Notch signaling in mouse blastocyst development and hatching

Research Areas: Developmental BiologyBackground: Mammalian early embryo development requires a well-orchestrated interplay of cell signaling pathways. Notch is a major regulatory pathway involved in cell-fate determination in embryonic and adult scenarios. However, the role of Notch in embryonic pre-implantation development is controversial. In particular, Notch role on blastocyst development and hatching remains elusive, and a complete picture of the transcription and expression patterns of Notch components during this time-period is not available. Results: This study provided a comprehensive view on the dynamics of individual embryo gene transcription and protein expression patterns of Notch components (receptors Notch1–4; ligands Dll1 and Dll4, Jagged1–2; and effectors Hes1–2), and their relationship with transcription of gene markers of pluripotency and differentiation (Sox2, Oct4, Klf4, Cdx2) during mouse blastocyst development and hatching. Transcription of Notch1–2, Jagged1–2 and Hes1 was highly prevalent and dynamic along stages of development, whereas transcription of Notch3–4, Dll4 and Hes2 had a low prevalence among embryos. Transcription levels of Notch1, Notch2, Jagged2 and Hes1 correlated with each other and with those of pluripotency and differentiation genes. Gene transcription was associated to protein expression, except for Jagged2, where high transcription levels in all embryos were not translated into protein. Presence of Notch signaling activity was confirmed through nuclear NICD and Hes1 detection, and downregulation of Hes1 transcription following canonical signaling blockade with DAPT. In vitro embryo culture supplementation with Jagged1 had no effect on embryo developmental kinetics. In contrast, supplementation with Jagged2 abolished Jagged1 transcription, downregulated Cdx2 transcription and inhibited blastocyst hatching. Notch signaling blockade by DAPT downregulated transcription of Sox2, and retarded embryo hatching. Conclusion: Transcription of Notch genes showed a dynamic pattern along blastocyst development and hatching. Data confirmed Notch signaling activity, and lead to the suggestion that Notch canonical signaling may be operating through Notch1, Notch3, Jagged1 and Hes1. Embryo culture supplementation with Jagged1 and Jagged2 unveiled a possible regulatory effect between Jagged1, Cdx2 and blastocyst hatching. Overall, results indicate that a deregulation in Notch signaling, either by its over or under-activation, affects blastocyst development and hatching.info:eu-repo/semantics/publishedVersio

SPPS: A Sequence-Based Method for Predicting Probability of Protein-Protein Interaction Partners

Background: The molecular network sustained by different types of interactions among proteins is widely manifested as the fundamental driving force of cellular operations. Many biological functions are determined by the crosstalk between proteins rather than by the characteristics of their individual components. Thus, the searches for protein partners in global networks are imperative when attempting to address the principles of biology. Results: We have developed a web-based tool ‘‘Sequence-based Protein Partners Search’ ’ (SPPS) to explore interacting partners of proteins, by searching over a large repertoire of proteins across many species. SPPS provides a database containing more than 60,000 protein sequences with annotations and a protein-partner search engine in two modes (Single Query and Multiple Query). Two interacting proteins of human FBXO6 protein have been found using the service in the study. In addition, users can refine potential protein partner hits by using annotations and possible interactive network in the SPPS web server. Conclusions: SPPS provides a new type of tool to facilitate the identification of direct or indirect protein partners which may guide scientists on the investigation of new signaling pathways. The SPPS server is available to the public a

Increased cortical surface area and gyrification following long-term survival from early monocular enucleation

AbstractPurposeRetinoblastoma is typically diagnosed before 5 years of age and is often treated by enucleation (surgical removal) of the cancerous eye. Here, we sought to characterize morphological changes of the cortex following long-term survival from early monocular enucleation.MethodsNine adults with early right-eye enucleation (≤48 months of age) due to retinoblastoma were compared to 18 binocularly intact controls. Surface area, cortical thickness, and gyrification estimates were obtained from T1 weighted images and group differences were examined.ResultsEarly monocular enucleation was associated with increased surface area and/or gyrification in visual (i.e., V1, inferior temporal), auditory (i.e., supramarginal), and multisensory (i.e., superior temporal, inferior parietal, superior parietal) cortices compared with controls. Visual cortex increases were restricted to the right hemisphere contralateral to the remaining eye, consistent with previous subcortical data showing asymmetrical lateral geniculate nucleus volume following early monocular enucleation.ConclusionsAltered morphological development of visual, auditory, and multisensory regions occurs subsequent to long-time survival from early eye loss

Genetic modifiers ameliorate endocytic and neuromuscular defects in a model of spinal muscular atrophy

© 2020 The Author(s). This article is licensed under a Creative Commons Attribution 4.0 International License (https://creativecommons.org/licenses/by/4.0/), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made.Background: Understanding the genetic modifiers of neurodegenerative diseases can provide insight into the mechanisms underlying these disorders. Here, we examine the relationship between the motor neuron disease spinal muscular atrophy (SMA), which is caused by reduced levels of the survival of motor neuron (SMN) protein, and the actin-bundling protein Plastin 3 (PLS3). Increased PLS3 levels suppress symptoms in a subset of SMA patients and ameliorate defects in SMA disease models, but the functional connection between PLS3 and SMN is poorly understood.Results: We provide immunohistochemical and biochemical evidence for large protein complexes localized in vertebrate motor neuron processes that contain PLS3, SMN and members of the hnRNP F/H family of proteins. Using a Caenorhabditis elegans (C. elegans) SMA model, we determine that overexpression of PLS3 or loss of the C. elegans hnRNP F/H ortholog SYM-2 enhances endocytic function and ameliorates neuromuscular defects caused by decreased SMN-1 levels. Furthermore, either increasing PLS3 or decreasing SYM-2 levels suppresses defects in a C. elegans ALS model.Conclusions: We propose that hnRNP F/H act in the same protein complex as PLS3 and SMN and that the function of this complex is critical for endocytic pathways, suggesting that hnRNP F/H proteins could be potential targets for therapy development.Peer reviewe

Self-oligomerization regulates stability of survival motor neuron protein isoforms by sequestering an SCF^Slmb degron

Spinal muscular atrophy (SMA) is caused by homozygous mutations in human SMN1. Expression of a duplicate gene (SMN2) primarily results in skipping of exon 7 and production of an unstable protein isoform, SMNΔ7. Although SMN2 exon skipping is the principal contributor to SMA severity, mechanisms governing stability of survival motor neuron (SMN) isoforms are poorly understood. We used a Drosophila model system and label-free proteomics to identify the SCFSlmb ubiquitin E3 ligase complex as a novel SMN binding partner. SCFSlmb interacts with a phosphor degron embedded within the human and fruitfly SMN YG-box oligomerization domains. Substitution of a conserved serine (S270A) interferes with SCFSlmb binding and stabilizes SMNΔ7. SMA-causing missense mutations that block multimerization of full-length SMN are also stabilized in the degron mutant background. Overexpression of SMNΔ7S270A, but not wild-type (WT) SMNΔ7, provides a protective effect in SMA model mice and human motor neuron cell culture systems. Our findings support a model wherein the degron is exposed when SMN is monomeric and sequestered when SMN forms higher-order multimers

The clerodane diterpene casearin J induces apoptosis of T-ALL cells through SERCA inhibition, oxidative stress, and interference with Notch1 signaling

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic malignancy that preferentially affects children and adolescents. Over 50% of human T-ALLs possess activating mutations of Notch1. The clerodane diterpene casearin J (CJ) is a natural product that inhibits the sarcoendoplasmatic reticulum calcium ATPase (SERCA) pump and induces cell death in leukemia cells, but the molecular mechanism of cytotoxicity remains poorly understood. Here we show that owing to SERCA pump inhibition, CJ induces depletion of the endoplasmic reticulum calcium pools, oxidative stress, and apoptosis via the intrinsic signaling pathway. Moreover, Notch1 signaling is reduced in T-ALL cells with auto-activating mutations in the HD-domain of Notch1, but not in cells that do not depend on Notch1 signaling. CJ also provoked a slight activation of NF-κB, and consistent with this notion a combined treatment of CJ and the NF-κB inhibitor parthenolide (Pt) led to a remarkable synergistic cell death in T-ALL cells. Altogether, our data support the concept that inhibition of the SERCA pump may be a novel strategy for the treatment of T-ALL with HD-domain-mutant Notch1 receptors and that additional treatment with the NF-κB inhibitor parthenolide may have further therapeutic benefits.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigaciones en Productos Naturales (CIPRONA)UCR::Vicerrectoría de Docencia::Ciencias Básicas::Facultad de Ciencias::Escuela de Químic

Mouse Rif1 is a regulatory subunit of protein phosphatase 1 (PP1)

International audienceRif1 is a conserved protein that plays essential roles in orchestrating DNA replication timing, controlling nuclear architecture, telomere length and DNA repair. However, the relationship between these different roles, as well as the molecular basis of Rif1 function is still unclear. The association of Rif1 with insoluble nuclear lamina has thus far hampered exhaustive characterization of the associated protein complexes. We devised a protocol that overcomes this problem, and were thus able to discover a number of novel Rif1 interactors, involved in chromatin metabolism and phosphorylation. Among them, we focus here on PP1. Data from different systems have suggested that Rif1-PP1 interaction is conserved and has important biological roles. Using mutagenesis, NMR, isothermal calorimetry and surface plasmon resonance we demonstrate that Rif1 is a high-affinity PP1 adaptor, able to out-compete the well-established PP1-inhibitor I2 in vitro. Our conclusions have important implications for understanding Rif1 diverse roles and the relationship between the biological processes controlled by Rif1