Activation of meiotic recombination by nuclear import of the DNA break hotspot-determining complex in fission yeast.

Meiotic recombination forms crossovers important for proper chromosome segregation and offspring viability. This complex process involves many proteins acting at each of the multiple steps of recombination. Recombination initiates by formation of DNA double-strand breaks (DSBs), which in the several species examined occur with high frequency at special sites (DSB hotspots). In Schizosaccharomyces pombe, DSB hotspots are bound with high specificity and strongly activated by linear element (LinE) proteins Rec25, Rec27 and Mug20, which form colocalized nuclear foci with Rec10, essential for all DSB formation and recombination. Here, we test the hypothesis that the nuclear localization signal (NLS) of Rec10 is crucial for coordinated nuclear entry after forming a complex with other LinE proteins. In NLS mutants, all LinE proteins were abundant in the cytoplasm, not the nucleus; DSB formation and recombination were much reduced but not eliminated. Nuclear entry of limited amounts of Rec10, apparently small enough for passive nuclear entry, can account for residual recombination. LinE proteins are related to synaptonemal complex proteins of other species, suggesting that they also share an NLS, not yet identified, and undergo protein complex formation before nuclear entry.This article has an associated First Person interview with Mélody Wintrebert, joint first author of the paper.

Exo1 recruits Cdc5 polo kinase to MutLγ to ensure efficient meiotic crossover formation.

Crossovers generated during the repair of programmed meiotic double-strand breaks must be tightly regulated to promote accurate homolog segregation without deleterious outcomes, such as aneuploidy. The Mlh1-Mlh3 (MutLγ) endonuclease complex is critical for crossover resolution, which involves mechanistically unclear interplay between MutLγ and Exo1 and polo kinase Cdc5. Using budding yeast to gain temporal and genetic traction on crossover regulation, we find that MutLγ constitutively interacts with Exo1. Upon commitment to crossover repair, MutLγ-Exo1 associate with recombination intermediates, followed by direct Cdc5 recruitment that triggers MutLγ crossover activity. We propose that Exo1 serves as a central coordinator in this molecular interplay, providing a defined order of interaction that prevents deleterious, premature activation of crossovers. MutLγ associates at a lower frequency near centromeres, indicating that spatial regulation across chromosomal regions reduces risky crossover events. Our data elucidate the temporal and spatial control surrounding a constitutive, potentially harmful, nuclease. We also reveal a critical, noncatalytic role for Exo1, through noncanonical interaction with polo kinase. These mechanisms regulating meiotic crossovers may be conserved across species.

Evaluation of the Contrast Enhancement Performance of Gadopiclenol for Magnetic Resonance Angiography in Healthy Rabbits and Pigs.

OBJECTIVES: Unexpected accumulations of gadolinium in various organs were reported after the administration of gadolinium-based contrast agents, making desirable to reduce the dose while maintaining equivalent diagnostic performance. The aim of this study was to evaluate the contrast enhancement performance of high relaxivity gadopiclenol compared with gadoterate meglumine in abdominal contrast-enhanced magnetic resonance angiography (CE-MRA). MATERIALS AND METHODS: In a first study in healthy rabbits, axial 3D gradient echo sequences were applied at 4.7 T to study arterial enhancement as a function of gadopiclenol dose (0.025, 0.05, 0.075, and 0.1 mmol Gd/kg) or gadoterate meglumine at 0.1 mmol Gd/kg (n = 5-6/group). The increase in signal-to-noise ratio (ΔSNR) in the aorta at the first pass was measured and compared. In a second, crossover study in 6 healthy pigs, abdominal CE-MRA sequences were acquired at 3 T with gadopiclenol at 0.05 mmol Gd/kg or gadoterate meglumine at 0.1 mmol Gd/kg at a 1-week interval. Quantitatively on the maximum intensity projection (MIP) images, the mean MIP SNR within the aorta of both groups was compared. Qualitatively, a blinded comparison of the angiograms was performed by an experienced radiologist to determine the preferred contrast agent. RESULTS: In the rabbit, ∆SNR is linearly correlated with the gadopiclenol dose (P = 0.0010). Compared with gadoterate meglumine 0.1 mmol Gd/kg, an increase in the ∆SNR is observed after 0.05, 0.075, and 0.1 mmol Gd/kg of gadopiclenol (+63% P = 0.0731, +78% P = 0.0081, and +72% P = 0.0773, respectively), whereas at 0.025 mmol Gd/kg, ∆SNR is in the same range as with gadoterate meglumine 0.1 mmol Gd/kg (+15% P > 0.9999). In pigs, contrast enhancement after gadopiclenol at 0.05 mmol/kg is +22% superior to MIP SNR after gadoterate meglumine at 0.1 mmol Gd/kg (P = 0.3095). Qualitatively, a preference was shown for gadopiclenol images (3/6) over the gadoterate meglumine examinations (1/6), with no preference being shown for the remainder (2/6). CONCLUSIONS: First-pass CE-MRA is feasible with gadopiclenol at 0.05 mmol Gd/kg with at least the same arterial signal enhancement and image quality as gadoterate meglumine at 0.1 mmol Gd/kg.

Hacd2 deficiency in mice leads to an early and lethal mitochondrial disease.

OBJECTIVE: Mitochondria fuel most animal cells with ATP, ensuring proper energetic metabolism of organs. Early and extensive mitochondrial dysfunction often leads to severe disorders through multiorgan failure. Hacd2 gene encodes an enzyme involved in very long chain fatty acid (C ≥ 18) synthesis, yet its roles in vivo remain poorly understood. Since mitochondria function relies on specific properties of their membranes conferred by a particular phospholipid composition, we investigated if Hacd2 gene participates to mitochondrial integrity. METHODS: We generated two mouse models, the first one leading to a partial knockdown of Hacd2 expression and the second one, to a complete knockout of Hacd2 expression. We performed an in-depth analysis of the associated phenotypes, from whole organism to molecular scale. RESULTS: Thanks to these models, we show that Hacd2 displays an early and broad expression, and that its deficiency in mice is lethal. Specifically, partial knockdown of Hacd2 expression leads to death within one to four weeks after birth, from a sudden growth arrest followed by cachexia and lethargy. The total knockout of Hacd2 is even more severe, characterized by embryonic lethality around E9.5 following developmental arrest and pronounced cardiovascular malformations. In-depth mechanistic analysis revealed that Hacd2 deficiency causes altered mitochondrial efficiency and ultrastructure, as well as accumulation of oxidized cardiolipin. CONCLUSIONS: Altogether, these data indicate that the Hacd2 gene is essential for energetic metabolism during embryonic and postnatal development, acting through the control of proper mitochondrial organization and function.

Antitumoral effect of local injection of TLR-9 agonist emulsified in Lipiodol with systemic anti-PD-1 in a murine model of colorectal carcinoma.

Introduction: Local treatments of cancer, including transarterial chemoembolization, could enhance responses to systemic immune checkpoint inhibitors such as anti-PD-1 antibodies. Lipiodol, a radiopaque oil, is widely used for transarterial chemoembolization as a tumor-targeting drug carrier and could be used in emulsion with immunomodulators. This study aimed at evaluating the antitumoral effect of intra-tumoral injection of Lipiodol-immunomodulator emulsions combined with systemic anti-PD-1 therapy in a murine model of colorectal carcinoma. Method: Mice (male BALB/c) with anti-PD-1-resistant subcutaneous CT26 tumors were injected with immunomodulators, emulsified or not with Lipiodol (N=10-12/group). Results: The TLR-9 agonist CpG displayed antitumor effects, while Poly I:C and QS21 did not. The Lipiodol-CpG emulsion appeared to be stable and maintained CpG within tumors for a longer time. Repeated intra-tumoral injections, combined with anti-PD-1, induced responses towards the tumor as well as to a distant metastatic-like nodule. This treatment was associated with an increase in proliferative CD8+ T cells and of IFN-γ expression, a decrease in proliferative regulatory T cells but also, surprisingly, an increase in myeloid derived suppressor cells. Conclusions: Local administration of CpG emulsified with Lipiodol led to an effective antitumoral effect when combined to systemic anti-PD-1 therapy. Lipiodol, apart from its radiopaque properties, is an efficient drug-delivery system. The formulated oil-in-water emulsion allows efficient loading and control release of CpG, which induces favorable immune modifications in this murine tumor model.