Pediatric neurological disorders are frequently devastating and present unmet needs for effective medicine. The successful treatment of spinal muscular atrophy with splice-switching antisense ...oligonucleotides (SSO) indicates a feasible path to targeting neurological disorders by redirecting pre-mRNA splicing. One direct outcome is the development of SSOs to treat haploinsufficient disorders by targeting naturally occurring non-productive splice isoforms. The development of personalized SSO treatment further inspired the therapeutic exploration of rare diseases. This review will discuss the recent advances that utilize SSOs to treat pediatric neurological disorders.
RNA-binding proteins (RBPs) are essential for gene regulation, but it remains a challenge to identify their RNA targets across cell types. Here we present PIE-Seq to investigate Protein-RNA ...Interaction with dual-deaminase Editing and Sequencing by conjugating C-to-U and A-to-I base editors to RBPs. We benchmark PIE-Seq and demonstrate its sensitivity in single cells, its application in the developing brain, and its scalability with 25 human RBPs. Bulk PIE-Seq identifies canonical binding features for RBPs such as PUM2 and NOVA1, and nominates additional target genes for most tested RBPs such as SRSF1 and TDP-43/TARDBP. Homologous RBPs frequently edit similar sequences and gene sets in PIE-Seq while different RBP families show distinct targets. Single-cell PIE-PUM2 uncovers comparable targets to bulk samples and applying PIE-PUM2 to the developing mouse neocortex identifies neural-progenitor- and neuron-specific target genes such as App. In summary, PIE-Seq provides an orthogonal approach and resource to uncover RBP targets in mice and human cells.
In the mammalian neocortex, projection neuron types are sequentially generated by the same pool of neural progenitors. How neuron type specification is related to developmental timing remains ...unclear. To determine whether temporal gene expression in neural progenitors correlates with neuron type specification, we performed single-cell RNA sequencing (scRNA-Seq) analysis of the developing mouse neocortex. We uncovered neuroepithelial cell enriched genes such as
and
when compared to radial glial cells (RGCs). RGCs display dynamic gene expression over time; for instance, early RGCs express higher levels of
, and late RGCs show higher expression of
Interestingly, intermediate progenitor cell marker gene
coexpresses temporally with known neuronal identity genes at different developmental stages, though mostly in postmitotic cells. Our results delineate neural progenitor cell diversity in the developing mouse neocortex and support that neuronal identity genes are transcriptionally evident in
-positive cells.
Dysregulation of alternative splicing has been repeatedly associated with neurodevelopmental disorders, but the extent of cell-type-specific splicing in human neural development remains largely ...uncharted. Here, single-cell long-read sequencing in induced pluripotent stem cell (iPSC)-derived cerebral organoids identifies over 31,000 uncatalogued isoforms and 4,531 cell-type-specific splicing events. Long reads uncover coordinated splicing and cell-type-specific intron retention events, which are challenging to study with short reads. Retained neuronal introns are enriched in RNA splicing regulators, showing shorter lengths, higher GC contents, and weaker 5′ splice sites. We use this dataset to explore the biological processes underlying neurological disorders, focusing on autism. In comparison with prior transcriptomic data, we find that the splicing program in autistic brains is closer to the progenitor state than differentiated neurons. Furthermore, cell-type-specific exons harbor significantly more de novo mutations in autism probands than in siblings. Overall, these results highlight the importance of cell-type-specific splicing in autism and neuronal gene regulation.
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•Over 31,000 uncatalogued isoforms are identified in human cerebral organoids•4,531 cell-type-specific splicing events are identified in human cerebral organoids•Alternative exons are enriched for autism-associated exons and de novo mutations•142 autism genes display cell-type-specific isoforms
Yang et al. perform single-cell long-read sequencing in the human cerebral organoids, identifying thousands of uncatalogued isoforms, differentially spliced exons, and retained introns. Their results reveal the enrichement of autism-associated exons and de novo mutations in cell-type-specific exons.
Chronic Hepatitis B virus (CHB) infection is a global health challenge, causing damage ranging from hepatitis to cirrhosis and hepatocellular carcinoma. In our study, single-cell RNA sequencing ...(scRNA-seq) analysis was performed in livers from mice models with chronic inflammation induced by CHB infection and we found that endothelial cells (ECs) exhibited the largest number of differentially expressed genes (DEGs) among all ten cell types. NF-κB signaling was activated in ECs to induce cell dysfunction and subsequent hepatic inflammation, which might be mediated by the interaction of macrophage-derived and cholangiocyte-derived VISFATIN/Nampt signaling. Moreover, we divided ECs into three subclusters, including periportal ECs (EC_Z1), midzonal ECs (EC_Z2), and pericentral ECs (EC_Z3) according to hepatic zonation. Functional analysis suggested that pericentral ECs and midzonal ECs, instead of periportal ECs, were more vulnerable to HBV infection, as the VISFATIN/Nampt- NF-κB axis was mainly altered in these two subpopulations. Interestingly, pericentral ECs showed increasing communication with macrophages and cholangiocytes via the Nampt-Insr and Nampt-Itga5/Itgb1 axis upon CHB infection, which contribute to angiogenesis and vascular capillarization. Additionally, ECs, especially pericentral ECs, showed a close connection with nature killer (NK) cells and T cells via the Cxcl6-Cxcr6 axis, which is involved in shaping the microenvironment in CHB mice livers. Thus, our study described the heterogeneity and functional alterations of three subclusters in ECs. We revealed the potential role of VISFATIN/Nampt signaling in modulating ECs characteristics and related hepatic inflammation, and EC-derived chemokine Cxcl16 in shaping NK and T cell recruitment, providing key insights into the multifunctionality of ECs in CHB-associated pathologies.
Environmental stresses and nutrition availability critically affect animal development. Numerous animal species across multiple phyla enter developmental arrest for long-term survival in unfavorable ...environments and resume development upon stress removal. Here we show that compromising overall microRNA (miRNA) functions or mutating certain individual miRNAs impairs the long-term survival of nematodes during starvation-induced L1 diapause. We provide evidence that miRNA miR-71 is not required for the animals’ entry into L1 diapause, but plays a critical role in long-term survival by repressing the expression of insulin receptor/PI3K pathway genes and genes acting downstream or in parallel to the pathway. Furthermore, miR-71 plays a prominent role in developmental recovery from L1 diapause partly through repressing the expression of certain heterochronic genes. The presented results indicate that interactions between multiple miRNAs and likely a large number of their mRNA targets in multiple pathways regulate the response to starvation-induced L1 diapause.
Somatic Mutations in Cerebral Cortical Malformations Jamuar, Saumya S; Lam, Anh-Thu N; Kircher, Martin ...
New England journal of medicine/The New England journal of medicine,
08/2014, Letnik:
371, Številka:
8
Journal Article
Recenzirano
Odprti dostop
Somatic mutations can cause brain malformations but may escape detection if their prevalence in blood is low. The authors of this study used deep-coverage targeting sequencing to gauge the extent to ...which somatic mutations cause relatively common forms of brain malformation.
Somatic mutation, a postzygotic event, leads to two or more populations of cells with distinct genotypes in an organism, despite development from a single fertilized egg.
1
,
2
Although the role of somatic mutation in cancer cells is well established,
3
an analogous role for somatic mutations that occur randomly during the normal mitotic cell divisions of embryonic development — and that are therefore present in clones of cells in one or more tissues of the body — has been recognized only recently. Somatic mutations have been described in several noncancerous disorders, including the McCune–Albright syndrome,
4
the Sturge–Weber syndrome,
5
the Proteus syndrome, . . .
Several laboratory and imaging assays are required to diagnose multiple myeloma (MM). Serum and urine immunofixation electrophoresis are two key assays to diagnose MM, while they have not been ...extensively utilized in Chinese hospitals. Serum light chain (sLC), β2 microglobulin (β2-MG), lactic dehydrogenase (LDH), and immunoglobulin (Ig) are routinely measured in the majority of Chinese hospitals. Imbalance of sLC ratio (involved light chain/uninvolved light chain) is frequently observed in MM patients. This study aimed to evaluate the screening value of sLC ratio, β2-MG, LDH, and Ig in MM patients using receiver operating characteristic (ROC) curves.
Data of 303 suspected MM patients, who were admitted to the Taizhou Central Hospital between March 2015 and July 2021, were retrospectively analyzed. In total, 69 patients (MM arm) met the International Myeloma Working Group (IMWG) updated criteria for the diagnosis of MM, while 234 patients were non-MM (non-MM arm). All patients' sLC, β2-MG, LDH, and Ig were measured using commercially available kits according to the manufacturer's instructions. The ROC curve analysis was employed to assess the screening value of sLC ratio, β2-MG, LDH, creatinine (Cr) and Ig. The statistical analysis was carried out by SPSS 26.0 (IBM, Armonk, NY, USA) and MedCalc 19.0.4 (Ostend, Belgium) software.
There was no significant difference between the MM and non-MM arms in terms of gender, age and Cr. The median sLC ratio in the MM arm was 11.5333, which was significantly higher than that of 1.9293 in the non-MM arm (P<0.001). The area under the curve (AUC) of sLC ratio was 0.875, which indicated a robust screening value. The optimal sensitivity and specificity were 81.16% and 94.87% respectively, when the sLC ratio was set as 3.2121. The serum levels of β2-MG and Ig were higher in the MM arm than those in the non-MM arm (P<0.001). The AUC values of β2-MG, LDH, and Ig were 0.843 (P<0.001), 0.547 (P = 0.2627), and 0.723 (P<0.001), respectively. The optimal cutoff values of β2-MG, LDH, and Ig were 1.95 mg/L, 220 U/L, and 46.4 g/L respectively, in the context of screening value. The triple combination of sLC ratio (3.2121), β2-MG (1.95 mg/L), and Ig (46.4 g/L) yielded a higher screening value compared with that of sLC ratio alone (AUC, 0.952; P<0.0001). The triple combination had a sensitivity of 94.20% and a specificity of 86.75%. The addition of LDH to the triple combination and formation of quadruple combination did not optimize the screening value, with AUC, sensitivity, and specificity of 0.952, 94.20%, and 85.47%, respectively.
The triple combination strategy (sLC ratio, 3.2121; β2-MG, 1.95 mg/L; Ig, 46.4 g/L) is accompanied by remarkable sensitivity and specificity for screening MM in Chinese hospitals.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
How the nuclei in mammalian skeletal muscle fibers properly position themselves relative to the cell body is an interesting and important cell biology question. In the syncytial skeletal muscle ...cells, more than 100 nuclei are evenly distributed at the periphery of each cell, with 3-8 nuclei anchored beneath the neuromuscular junction (NMJ). Our previous studies revealed that the KASH domain-containing Syne-1/Nesprin-1 protein plays an essential role in anchoring both synaptic and nonsynaptic myonuclei in mice. SUN domain-containing proteins (SUN proteins) have been shown to interact with KASH domain-containing proteins (KASH proteins) at the nuclear envelope (NE), but their roles in nuclear positioning in mice are unknown. Here we show that the synaptic nuclear anchorage is partially perturbed in Sun1, but not in Sun2, knockout mice. Disruption of 3 or all 4 Sun1/2 wild-type alleles revealed a gene dosage effect on synaptic nuclear anchorage. The organization of nonsynaptic nuclei is disrupted in Sun1/2 double-knockout (DKO) mice as well. We further show that the localization of Syne-1 to the NE of muscle cells is disrupted in Sun1/2 DKO mice. These results clearly indicate that SUN1 and SUN2 function critically in skeletal muscle cells for Syne-1 localization at the NE, which is essential for proper myonuclear positioning.
Background
Mucosal-associated invariant T (MAIT) cells are considered to participate of the host immune response against acute severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection; ...however, single-cell transcriptomic profiling of MAIT cells in patients with COVID-19 remains unexplored.
Methods
We performed single-cell RNA sequencing analyses on peripheral MAIT cells from 13 patients with COVID-19 and 5 healthy donors. The transcriptional profiles of MAIT cells, together with assembled T-cell receptor sequences, were analyzed. Flow cytometry analysis was also performed to investigate the properties of MAIT cells.
Results
We identified that differentially expressed genes (DEGs) of MAIT cells were involved in myeloid leukocyte activation and lymphocyte activation in patients with COVID-19. In addition, in MAIT cells from severe cases, more DEGs were enriched in adaptive cellular and humoral immune responses compared with those in moderate cases. Further analysis indicated that the increase of cell cytotoxicity (killing), chemotaxis, and apoptosis levels in MAIT cells were consistent with disease severity and displayed the highest levels in patients with severe disease. Interestingly, flow cytometry analysis showed that the frequencies of pyroptotic MAIT cells, but not the frequencies of apoptotic MAIT cells, were increased significantly in patients with COVID-19, suggesting pyroptosis is one of leading causes of MAIT cell deaths during SARS-CoV-2 infection. Importantly, there were more clonal expansions of MAIT cells in severe cases than in moderate cases.
Conclusions
The results of the present study suggest that MAIT cells are likely to be involved in the host immune response against SARS-CoV-2 infection. Simultaneously, the transcriptomic data from MAIT cells provides a deeper understanding of the immune pathogenesis of the disease.
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