signaling is conserved. Together, these data suggest that Fat4 regulates vertebrate PCP and that loss of PCP signaling may underlie some cystic diseases in humans.
Hepatocyte nuclear factor-1beta (HNF-1beta) is a transcription factor required for the expression of several renal cystic genes and whose prenatal deletion leads to polycystic kidney disease (PKD). ...We show here that inactivation of Hnf1b from postnatal day 10 onward does not elicit cystic dilations in tubules after their proliferative morphogenetic elongation is over. Cystogenic resistance is intrinsically linked to the quiescent state of cells. In fact, when Hnf1b deficient quiescent cells are forced to proliferate by an ischemia-reperfusion injury, they give rise to cysts, owing to loss of oriented cell division. Remarkably, in quiescent cells, the transcription of crucial cystogenic target genes is maintained even in the absence of HNF-1beta. However, their expression is lost as soon as cells proliferate and the chromatin of target genes acquires heterochromatin marks. These results unveil a previously undescribed aspect of gene regulation. It is well established that transcription is shut off during the mitotic condensation of chromatin. We propose that transcription factors such as HNF-1beta might be involved in reprogramming gene expression after transcriptional silencing is induced by mitotic chromatin condensation. Notably, HNF-1beta remains associated with the mitotically condensed chromosomal barrels. This association suggests that HNF-1beta is a bookmarking factor that is necessary for reopening the chromatin of target genes after mitotic silencing.
•New lineage analyses support the notion of fate restricted progenitors in the neocortex.•Reversibility of transition from proliferation to neurogenesis in individual progenitors is ...discussed.•Combination of clonal tracing and modelling will unravel the logic of neurogenesis.
The vertebrate central nervous system (CNS) is a fantastically complex organ composed of dozens of cell types within the neural and glial lineages. Its organization is laid down during development, through the localized and sequential production of subsets of neurons with specific identities. The principles and mechanisms that underlie the timely production of adequate classes of cells are only partially understood. Recent advances in molecular profiling describe the developmental trajectories leading to this amazing cellular diversity and provide us with cell atlases of an unprecedented level of precision. Yet, some long-standing questions pertaining to lineage relationships between neural progenitor cells and their differentiated progeny remain unanswered. Here, we discuss questions related to proliferation potential, timing of fate choices and restriction of neuronal output potential of individual CNS progenitors through the lens of lineage relationship. Unlocking methodological barriers will be essential to accurately describe CNS development at a cellular resolution.
Mutations in cystic kidney disease genes represent a major genetic cause of end‐stage renal disease. However, the molecular cascades controlling the expression of these genes are still poorly ...understood. Hepatocyte Nuclear Factor 1β (HNF1β) is a homeoprotein predominantly expressed in renal, pancreatic and hepatic epithelia. We report here that mice with renal‐specific inactivation of HNF1β develop polycystic kidney disease. We show that renal cyst formation is accompanied by a drastic defect in the transcriptional activation of Umod, Pkhd1 and Pkd2 genes, whose mutations are responsible for distinct cystic kidney syndromes. In vivo chromatin immunoprecipitation experiments demonstrated that HNF1β binds to several DNA elements in murine Umod, Pkhd1, Pkd2 and Tg737/Polaris genomic sequences. Our results uncover a direct transcriptional hierarchy between HNF1β and cystic disease genes. Interestingly, most of the identified HNF1β target gene products colocalize to the primary cilium, a crucial organelle that plays an important role in controlling the proliferation of tubular cells. This may explain the increased proliferation of cystic cells in MODY5 patients carrying autosomal dominant mutations in HNF1β.
Congenital nephron number varies widely in the human population and individuals with low nephron number are at risk of developing hypertension and chronic kidney disease. The development of the ...kidney occurs via an orchestrated morphogenetic process where metanephric mesenchyme and ureteric bud reciprocally interact to induce nephron formation. The genetic networks that modulate the extent of this process and set the final nephron number are mostly unknown. Here, we identified a specific isoform of MITF (MITF-A), a bHLH-Zip transcription factor, as a novel regulator of the final nephron number. We showed that overexpression of MITF-A leads to a substantial increase of nephron number and bigger kidneys, whereas Mitfa deficiency results in reduced nephron number. Furthermore, we demonstrated that MITF-A triggers ureteric bud branching, a phenotype that is associated with increased ureteric bud cell proliferation. Molecular studies associated with an in silico analyses revealed that amongst the putative MITF-A targets, Ret was significantly modulated by MITF-A. Consistent with the key role of this network in kidney morphogenesis, Ret heterozygosis prevented the increase of nephron number in mice overexpressing MITF-A. Collectively, these results uncover a novel transcriptional network that controls branching morphogenesis during kidney development and identifies one of the first modifier genes of nephron endowment.
Hepatocyte nuclear factor-1β (HNF-1β) is a Pit-1, Oct-1/2, Unc-86 (POU) homeodomain-containing transcription factor expressed in the kidney, liver, pancreas, and other epithelial organs. Mutations of ...HNF-1β cause maturity-onset diabetes of the young, type 5 (MODY5), which is characterized by early-onset diabetes mellitus and congenital malformations of the kidney, pancreas, and genital tract. Knockout of HNF-1β in the mouse kidney results in cyst formation. However, the signaling pathways and transcriptional programs controlled by HNF-1β are poorly understood. Using genome-wide chromatin immunoprecipitation and DNA microarray (ChIP-chip) and microarray analysis of mRNA expression, we identified SOCS3 (suppressor of cytokine signaling-3) as a previously unrecognized target gene of HNF-1β in the kidney. HNF-1β binds to the SOCS3 promoter and represses SOCS3 transcription. The expression of SOCS3 is increased in HNF-1β knockout mice and in renal epithelial cells expressing dominant-negative mutant HNF-1β. Increased levels of SOCS-3 inhibit HGF-induced tubulogenesis by decreasing phosphorylation of Erk and STAT-3. Conversely, knockdown of SOCS-3 in renal epithelial cells expressing dominant-negative mutant HNF-1β rescues the defect in HGF-induced tubulogenesis by restoring phosphorylation of Erk and STAT-3. Thus, HNF-1β regulates tubulogenesis by controlling the levels of SOCS-3 expression. Manipulating the levels of SOCS-3 may be a useful therapeutic approach for human diseases induced by HNF-1β mutations.
The thermodynamic modelling of the binary Al-Zr system has been carried out using the CALPHAD approach, based on available experimental and theoretical data for phase diagram and thermodynamic ...properties. A set of self-consistent thermodynamic parameters was established. The liquid phase and the terminal fcc_A1(Al), bcc_A2(Zr), hcp_A3(Zr) solid solutions were treated as disordered solutions, using the Redlich-Kister expressions for the excess Gibbs energy, and the intermetallic phases were considered to be line compounds. A satisfactory agreement was achieved between the experimental data and the calculated results.
Unequal centrosome maturation correlates with asymmetric division in multiple cell types. Nevertheless, centrosomal fate determinants have yet to be identified. Here, we show that the Notch pathway ...regulator Mindbomb1 co-localizes asymmetrically with centriolar satellite proteins PCM1 and AZI1 at the daughter centriole in interphase. Remarkably, while PCM1 and AZI1 remain asymmetric during mitosis, Mindbomb1 is associated with either one or both spindle poles. Asymmetric Mindbomb1 correlates with neurogenic divisions and Mindbomb1 is inherited by the prospective neuron. By contrast, in proliferative divisions, a supplementary pool of Mindbomb1 associated with the Golgi apparatus in interphase is released during mitosis and compensates for Mindbomb1 centrosomal asymmetry. Finally, we show that preventing Mindbomb1 centrosomal association induces reciprocal Notch activation between sister cells and promotes symmetric divisions. Thus, we uncover a link between differential centrosome maturation and Notch signaling and reveal an unexpected compensatory mechanism involving the Golgi apparatus in restoring symmetry in proliferative divisions.
•Centriolar satellites constitute a docking point for Mib1 at the daughter centriole•Mib1 is inherited by the prospective neuron following asymmetric divisions•Mib1 asymmetry regulates fate choices through unequal Notch activation in daughter cells•In proliferative divisions, a Golgi apparatus pool of Mib1 compensates for the asymmetry
Tozer et al. report that the Notch regulator Mib1, enriched via centriolar satellites at the daughter centriole, is inherited by the prospective neuron and regulates asymmetric fate choices. In proliferative divisions, a Golgi apparatus pool of Mib1 compensates for this asymmetry.
RuO4 oxide appears much less stable than RuO2(s) in the Ru–O binary system with a melting point close to room temperature and a certain propensity to vaporize or decompose at low temperatures. Ab ...initio simulations in the framework of density functional theory (DFT) on RuO4(s) are performed to analyze the cubic and monoclinic structures and to evaluate the heat capacities at low temperatures. Then, a critical evaluation of thermodynamic data from calorimetry and vapor pressure determinations - was carried out coupled with ab-initio calculations to propose new thermodynamic data: the entropy.
S° (RuO4, s, cubic, 298K) = 132.7 J·K−1mol−1 and formation enthalpy.
ΔfH° (RuO4, s, cubic, 298K) = −252.4 ± 5.5 kJ mol−1.
•DFT calculations on RuO4(s) are performed to analyze cubic and monoclinic structures.•Heat capacities of RuO4(s) are calculated.•A critical evaluation is done from high temperature chemistry determinations.•RuO4(s) entropy and formation enthalpy at 298k are proposed and thermodynamic data for the liquid.
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