Differentiation Events Research Articles

BRIE2: computational identification of splicing phenotypes from single-cell transcriptomic experiments

RNA splicing is an important driver of heterogeneity in single cells through the expression of alternative transcripts and as a determinant of transcriptional kinetics. However, the intrinsic coverage limitations of scRNA-seq technologies make it challenging to associate specific splicing events to cell-level phenotypes. BRIE2 is a scalable computational method that resolves these issues by regressing single-cell transcriptomic data against cell-level features. We show that BRIE2 effectively identifies differential disease-associated alternative splicing events and allows a principled selection of genes that capture heterogeneity in transcriptional kinetics and improve RNA velocity analyses, enabling the identification of splicing phenotypes associated with biological changes.

Early signalling processes in roots play a crucial role in the differential salt tolerance in contrasting Chenopodium quinoa accessions.

Significant variation in epidermal bladder cell (EBC) density and salt tolerance (ST) exists amongst quinoa accessions, suggesting that salt sequestration in EBCs is not the only mechanism conferring ST in this halophyte. In order to reveal other traits that may operate in tandem with salt sequestration in EBCs and whether these additional tolerance mechanisms acted mainly at the root or shoot level, two quinoa (Chenopodium quinoa) accessions with contrasting ST and EBC densities (Q30, low ST with high EBC density versus Q68, with high ST and low EBC density) were studied. The results indicate that responses in roots, rather than in shoots, contributed to the greater ST in the accession with low EBC density. In particular, the tolerant accession had improved root plasma membrane integrity and K+ retention in the mature root zone in response to salt. Furthermore, superior ST in the tolerant Q68 was associated with faster and root-specific H2O2 accumulation and reactive oxygen species-induced K+ and Ca2+ fluxes in the root apex within 30 min after NaCl application. This was found to be associated with the constitutive up-regulation of the membrane-localized receptor kinases regulatory protein FERONIA in the tolerant accession. Taken together, this study shows that differential root signalling events upon salt exposure are essential for the halophytic quinoa; the failure to do this limits quinoa adaptation to salinity, independently of salt sequestration in EBCs.

The endogenous mex-3 3´UTR is required for germline repression and contributes to optimal fecundity in C. elegans.

RNA regulation is essential to successful reproduction. Messenger RNAs delivered from parent to progeny govern early embryonic development. RNA-binding proteins (RBPs) are the key effectors of this process, regulating the translation and stability of parental transcripts to control cell fate specification events prior to zygotic gene activation. The KH-domain RBP MEX-3 is conserved from nematode to human. It was first discovered in Caenorhabditis elegans, where it is essential for anterior cell fate and embryo viability. Here, we show that loss of the endogenous mex-3 3´UTR disrupts its germline expression pattern. An allelic series of 3´UTR deletion variants identify repressing regions of the UTR and demonstrate that repression is not precisely coupled to reproductive success. We also show that several RBPs regulate mex-3 mRNA through its 3´UTR to define its unique germline spatiotemporal expression pattern. Additionally, we find that both poly(A) tail length control and the translation initiation factor IFE-3 contribute to its expression pattern. Together, our results establish the importance of the mex-3 3´UTR to reproductive health and its expression in the germline. Our results suggest that additional mechanisms control MEX-3 function when 3´UTR regulation is compromised.

Cell type diversity in scallop adductor muscles revealed by single-cell RNA-Seq

Studies on cell atlas in marine invertebrates provide a better understanding of cell types, stem cell maintenance, and lineages of cell differentiation. To investigate the molecular features of various cell types in molluscan muscles, we performed single-cell RNA sequencing (scRNA-seq) to map cell types in scallop adductor muscles. We uncovered the cell type-specific features of 20 cell clusters defined by the expression of multiple specific molecular markers. These cell clusters are mainly classified into four broad classes, including mesenchymal stem cells, muscle cells, neurons, and haemolymph cells. In particular, we identified a diverse repertoire of neurons in the striated adductor muscle, but not in the smooth muscle. We further reconstructed the cell differentiation events using all the cell clusters by single-cell pseudotemporal trajectories. By integrating dual BrdU-PCNA immunodetection, neuron-specific staining and electron microscopy observation, we showed the spatial distribution of mesenchymal stem cells and neurons in striated adductor muscle of scallops. The present findings will not only be useful to address the cell type-specific gene expression profiles in scallop muscles, but also provide valuable resources for cross-species comparison of marine organisms.

Biomaterials and Adipose-Derived Mesenchymal Stem Cells for Regenerative Medicine: A Systematic Review.

The use of biological templates for the suitable growth of adipose-derived mesenchymal stem cells (AD-MSC) and “neo-tissue” construction has exponentially increased over the last years. The bioengineered scaffolds still have a prominent and biocompatible framework playing a role in tissue regeneration. In order to supply AD-MSCs, biomaterials, as the stem cell niche, are more often supplemented by or stimulate molecular signals that allow differentiation events into several strains, besides their secretion of cytokines and effects of immunomodulation. This systematic review aims to highlight the details of the integration of several types of biomaterials used in association with AD-MSCs, collecting notorious and basic data of in vitro and in vivo assays, taking into account the relevance of the interference of the cell lineage origin and handling cell line protocols for both the replacement and repairing of damaged tissues or organs in clinical application. Our group analyzed the quality and results of the 98 articles selected from PubMed, Scopus and Web of Science. A total of 97% of the articles retrieved demonstrated the potential in clinical applications. The synthetic polymers were the most used biomaterials associated with AD-MSCs and almost half of the selected articles were applied on bone regeneration.

Transcriptome-Wide Off-Target Effects of Steric-Blocking Oligonucleotides.

Steric-blocking oligonucleotides (SBOs) are short, single-stranded nucleic acids designed to modulate gene expression by binding to RNA transcripts and blocking access from cellular machinery such as splicing factors. SBOs have the potential to bind to near-complementary sites in the transcriptome, causing off-target effects. In this study, we used RNA-seq to evaluate the off-target differential splicing events of 81 SBOs and differential expression events of 46 SBOs. Our results suggest that differential splicing events are predominantly hybridization driven, whereas differential expression events are more common and driven by other mechanisms (including spurious experimental variation). We further evaluated the performance of in silico screens for off-target splicing events, and found an edit distance cutoff of three to result in a sensitivity of 14% and false discovery rate (FDR) of 99%. A machine learning model incorporating splicing predictions substantially improved the ability to prioritize low edit distance hits, increasing sensitivity from 4% to 26% at a fixed FDR of 90%. Despite these large improvements in performance, this approach does not detect the majority of events at an FDR <99%. Our results suggest that in silico methods are currently of limited use for predicting the off-target effects of SBOs, and experimental screening by RNA-seq should be the preferred approach.

Shallow population structure of southern bluefin tuna, Thunnus maccoyii (Castelnau, 1872) between Indian and Atlantic Oceans inferred from mitochondrial DNA sequences

Southern bluefin tuna (Thunnus maccoyii Castelnau, 1872) is distributed across most of the southern temperate ocean and migrates extensively between 30°S and 50°S. Since T. maccoyii has been continually and heavily exploited, it is necessary to investigate the genetic diversity, population structure and demographic history of T. maccoyii for effective management and conservation. Thirty-seven gonad tissues of T. maccoyii were sampled from two locations, which were in the eastern Indian Ocean and the eastern Atlantic Ocean, by scientific observers onboard Korean T. maccoyii longline vessels in 2015. We compared 1240-bp sequences of combined mitochondrial DNA (mtDNA) from the cytochrome c oxidase subunit I (COI, 504-bp) and control region (CR, 736-bp) sequences. The pairwise fixation index (F ST) and maximum-likelihood tree showed that two clades (A and B) were formed regardless of locations. Clade A occurred more commonly than clade B in both localities: the occurrence ratio of clade A was 69% in the Indian Ocean, and 79% in the Atlantic Ocean, respectively. Our findings suggest that a historic differentiation event may have occurred in T. maccoyii, but recently the connectivity between the two oceans may be possible in T. maccoyii populations.

Peri-endothelial cells, the versatile players in the alternating circular events of vascular growth and differentiation, a switch guided by lycat and cloche, fine tuning angiogenic responsiveness beyond linear hypoxia stimulation

Peri-endothelial cells, the versatile players in the alternating circular events of vascular growth and differentiation, a switch guided by lycat and cloche, fine tuning angiogenic responsiveness beyond linear hypoxia stimulation

Differential Dermatologic Adverse Events Associated With Checkpoint Inhibitor Monotherapy and Combination Therapy: A Meta-Analysis of Randomized Control Trials.

Background: As immune checkpoint inhibitors (ICIs) transition to the forefront of cancer treatment, a better understanding of immune related adverse events (IRAEs) is essential to promote safe clinical practice. Dermatologic adverse events are the most common IRAEs and can lead to drug withdrawal and decreased quality of life. This meta-analysis aimed to investigate the risk of the most prevalent dermatologic adverse events (pruritus and rash) among various ICI treatment regimens.Methods: A systematic search of electronic databases was performed to identify qualified randomized controlled trials (RCTs). Data for any grade and high grade pruritus and rash were extracted for meta-analysis. Two reviewers independently assessed methodological quality. The relative risk summary and 95% confidence interval were calculated.Results: 50 RCTs involving 29941 patients were analyzed. The risk of pruritus (2.15 and 4.21 relative risk respectively) and rash (1.61 and 3.89 relative risk respectively) developing from CTLA-4 or PD-1/-L1 inhibitor were increased compared to placebo, but this effect was not dose-dependent. PD-1/-L1 plus CTLA-4 inhibitor was associated with increased risk of pruritus (1.76 and 0.98 relative risk respectively) and rash (1.72 and 1.37 relative risk respectively) compared to either monotherapy. Compared with CTLA-4 inhibitor, PD-1/-L1 inhibitor had a significantly decreased risk of pruritus and rash in both monotherapy and combination therapy (0.65 and 0.29 relative risk respectively). No significant difference was found between PD-1/-L1 inhibitor combined with chemotherapy and PD-1/-L1 monotherapy in any grade and high grade rash (0.84 and 1.43 relative risk respectively). In subgroup analyses, PD-1 inhibitor was associated with reduced risk of pruritus and rash compared to PD-L1 inhibitor.Conclusion: Our meta-analysis demonstrates a better safety profile for PD-1/-L1 inhibitor compared to CTLA-4 inhibitor in terms of pruritus and rash among both monotherapy and multiple combination therapies. PD-L1 inhibitor may contribute to an increased risk of pruritus and rash compared to PD-1 inhibitor.

Generation of Trophoblast-Like Cells From Hypomethylated Porcine Adult Dermal Fibroblasts.

The first differentiation event in mammalian embryos is the formation of the trophectoderm, which is the progenitor of the outer epithelial components of the placenta, and which supports the fetus during the intrauterine life. However, the epigenetic and paracrine controls at work in trophectoderm differentiation are still to be fully elucidated and the creation of dedicated in vitro models is desirable to increase our understanding. Here we propose a novel approach based on the epigenetic conversion of adult dermal fibroblasts into trophoblast-like cells. The method combines the use of epigenetic erasing with an ad hoc differentiation protocol. Dermal fibroblasts are erased with 5-azacytidine (5-aza-CR) that confers cells a transient high plasticity state. They are then readdressed toward the trophoblast (TR) phenotype, using MEF conditioned medium, supplemented with bone morphogenetic protein 4 (BMP4) and inhibitors of the Activin/Nodal and FGF2 signaling pathways in low O2 conditions. The method here described allows the generation of TR-like cells from easily accessible material, such as dermal fibroblasts, that are very simply propagated in vitro. Furthermore, the strategy proposed is free of genetic modifications that make cells prone to instability and transformation. The TR model obtained may also find useful application in order to better characterize embryo implantation mechanisms and developmental disorders based on TR defects.

Identification of Biomarkers Controlling Cell Fate In Blood Cell Development.

A blood cell lineage consists of several consecutive developmental stages starting from the pluri- or multipotent stem cell to a state of terminal differentiation. Despite their importance for human biology, the regulatory pathways and gene networks that govern these differentiation processes are not yet fully understood. This is in part due to challenges associated with delineating the interactions between transcription factors (TFs) and their corresponding target genes. A possible step forward in this case is provided by the increasing amount of expression data, as a basis for linking differentiation stages and gene activities. Here, we present a novel hierarchical approach to identify characteristic expression peak patterns that global regulators excert along the differentiation path of cell lineages. Based on such simple patterns, we identified cell state-specific marker genes and extracted TFs that likely drive their differentiation. Integration of the mean expression values of stage-specific “key player” genes yielded a distinct peaking pattern for each lineage that was used to identify further genes in the dataset which behave similarly. Incorporating the set of TFs that regulate these genes led to a set of stage-specific regulators that control the biological process of cell fate. As proof of concept, we considered two expression datasets covering key differentiation events in blood cell formation of mice.

Sex Maintenance in Mammals.

The crucial event in mammalian sexual differentiation occurs at the embryonic stage of sex determination, when the bipotential gonads differentiate as either testes or ovaries, according to the sex chromosome constitution of the embryo, XY or XX, respectively. Once differentiated, testes produce sexual hormones that induce the subsequent differentiation of the male reproductive tract. On the other hand, the lack of masculinizing hormones in XX embryos permits the formation of the female reproductive tract. It was long assumed that once the gonad is differentiated, this developmental decision is irreversible. However, several findings in the last decade have shown that this is not the case and that a continuous sex maintenance is needed. Deletion of Foxl2 in the adult ovary lead to ovary-to-testis transdifferentiation and deletion of either Dmrt1 or Sox9/Sox8 in the adult testis induces the opposite process. In both cases, mutant gonads were genetically reprogrammed, showing that both the male program in ovaries and the female program in testes must be actively repressed throughout the individual’s life. In addition to these transcription factors, other genes and molecular pathways have also been shown to be involved in this antagonism. The aim of this review is to provide an overview of the genetic basis of sex maintenance once the gonad is already differentiated.

The Role of TCA Cycle Compounds in Erythroid Heme Synthesis and Its Regulation During an Inflammatory Response

One early hallmark event of terminal erythroid differentiation is the induction of heme and globin synthesis. Onset of heme synthesis occurs when the first pathway enzyme, erythroid-specific 5-aminolevulinate synthase (ALAS2), is induced. The steps of heme biosynthesis are identical between non-erythroid (housekeeping) and differentiating erythroid cells, but two genes encode the first enzyme, 5-aminolevulinate synthase (ALAS). The housekeeping gene (ALAS1) is expressed in non-erythroid cells while the erythroid-specific gene (ALAS2) is expressed only in the erythron. Heme synthesis in metazoan cells requires iron, glycine, and succinyl CoA. The supply of iron required for heme synthesis has been well studied and a reasonable view of the components involved in this process exists. Glycine is abundant in plasma (~250 µM) and studies of glycine transporters are consistent with glycine being supplied from extracellular sources. For succinyl CoA the general assumption is that it is produced by the TCA cycle. This may be possible for housekeeping heme synthesis where only modest levels of heme are made at a given time, but this mechanism would clearly place cells under strain during erythroid differentiation when cells make ~109 molecules of heme in a span of 2-3 days.ALAS2, but not ALAS1, physically interacts with the succinyl CoA synthetase (SCS) ATP-dependent subunit SUCLA2. Since ALAS utilizes succinyl CoA to synthesize aminolevulinic acid (ALA), and because SUCLA2 as a subunit of SCS may be involved in the ATP-dependent reverse reaction of SCS to generate succinyl CoA from succinate, one popular hypothesis is that the ALAS2-SUCLA2 interaction exists to provide succinyl CoA for ALAS2. While the evidence for an association between ALAS2 and SUCLA2 is strong, there is no experimental data showing that SUCLA2 has an impact on ALAS2 activity or that there is a role for this interaction in the supply of succinyl CoA for ALAS2.In the current work we have sought to determine if succinyl CoA for heme synthesis during erythropoiesis in MEL cells originates from the TCA cycle or more directly from metabolism of glutamine via transaminases and α-ketoglutarate dehydrogenase (α-KGDH). Employing 13C-labeled glucose, diethyl succinate, or glutamine we determined that succinate is a poor source of carbons for erythroid heme synthesis, providing a maximum of 5 carbons out of a possible 26 (by isotopomer analysis) and labeling only 2.5% of carbons of isolated heme. Glucose contributes up to 18 carbons and a total of 8.3%, and glutamine contributes to 23 carbons and a total of 23%. Enzyme assays of TCA cycle enzymes revealed that during erythropoiesis only aconitase, isocitrate dehydrogenase, and α-KGDH are induced, which is consistent with glutamine, and not succinate, being the major source of carbon for heme.We also examined the impact of itaconate, a compound produced from the TCA cycle in LPS-induced macrophages, on erythroid differentiation. Itaconate is known to inhibit succinate dehydrogenase, and so might be expected to induce heme synthesis if succinyl CoA is being supplied from accumulating succinate via ATP-dependent SCS. However, dimethyl itaconate (a cell permeable form of itaconate) inhibits heme synthesis in differentiating erythroid K562 cells and this inhibition is reversed by ALA. It has long been assumed that SCS has broad specificity and is capable of making itaconyl CoA (Adler J. et al. 1957, JBC 229:865). However, we found that purified human and porcine SCS do not synthesize itaconyl CoA, but that the enzyme succinyl CoA:glutarate CoA transferase (SUGCT) generates itaconyl CoA from itaconate and succinyl CoA. Additionally we demonstrated that itaconyl CoA inhibits purified ALAS2 at µM concentrations. Previously published data along with our studies on K562 cells in the presence of LPS-induced RAW cells suggest that during the inflammatory response macrophages resident in erythroblastic islands produce itaconate as well as cytokines that induce proximal developing red cells to express a plasmalemmal transporter (possibly SLC13A5) required for itaconate import. Intracellular itaconate is then converted to itaconyl CoA via SUGCT, resulting in ALAS2 inhibition and a concomitant decrease in heme synthesis. This process restricts the amount of protoporphyrin synthesized during infection and, thereby, complements mechanisms that restrict the availability of iron to microbial pathogens. DisclosuresNo relevant conflicts of interest to declare.

Impact of Delayed Anticoagulant Initiation in Patients with Sickle Cell Disease and Newly Diagnosed Venous Thromboembolism: A Population-Based Cohort Study

Background:Timely anticoagulant (AC) initiation is critical in patients with newly diagnosed venous thromboembolism (VTE) to avert recurrence and associated mortality. However, data on the impact of anticoagulant timing on clinical outcomes in sickle cell disease (SCD) patients with newly diagnosed VTE are scarce.Objective:To assess the impact of delayed AC initiation on clinical outcomes in SCD patients with newly diagnosed VTE.Methods:We conducted a retrospective matched-cohort study using the 2009-2015 Truven Health MarketScan® databases to identify subjects with newly diagnosed VTE (VTE diagnosis date used as the index date) and SCD based on International Classification of Diseases (ICD) codes for inpatient and outpatient encounters. VTE patients with SCD were 1:5 individually matched to non-SCD patients on sex, age, index date, and VTE type. Timing of AC initiation was defined as: (a) timely initiation (treatment started within 24 hours post index; reference group), (b) delayed initiation (treatment started within 2-7 days post-index) and (c) no treatment (treatment not started within 7 days post-index). Clinical outcomes were defined as: (i) effectiveness endpoint (time to recurrent VTE), (ii) safety endpoint (time to clinically relevant bleeding) and (iii) composite endpoint (first recurrent VTE or clinically relevant bleeding) within 365 days post index.Kaplan-Meier survivor functions were estimated for all three endpoints by anticoagulant initiation patterns and censored for end of continuous enrollment, end of observation period and occurrence of the opposite outcome. Multivariable Cox proportional hazards models were used to compare hazard ratios (HRs) between groups with adjustment for potential confounders.Results:We identified 434 SCD patients and 2170 non-SCD controls with newly diagnosed VTE. The median age was 45 years (range 15-98) and 63.4% were female. Compared to non-SCD patients, timely initiation of AC therapy was significantly lower in those patients with SCD (50.8% vs 32.7%, p<0.0001). Further, in the SCD cohort, 142 (32.7%) patients had timely AC initiation, 55 (12.7%) had delayed AC initiation and 237 (54.6%) had no AC initiated within 7 days post-index. VTE recurrence and/or bleeding occurred in 36/142 (25.4%) patients with timely AC initiation versus 21/55 (38.2%) with delayed initiation and 59/237 (23.2%) with no AC initiation (p=0.1). (Figure 1) In multivariable analyses, delayed AC was associated with higher risk of VTE recurrence compared to those with timely AC initiation (HR [95% CI] = 3.46 [1.12-10.7]) within 90 days post index. Within 180 days post index, results remained suggestive of increased risk of VTE recurrence (HR [95% CI] = 2.85 [0.97-8.41]) and significantly higher risk of clinically relevant bleeding (HR [95% CI] = 2.39 [1.03-5.51]). Delayed anticoagulation was associated with significantly greater risk of the composite outcome compared to those with timely anticoagulant initiation (HR [95% CI] = 2.56 [1.22-5.37] and 2.58 [1.34-4.95]) within 90 and 180 days post index, respectively. We did not find an association in relation to outcomes between those with timely AC initiation and those with no AC initiation within 7 days post index. (Table 1)Conclusions:It is of concern that SCD patients with a newly diagnosed VTE were less likely to receive timely AC initiation. Delayed AC initiation in SCD patients was associated with greater risk of VTE recurrence and clinically significant bleeding at 90 and 180-days post index, respectively. Paradoxically, we found no statistically significant difference in clinical events between those with timely AC initiation and no AC initiation within 7 days post index. This discrepancy may be attributed to provider preferences in terms of risk-benefit guided treatment initiation and differential event reporting in those without a record of AC initiation in our database. Further investigation is warranted to evaluate provider treatment initiation preferences in relation to patient risk and AC treatment patterns vs risk-benefit balance in SCD patients with VTE. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Alternative splicing associated with cancer stemness in kidney renal clear cell carcinoma

BackgroudCancer stemness is associated with metastases in kidney renal clear cell carcinoma (KIRC) and negatively correlates with immune infiltrates. Recent stemness evaluation methods based on the absolute expression have been proposed to reveal the relationship between stemness and cancer. However, we found that existing methods do not perform well in assessing the stemness of KIRC patients, and they overlooked the impact of alternative splicing. Alternative splicing not only progresses during the differentiation of stem cells, but also changes during the acquisition of the stemness features of cancer stem cells. There is an urgent need for a new method to predict KIRC-specific stemness more accurately, so as to provide help in selecting treatment options.MethodsThe corresponding RNA-Seq data were obtained from the The Cancer Genome Atlas (TCGA) data portal. We also downloaded stem cell RNA sequence data from the Progenitor Cell Biology Consortium (PCBC) Synapse Portal. Independent validation sets with large sample size and common clinic pathological characteristics were obtained from the Gene Expression Omnibus (GEO) database. we constructed a KIRC-specific stemness prediction model using an algorithm called one-class logistic regression based on the expression and alternative splicing data to predict stemness indices of KIRC patients, and the model was externally validated. We identify stemness-associated alternative splicing events (SASEs) by analyzing different alternative splicing event between high- and low- stemness groups. Univariate Cox and multivariable logistic regression analysisw as carried out to detect the prognosis-related SASEs respectively. The area under curve (AUC) of receiver operating characteristic (ROC) was performed to evaluate the predictive values of our model.ResultsHere, we constructed a KIRC-specific stemness prediction model with an AUC of 0.968,and to provide a user-friendly interface of our model for KIRC stemness analysis, we have developed KIRC Stemness Calculator and Visualization (KSCV), hosted on the Shiny server, can most easily be accessed via web browser and the url https://jiang-lab.shinyapps.io/kscv/. When applied to 605 KIRC patients, our stemness indices had a higher correlation with the gender, smoking history and metastasis of the patients than the previous stemness indices, and revealed intratumor heterogeneity at the stemness level. We identified 77 novel SASEs by dividing patients into high- and low- stemness groups with significantly different outcome and they had significant correlations with expression of 17 experimentally validated splicing factors. Both univariate and multivariate survival analysis demonstrated that SASEs closely correlated with the overall survival of patients.ConclusionsBasing on the stemness indices, we found that not only immune infiltration but also alternative splicing events showed significant different at the stemness level. More importantly, we highlight the critical role of these differential alternative splicing events in poor prognosis, and we believe in the potential for their further translation into targets for immunotherapy.

Fmrp regulates oligodendrocyte lineage cell specification and differentiation.

Neurodevelopment requires the precise integration of a wide variety of neuronal and glial cell types. During early embryonic development, motor neurons and then oligodendrocyte precursor cells (OPCs) are specified from neural progenitors residing in the periventricular pMN progenitor domain of the spinal cord. Following gliogenesis, OPCs can differentiate as oligodendrocytes (OLs)-the myelinating glial cells of the central nervous system-or remain as OPCs. To generate unique cell types capable of highly divergent functions, these specification and differentiation events require specialized gene expression programs. RNA binding proteins (RBPs) regulate mRNA localization and translation in the developing nervous system and are linked to many neurodevelopmental disorders. One example is Fragile X syndrome (FXS), caused by the loss of the RBP fragile X mental retardation protein (FMRP). Importantly, infants with FXS have reduced white matter and we previously showed that zebrafish Fmrp is autonomously required in OLs to promote myelin sheath growth. We now find that Fmrp regulates cell specification in pMN progenitor cells such that fmr1 mutant zebrafish generate fewer motor neurons and excess OPCs. Fmrp subsequently promotes differentiation of OPCs, leading to fewer differentiating OLs in the developing spinal cord of fmr1 larvae. Although the early patterning of spinal progenitor domains appears largely normal in fmr1 mutants during early embryogenesis, Shh signaling is greatly diminished. Taken together, these results suggest cell stage-specific requirements for Fmrp in the specification and differentiation of oligodendrocyte lineage cells.

Differential Epigenetic Signature of Corticospinal Motor Neurons in ALS

Corticospinal motor neurons (CSMN) are an indispensable neuron population for the motor neuron circuitry. They are excitatory projection neurons, which collect information from different regions of the brain and transmit it to spinal cord targets, initiating and controlling motor function. CSMN degeneration is pronounced cellular event in motor neurons diseases, such as amyotrophic lateral sclerosis (ALS). Genetic mutations contribute to only about ten percent of ALS. Thus understanding the involvement of other factors, such as epigenetic controls, is immensely valuable. Here, we investigated epigenomic signature of CSMN that become diseased due to misfolded SOD1 toxicity and TDP-43 pathology, by performing quantitative analysis of 5-methylcytosine (5mC) and 5-hydroxymethycytosine (5hmC) expression profiles during end-stage of the disease in hSOD1G93A, and prpTDP-43A315T mice. Our analysis revealed that expression of 5mC was specifically reduced in CSMN of both hSOD1G93A and prpTDP-43A315T mice. However, 5hmC expression was increased in the CSMN that becomes diseased due to misfolded SOD1 and decreased in CSMN that degenerates due to TDP-43 pathology. These results suggest the presence of a distinct difference between different underlying causes. These differential epigenetic events might modulate the expression profiles of select genes, and ultimately contribute to the different paths that lead to CSMN vulnerability in ALS.

Cell Tracking for Organoids: Lessons From Developmental Biology.

Organoids have emerged as powerful model systems to study organ development and regeneration at the cellular level. Recently developed microscopy techniques that track individual cells through space and time hold great promise to elucidate the organizational principles of organs and organoids. Applied extensively in the past decade to embryo development and 2D cell cultures, cell tracking can reveal the cellular lineage trees, proliferation rates, and their spatial distributions, while fluorescent markers indicate differentiation events and other cellular processes. Here, we review a number of recent studies that exemplify the power of this approach, and illustrate its potential to organoid research. We will discuss promising future routes, and the key technical challenges that need to be overcome to apply cell tracking techniques to organoid biology.

Alternative splicing and gene co-expression network-based analysis of dizygotic twins with autism-spectrum disorder and their parents

Autism spectrum disorder (ASD) is a neurodevelopmental disorder with high heritability, however, understanding the complexity of the underlying genetic basis has proven to be a challenging task. We hypothesized that dissecting the aberrations in alternative splicing (AS) and their effects on expression networks might provide insight. Therefore, we performed AS and co-expression analyses of total RNA isolated from Peripheral Blood Mononuclear Cells (PBMCs) of two pairs of dizygotic (DZ) twins with non-syndromic autism and their parents. We identified 183 differential AS events in 146 genes, seven of them being Simons Foundation Autism Research Initiative (SFARI) Category 1–3 genes, three of which had previously been reported to be alternatively spliced in ASD post-mortem brains. Gene co-expression analysis identified 7 modules with 513 genes, 5 of which were SFARI Category 1 or Category 2 genes. Among differentially AS genes within the modules, ZNF322 and NR4A1 could be potentially interesting targets for further investigations.

Dynamic methylation of histone H3K18 in differentiating Theileria parasites

Lysine methylation on histone tails impacts genome regulation and cell fate determination in many developmental processes. Apicomplexa intracellular parasites cause major diseases and they have developed complex life cycles with fine-tuned differentiation events. Yet, apicomplexa genomes have few transcription factors and little is known about their epigenetic control systems. Tick-borne Theileria apicomplexa species have relatively small, compact genomes and a remarkable ability to transform leucocytes in their bovine hosts. Here we report enriched H3 lysine 18 monomethylation (H3K18me1) on the gene bodies of repressed genes in Theileria macroschizonts. Differentiation to merozoites (merogony) leads to decreased H3K18me1 in parasite nuclei. Pharmacological manipulation of H3K18 acetylation or methylation impacted parasite differentiation and expression of stage-specific genes. Finally, we identify a parasite SET-domain methyltransferase (TaSETup1) that can methylate H3K18 and represses gene expression. Thus, H3K18me1 emerges as an important epigenetic mark which controls gene expression and stage differentiation in Theileria parasites.