RESUMEN
Meiotic defects derived from incorrect DNA repair during gametogenesis can lead to mutations, aneuploidies and infertility. The coordinated resolution of meiotic recombination intermediates is required for crossover formation, ultimately necessary for the accurate completion of both rounds of chromosome segregation. Numerous master kinases orchestrate the correct assembly and activity of the repair machinery. Although much less is known, the reversal of phosphorylation events in meiosis must also be key to coordinate the timing and functionality of repair enzymes. Cdc14 is a crucial phosphatase required for the dephosphorylation of multiple CDK1 targets in many eukaryotes. Mutations that inactivate this phosphatase lead to meiotic failure, but until now it was unknown if Cdc14 plays a direct role in meiotic recombination. Here, we show that the elimination of Cdc14 leads to severe defects in the processing and resolution of recombination intermediates, causing a drastic depletion in crossovers when other repair pathways are compromised. We also show that Cdc14 is required for the correct activity and localization of the Holliday Junction resolvase Yen1/GEN1. We reveal that Cdc14 regulates Yen1 activity from meiosis I onwards, and this function is essential for crossover resolution in the absence of other repair pathways. We also demonstrate that Cdc14 and Yen1 are required to safeguard sister chromatid segregation during the second meiotic division, a late action that is independent of the earlier role in crossover formation. Thus, this work uncovers previously undescribed functions of the evolutionary conserved Cdc14 phosphatase in the regulation of meiotic recombination.
Asunto(s)
Proteína Quinasa CDC2/genética , Proteínas de Ciclo Celular/genética , Resolvasas de Unión Holliday/genética , Meiosis/genética , Proteínas Tirosina Fosfatasas/genética , Proteínas de Saccharomyces cerevisiae/genética , Segregación Cromosómica/genética , Intercambio Genético/genética , Reparación del ADN/genética , ADN Cruciforme/genética , Gametogénesis/genética , Recombinación Homóloga/genética , Mutación/genética , Fosforilación/genética , Saccharomyces cerevisiae/genéticaRESUMEN
Phospholipids rich in omega-3 fatty acids from Argentine red shrimp waste were explored as a source to produce food-grade liposomes. Partially purified phospholipids (PL-AS), hexane-soluble (HxSE) and acetone-soluble (Ac-SE) lipid co-extracts, were characterized in terms of extraction yield (2.0%, 1.46% and 4.51%, respectively), chemical composition (fatty acids, tocopherols, sterols, astaxanthin) and thermal stability. Based on lipid fractionation, PL-AS presented 85% phospholipids, while neutral lipids were mostly present in HxSE (75%) and free FA in AcSE (34%), the latter suggesting significant fat hydrolysis. Palmitic, oleic, eicosapentaenoic (EPA) and docosahexaenoic (DHA) acids predominated in the phospholipid fraction of PL-AS, mainly constituted by phosphatidylcholine (PC) (96%). The most abundant phospholipid was identified at m/z 760.59, composed of PC, with C16:0/C18:1 as the most probable FA combination. Unilamellar spherical liposomes were successfully made of PL-AS (≈140 nm, 0.248 PDI, -68.5 mV ζ potential), showing high stability for 28 days at 4 °C.
Asunto(s)
Ácidos Grasos Omega-3 , Liposomas , Acetona , Ácidos Docosahexaenoicos , Ácido Eicosapentaenoico , Ácidos Grasos , FosfolípidosRESUMEN
Liposomes made of partially purified phospholipids (PL) from Argentine red shrimp waste oil were loaded with two antioxidant lipid co-extracts (hexane-soluble, Hx and acetone-soluble, Ac) to provide a higher content of omega-3 fatty acids. The physical properties of the liposomes were characterized by Transmission Electron Microscopy (TEM), Dynamic Light Scattering (DLS) and Differential Scanning Calorimetry (DSC). The antioxidant and anti-inflammatory activity of the lipid extracts and liposomal suspensions were evaluated in terms of Superoxide and ABTS radical scavenging capacities and TNF-α inhibition. Uni-lamellar spherical liposomes (z-average ≈ 145 nm) with strong negative ζ potential (≈ -67 mV) were obtained in all cases. The high content of neutral lipids in the Hx extract caused structural changes in the bilayer membrane and decreased entrapment efficiency regarding astaxanthin and EPA + DHA contents. The liposomes loaded with the Hx/Ac extracts showed higher antioxidant and anti-inflammatory activity compared with empty liposomes. The liposomal dispersions improved the physical and oxidative stability of water-in-oil emulsions as compared with the PL extract, inducing pronounced close packing of water droplets. The liposomes decreased hydroperoxide formation in freshly made emulsions and prevented thio-barbituric acid-reactive substances (TBARS) accumulation during chilled storage. Liposomes from shrimp waste could be valuable nanocarriers and stabilizers in functional food emulsions.