Control of ribosomal RNA synthesis by hematopoietic transcription factors.

Ribosomal RNAs (rRNAs) are the most abundant cellular RNAs, and their synthesis from rDNA repeats by RNA polymerase I accounts for the bulk of all transcription. Despite substantial variation in rRNA transcription rates across cell types, little is known about cell-type-specific factors that bind rDNA and regulate rRNA transcription to meet tissue-specific needs. Using hematopoiesis as a model system, we mapped about 2,200 ChIP-seq datasets for 250 transcription factors (TFs) and chromatin proteins to human and mouse rDNA and identified robust binding of multiple TF families to canonical TF motifs on rDNA. Using a 47S-FISH-Flow assay developed for nascent rRNA quantification, we demonstrated that targeted degradation of C/EBP alpha (CEBPA), a critical hematopoietic TF with conserved rDNA binding, caused rapid reduction in rRNA transcription due to reduced RNA Pol I occupancy. Our work identifies numerous potential rRNA regulators and provides a template for dissection of TF roles in rRNA transcription.

PHF6 suppresses self-renewal of leukemic stem cells in AML.

Acute myeloid leukemia is characterized by uncontrolled proliferation of self-renewing myeloid progenitors. PHF6 is a chromatin-binding protein mutated in myeloid leukemias, and its loss increases mouse HSC self-renewal without malignant transformation. We report here that Phf6 knockout increases the aggressiveness of Hoxa9-driven AML over serial transplantation, and increases the frequency of leukemia initiating cells. We define the in vivo hierarchy of Hoxa9-driven AML and identify a population that we term the 'LIC-e' (leukemia initiating cells enriched) population. We find that Phf6 loss has context-specific transcriptional effects, skewing the LIC-e transcriptome to a more stem-like state. We demonstrate that LIC-e accumulation in Phf6 knockout AML occurs not due to effects on cell cycle or apoptosis, but due to an increase in the fraction of its progeny that retain LIC-e identity. Overall, our work indicates that Phf6 loss increases AML self-renewal through context-specific effects on leukemia stem cells.

FISH-Flow to quantify nascent and mature ribosomal RNA in mouse and human cells.

FISH-Flow (fluorescence in situ hybridization-flow cytometry) involves hybridizing fluorescent oligos to RNA and quantifying fluorescence at a single-cell level using flow cytometry. Here, we present a FISH-Flow protocol to quantify nascent 47S and mature 18S and 28S rRNAs in mouse and human cells, including rRNA quantification across cell cycle stages using DNA staining. We describe steps for cell preparation, hybridization of fluorescent probes against rRNA, and DNA staining. We then detail procedures for flow cytometry and data analysis. For complete details on the use and execution of this protocol, please refer to Antony et al. (2022).1.

A Review on Self-Assembly of Colloidal Nanoparticles into Clusters, Patterns, and Films: Emerging Synthesis Techniques and Applications.

The colloidal synthesis of functional nanoparticles has gained tremendous scientific attention in the last decades. In parallel to these advancements, another rapidly growing area is the self-assembly or self-organization of these colloidal nanoparticles. First, the organization of nanoparticles into ordered structures is important for obtaining functional interfaces that extend or even amplify the intrinsic properties of the constituting nanoparticles at a larger scale. The synthesis of large-scale interfaces using complex or intricately designed nanostructures as building blocks, requires highly controllable self-assembly techniques down to the nanoscale. In certain cases, for example, when dealing with plasmonic nanoparticles, the assembly of the nanoparticles further enhances their properties by coupling phenomena. In other cases, the process of self-assembly itself is useful in the final application such as in sensing and drug delivery, amongst others. In view of the growing importance of this field, this review provides a comprehensive overview of the recent developments in the field of nanoparticle self-assembly and their applications. For clarity, the self-assembled nanostructures are classified into two broad categories: finite clusters/patterns, and infinite films. Different state-of-the-art techniques to obtain these nanostructures are discussed in detail, before discussing the applications where the self-assembly significantly enhances the performance of the process.

CdS Nanorods Anchored with Crystalline FeP Nanoparticles for Efficient Photocatalytic Formic Acid Dehydrogenation.

Photocatalytic dehydrogenation of formic acid is a promising strategy for H2 generation. In this work, we report the use of crystalline iron phosphide (FeP) nanoparticles as an efficient and robust cocatalyst on CdS nanorods (FeP@CdS) for highly efficient photocatalytic formic acid dehydrogenation. The optimal H2 evolution rate can reach ∼556 µmol·h-1 at pH 3.5, which is more than 37 times higher than that of bare CdS. Moreover, the photocatalyst demonstrates excellent stability; no significant decrease of the catalytic activity was observed during continuous testing for more than four days. The apparent quantum yield is ∼54% at 420 nm, which is among the highest values obtained using noble-metal-free photocatalysts for formic acid dehydrogenation. This work provides a novel strategy for designing highly efficient and economically viable photocatalysts for formic acid dehydrogenation.

HectD1 controls hematopoietic stem cell regeneration by coordinating ribosome assembly and protein synthesis.

Impaired ribosome function is the underlying etiology in a group of bone marrow failure syndromes called ribosomopathies. However, how ribosomes are regulated remains poorly understood, as are approaches to restore hematopoietic stem cell (HSC) function loss because of defective ribosome biogenesis. Here we reveal a role of the E3 ubiquitin ligase HectD1 in regulating HSC function via ribosome assembly and protein translation. Hectd1-deficient HSCs exhibit a striking defect in transplantation ability and ex vivo maintenance concomitant with reduced protein synthesis and growth rate under stress conditions. Mechanistically, HectD1 ubiquitinates and degrades ZNF622, an assembly factor for the ribosomal 60S subunit. Hectd1 loss leads to accumulation of ZNF622 and the anti-association factor eIF6 on 60S, resulting in 60S/40S joining defects. Importantly, Znf622 depletion in Hectd1-deficient HSCs restored ribosomal subunit joining, protein synthesis, and HSC reconstitution capacity. These findings highlight the importance of ubiquitin-coordinated ribosome assembly in HSC regeneration.

Role of peak current in conversion of patients with ventricular fibrillation.

INTRODUCTION: Peak currents are the final arbiter of defibrillation in patients with ventricular fibrillation (VF). However, biphasic defibrillators continue to use energy in joules for electrical conversion in hopes that their impedance compensation properties will address transthoracic impedance (TTI), which must be overcome when a fixed amount of energy is delivered. However, optimal peak currents for conversion of VF remain unclear. We aimed to determine the role of peak current and optimal peak levels for conversion in collapsed VF patients. METHODS: Adult, non-pregnant patients presenting with non-traumatic VF were included in the study. All defibrillations that occurred were included. Impedance values during defibrillation were used to calculate peak current values. The endpoint was return of spontaneous circulation (ROSC). RESULTS: Of the 197 patients analysed, 105 had ROSC. Characteristics of patients with and without ROSC were comparable. Short duration of collapse < 10 minutes correlated positively with ROSC. Generally, patients with average or high TTI converted at lower peak currents. 25% of patients with high TTI converted at 13.3 ± 2.3 A, 22.7% with average TTI at 18.2 ± 2.5 A and 18.6% with low TTI at 27.0 ± 4.7 A (p = 0.729). Highest peak current conversions were at < 15 A and 15-20 A. Of the 44 patients who achieved first-shock ROSC, 33 (75.0%) received < 20 A peak current vs. > 20 A for the remaining 11 (25%) patients (p = 0.002). CONCLUSION: For best effect, priming biphasic defibrillators to deliver specific peak currents should be considered.

Prompt use of mechanical cardiopulmonary resuscitation in out-of-hospital cardiac arrest: the MECCA study report.

INTRODUCTION: Early use of mechanical cardiopulmonary resuscitation (CPR) for out-of-hospital cardiac arrest (OHCA) may improve survival outcomes. Current evidence for such devices uses outcomes from an intention-to-treat (ITT) perspective. We aimed to determine whether early use of mechanical CPR using a LUCAS 2 device results in better outcomes. METHODS: A prospective, randomised, multicentre study was conducted over one year with LUCAS 2 devices in 14 ambulances and manual CPR in 32 ambulances to manage OHCA. The primary outcome was return of spontaneous circulation (ROSC). Secondary outcomes were survival at 24 hours, discharge from hospital and 30 days. RESULTS: Of the 1,274 patients recruited, 1,191 were eligible for analysis. 889 had manual CPR and 302 had LUCAS CPR. From an ITT perspective, outcomes for manual and LUCAS CPR were: ROSC 29.2% and 31.1% (odds ratio [OR] 1.09, 95% confidence interval [CI] 0.82-1.45; p = 0.537); 24-hour survival 11.2% and 13.2% (OR 1.20, 95% CI 0.81-1.78; p = 0.352); survival to discharge 3.6% and 4.3% (OR 1.20, 95% CI 0.62-2.33; p = 0.579); and 30-day survival 3.0% and 4.0% (OR 1.32, 95% CI 0.66-2.64; p = 0.430), respectively. By as-treated analysis, outcomes for manual, early LUCAS and late LUCAS CPR were: ROSC 28.0%, 36.9% and 24.5%; 24-hour survival 10.6%, 15.5% and 8.2%; survival to discharge 2.9%, 5.8% and 2.0%; and 30-day survival 2.4%, 5.8% and 0.0%, respectively. Adjusted OR for survival with early LUCAS vs. manual CPR was 1.47 after adjustment for other variables (p = 0.026). CONCLUSION: This study showed a survival benefit with LUCAS CPR as compared to manual CPR only, when the device was applied early on-site.