Targeting of PI3K/AKT/mTOR pathway to inhibit T cell activation and prevent graft-versus-host disease development

Producción CientíficaBackground: Graft-versus-host disease (GvHD) remains the major obstacle to successful allogeneic hematopoietic stem cell transplantation, despite of the immunosuppressive regimens administered to control T cell alloreactivity. PI3K/AKT/mTOR pathway is crucial in T cell activation and function and, therefore, represents an attractive therapeutic target to prevent GvHD development. Recently, numerous PI3K inhibitors have been developed for cancer therapy. However, few studies have explored their immunosuppressive effect. Methods: The effects of a selective PI3K inhibitor (BKM120) and a dual PI3K/mTOR inhibitor (BEZ235) on human T cell proliferation, expression of activation-related molecules, and phosphorylation of PI3K/AKT/mTOR pathway proteins were analyzed. Besides, the ability of BEZ235 to prevent GvHD development in mice was evaluated. Results: Simultaneous inhibition of PI3K and mTOR was efficient at lower concentrations than PI3K specific targeting. Importantly, BEZ235 prevented naïve T cell activation and induced tolerance of alloreactive T cells, while maintaining an adequate response against cytomegalovirus, more efficiently than BKM120. Finally, BEZ235 treatment significantly improved the survival and decreased the GvHD development in mice. Conclusions: These results support the use of PI3K inhibitors to control T cell responses and show the potential utility of the dual PI3K/mTOR inhibitor BEZ235 in GvHD prophylaxis.Asociación Española Contra el Cáncer (Proyecto AIOA110296BLAN).Gerencia Regional de Salud de Castilla y León (Proyecto GRS 726/A13

Identification of the Photoreceptor Transcriptional Co-Repressor SAMD11 as Novel Cause of Autosomal Recessive Retinitis Pigmentosa

Retinitis pigmentosa (RP), the most frequent form of inherited retinal dystrophy is characterized by progressive photoreceptor degeneration. Many genes have been implicated in RP development, but several others remain to be identified. Using a combination of homozygosity mapping, whole-exome and targeted next-generation sequencing, we found a novel homozygous nonsense mutation in SAMD11 in five individuals diagnosed with adult-onset RP from two unrelated consanguineous Spanish families. SAMD11 is ortholog to the mouse major retinal SAM domain (mr-s) protein that is implicated in CRX-mediated transcriptional regulation in the retina. Accordingly, protein-protein network analysis revealed a significant interaction of SAMD11 with CRX. Immunoblotting analysis confirmed strong expression of SAMD11 in human retina. Immunolocalization studies revealed SAMD11 was detected in the three nuclear layers of the human retina and interestingly differential expression between cone and rod photoreceptors was observed. Our study strongly implicates SAMD11 as novel cause of RP playing an important role in the pathogenesis of human degeneration of photoreceptors.This work was supported by several grants from the Spanish Centre for Biomedical Network Research on Rare Diseases (CIBERER)(06/07/0036), Instituto de Salud Carlos III (ISCIII, Spanish Ministry of Health)/FEDER, including FIS (PI013/00226) and RETICS (RD09/0076/00101 and RD12/0034/0010), Ministry of Economy and Competitiveness (MINECO), including FEDER (BFU2012-36845), and BIO2011-27069, Conselleria de Educació of the Valencia Community (PROMETEOII/2014/025), Spanish National Organization of the Blind (ONCE) and the Spanish Fighting Blindness Foundation (FUNDALUCE). M.C. was sponsored by the Miguel Servet Program for Researchers in the Spanish National Health Service (CP12/03256) and RSA by Sara Borrel Postdoctoral Program (CD12/00676), both from the ISCIII/FEDER. A.A-F. was sponsored by CIBERER, RPC is supported by Fundación Conchita Rábago (FCR), L.C is sponsored by RETICS (RD12/0034/0010) from ISCIII and L.d.S. was supported by CAPES Foundation, Ministry of Education of Brazil

Frequent and Efficient Use of the Sister Chromatid for DNA Double-Strand Break Repair during Budding Yeast Meiosis

Studies of DNA double-strand break repair during meiosis reveal that a substantial fraction of recombination occurs between sister chromatids

Pch2 Links Chromosome Axis Remodeling at Future Crossover Sites and Crossover Distribution during Yeast Meiosis

Segregation of homologous chromosomes during meiosis I depends on appropriately positioned crossovers/chiasmata. Crossover assurance ensures at least one crossover per homolog pair, while interference reduces double crossovers. Here, we have investigated the interplay between chromosome axis morphogenesis and non-random crossover placement. We demonstrate that chromosome axes are structurally modified at future crossover sites as indicated by correspondence between crossover designation marker Zip3 and domains enriched for axis ensemble Hop1/Red1. This association is first detected at the zygotene stage, persists until double Holliday junction resolution, and is controlled by the conserved AAA+ ATPase Pch2. Pch2 further mediates crossover interference, although it is dispensable for crossover formation at normal levels. Thus, interference appears to be superimposed on underlying mechanisms of crossover formation. When recombination-initiating DSBs are reduced, Pch2 is also required for viable spore formation, consistent with further functions in chiasma formation. pch2Δ mutant defects in crossover interference and spore viability at reduced DSB levels are oppositely modulated by temperature, suggesting contributions of two separable pathways to crossover control. Roles of Pch2 in controlling both chromosome axis morphogenesis and crossover placement suggest linkage between these processes. Pch2 is proposed to reorganize chromosome axes into a tiling array of long-range crossover control modules, resulting in chiasma formation at minimum levels and with maximum spacing

Guidelines for the use and interpretation of assays for monitoring autophagy (3rd edition)

In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. For example, a key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process versus those that measure fl ux through the autophagy pathway (i.e., the complete process including the amount and rate of cargo sequestered and degraded). In particular, a block in macroautophagy that results in autophagosome accumulation must be differentiated from stimuli that increase autophagic activity, defi ned as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (inmost higher eukaryotes and some protists such as Dictyostelium ) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the fi eld understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. It is worth emphasizing here that lysosomal digestion is a stage of autophagy and evaluating its competence is a crucial part of the evaluation of autophagic flux, or complete autophagy. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. Along these lines, because of the potential for pleiotropic effects due to blocking autophagy through genetic manipulation it is imperative to delete or knock down more than one autophagy-related gene. In addition, some individual Atg proteins, or groups of proteins, are involved in other cellular pathways so not all Atg proteins can be used as a specific marker for an autophagic process. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field

Constraints on dark photon dark matter using data from LIGO's and Virgo's third observing run

We present a search for dark photon dark matter that could couple to gravitational-wave interferometers using data from Advanced LIGO and Virgo's third observing run. To perform this analysis, we use two methods, one based on cross-correlation of the strain channels in the two nearly aligned LIGO detectors, and one that looks for excess power in the strain channels of the LIGO and Virgo detectors. The excess power method optimizes the Fourier Transform coherence time as a function of frequency, to account for the expected signal width due to Doppler modulations. We do not find any evidence of dark photon dark matter with a mass between $m_{\rm A} \sim 10^{-14}-10^{-11}$ eV/$c^2$, which corresponds to frequencies between 10-2000 Hz, and therefore provide upper limits on the square of the minimum coupling of dark photons to baryons, i.e. $U(1)_{\rm B}$ dark matter. For the cross-correlation method, the best median constraint on the squared coupling is $\sim1.31\times10^{-47}$ at $m_{\rm A}\sim4.2\times10^{-13}$ eV/$c^2$; for the other analysis, the best constraint is $\sim 2.4\times 10^{-47}$ at $m_{\rm A}\sim 5.7\times 10^{-13}$ eV/$c^2$. These limits improve upon those obtained in direct dark matter detection experiments by a factor of $\sim100$ for $m_{\rm A}\sim [2-4]\times 10^{-13}$ eV/$c^2$, and are, in absolute terms, the most stringent constraint so far in a large mass range $m_A\sim$ $2\times 10^{-13}-8\times 10^{-12}$ eV/$c^2$.Comment: 20 pages, 7 figure

The population of merging compact binaries inferred using gravitational waves through GWTC-3

We report on the population properties of 76 compact binary mergers detected with gravitational waves below a false alarm rate of 1 per year through GWTC-3. The catalog contains three classes of binary mergers: BBH, BNS, and NSBH mergers. We infer the BNS merger rate to be between 10 $\rm{Gpc^{-3} yr^{-1}}$ and 1700 $\rm{Gpc^{-3} yr^{-1}}$ and the NSBH merger rate to be between 7.8 $\rm{Gpc^{-3}\, yr^{-1}}$ and 140 $\rm{Gpc^{-3} yr^{-1}}$ , assuming a constant rate density versus comoving volume and taking the union of 90% credible intervals for methods used in this work. Accounting for the BBH merger rate to evolve with redshift, we find the BBH merger rate to be between 17.9 $\rm{Gpc^{-3}\, yr^{-1}}$ and 44 $\rm{Gpc^{-3}\, yr^{-1}}$ at a fiducial redshift (z=0.2). We obtain a broad neutron star mass distribution extending from $1.2^{+0.1}_{-0.2} M_\odot$ to $2.0^{+0.3}_{-0.3} M_\odot$. We can confidently identify a rapid decrease in merger rate versus component mass between neutron star-like masses and black-hole-like masses, but there is no evidence that the merger rate increases again before 10 $M_\odot$. We also find the BBH mass distribution has localized over- and under-densities relative to a power law distribution. While we continue to find the mass distribution of a binary's more massive component strongly decreases as a function of primary mass, we observe no evidence of a strongly suppressed merger rate above $\sim 60 M_\odot$. The rate of BBH mergers is observed to increase with redshift at a rate proportional to $(1+z)^{\kappa}$ with $\kappa = 2.9^{+1.7}_{-1.8}$ for $z\lesssim 1$. Observed black hole spins are small, with half of spin magnitudes below $\chi_i \simeq 0.25$. We observe evidence of negative aligned spins in the population, and an increase in spin magnitude for systems with more unequal mass ratio