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Background: With the rise of single-cell RNA sequencing new bioinformatic tools have been developed to handle specific demands, such as quantifying unique molecular identifiers and correcting cell barcodes. Here, we benchmarked several datasets with the most common alignment tools for single-cell RNA sequencing data. We evaluated differences in the whitelisting, gene quantification, overall performance, and potential variations in clustering or detection of differentially expressed genes. We compared the tools Cell Ranger version 6, STARsolo, Kallisto, Alevin, and Alevin-fry on 3 published datasets for human and mouse, sequenced with different versions of the 10X sequencing protocol.
Results: Striking differences were observed in the overall runtime of the mappers. Besides that, Kallisto and Alevin showed variances in the number of valid cells and detected genes per cell. Kallisto reported the highest number of cells; however, we observed an overrepresentation of cells with low gene content and unknown cell type. Conversely, Alevin rarely reported such low-content cells. Further variations were detected in the set of expressed genes. While STARsolo, Cell Ranger 6, Alevin-fry, and Alevin produced similar gene sets, Kallisto detected additional genes from the Vmn and Olfr gene family, which are likely mapping artefacts. We also observed differences in the mitochondrial content of the resulting cells when comparing a prefiltered annotation set to the full annotation set that includes pseudogenes and other biotypes.
Conclusion: Overall, this study provides a detailed comparison of common single-cell RNA sequencing mappers and shows their specific properties on 10X Genomics data.
The 14-3-3 protein family are molecular chaperones involved in several biological functions and neurological diseases. We previously pinpointed YWHAZ (encoding 14-3-3ζ) as a candidate gene for autism spectrum disorder (ASD) through a whole-exome sequencing study, which identified a frameshift variant within the gene (c.659-660insT, p.L220Ffs*18). Here, we explored the contribution of the seven human 14-3-3 family members in ASD and other psychiatric disorders by investigating the: (i) functional impact of the 14-3-3ζ mutation p.L220Ffs*18 by assessing solubility, target binding and dimerization; (ii) contribution of common risk variants in 14-3-3 genes to ASD and additional psychiatric disorders; (iii) burden of rare variants in ASD and schizophrenia; and iv) 14-3-3 gene expression using ASD and schizophrenia transcriptomic data. We found that the mutant 14-3-3ζ protein had decreased solubility and lost its ability to form heterodimers and bind to its target tyrosine hydroxylase. Gene-based analyses using publicly available datasets revealed that common variants in YWHAE contribute to schizophrenia (p = 6.6 × 10−7), whereas ultra-rare variants were found enriched in ASD across the 14-3-3 genes (p = 0.017) and in schizophrenia for YWHAZ (meta-p = 0.017). Furthermore, expression of 14-3-3 genes was altered in post-mortem brains of ASD and schizophrenia patients. Our study supports a role for the 14-3-3 family in ASD and schizophrenia.
Social insects dominate arthropod communities worldwide due to cooperation and division of labor in their societies. This, however, makes them vulnerable to exploitation by social parasites, such as slave‐making ants. Slave‐making ant workers pillage brood from neighboring nests of related host ant species. After emergence, host workers take over all nonreproductive colony tasks, whereas slavemakers have lost the ability to care for themselves and their offspring. Here, we compared transcriptomes of different developmental stages (larvae, pupae, and adults), castes (queens and workers), and sexes of two related ant species, the slavemaker Temnothorax americanus and its host Temnothorax longispinosus. Our aim was to investigate commonalities and differences in group‐specific transcriptomes, whereupon across‐species differences possibly can be explained by their divergent lifestyles. Larvae and pupae showed the highest similarity between the two species and upregulated genes with enriched functions of translation and chitin metabolism, respectively. Workers commonly upregulated oxidation‐reduction genes, possibly indicative of their active lifestyle. Host workers, but not workers of the slavemaker, upregulated a “social behavior” gene. In slavemaker queens and workers, genes associated with the regulation of transposable elements were upregulated. Queens of both species showed transcriptomic signals of anti‐aging mechanisms, with hosts upregulating various DNA repair pathways and slavemaker queens investing in trehalose metabolism. The transcriptomes of males showed enriched functions for quite general terms realized in different genes and pathways in each species. In summary, the strong interspecific commonalities in larvae, pupae, and workers were reflected in the same enriched Gene Ontology (GO) terms. Less commonalities occurred in the transcriptomes of queens and males, which apparently utilize different pathways to achieve a long life and sperm production, respectively. We found that all analyzed groups in this study show characteristic GO terms, with similar patterns in both species.
Background: The European beech is arguably the most important climax broad-leaved tree species in Central Europe, widely planted for its valuable wood. Here, we report the 542 Mb draft genome sequence of an up to 300-year-old individual (Bhaga) from an undisturbed stand in the Kellerwald-Edersee National Park in central Germany.
Findings: Using a hybrid assembly approach, Illumina reads with short- and long-insert libraries, coupled with long Pacific Biosciences reads, we obtained an assembled genome size of 542 Mb, in line with flow cytometric genome size estimation. The largest scaffold was of 1.15 Mb, the N50 length was 145 kb, and the L50 count was 983. The assembly contained 0.12% of Ns. A Benchmarking with Universal Single-Copy Orthologs (BUSCO) analysis retrieved 94% complete BUSCO genes, well in the range of other high-quality draft genomes of trees. A total of 62,012 protein-coding genes were predicted, assisted by transcriptome sequencing. In addition, we are reporting an efficient method for extracting high-molecular-weight DNA from dormant buds, by which contamination by environmental bacteria and fungi was kept at a minimum.
Conclusions: The assembled genome will be a valuable resource and reference for future population genomics studies on the evolution and past climate change adaptation of beech and will be helpful for identifying genes, e.g., involved in drought tolerance, in order to select and breed individuals to adapt forestry to climate change in Europe. A continuously updated genome browser and download page can be accessed from beechgenome.net, which will include future genome versions of the reference individual Bhaga, as new sequencing approaches develop.
The aim of this study was to assess whether endosperm-specific carotenoid biosynthesis influenced core metabolic processes in maize embryo and endosperm and how global seed metabolism adapted to this expanded biosynthetic capacity. Although enhancement of carotenoid biosynthesis was targeted to the endosperm of maize kernels, a concurrent up-regulation of sterol and fatty acid biosynthesis in the embryo was measured. Targeted terpenoid analysis, and non-targeted metabolomic, proteomic, and transcriptomic profiling revealed changes especially in carbohydrate metabolism in the transgenic line. In-depth analysis of the data, including changes of metabolite pools and increased enzyme and transcript concentrations, gave a first insight into the metabolic variation precipitated by the higher up-stream metabolite demand by the extended biosynthesis capacities for terpenoids and fatty acids. An integrative model is put forward to explain the metabolic regulation for the increased provision of terpenoid and fatty acid precursors, particularly glyceraldehyde 3-phosphate and pyruvate or acetyl-CoA from imported fructose and glucose. The model was supported by higher activities of fructokinase, glucose 6-phosphate isomerase, and fructose 1,6-bisphosphate aldolase indicating a higher flux through the glycolytic pathway. Although pyruvate and acetyl-CoA utilization was higher in the engineered line, pyruvate kinase activity was lower. A sufficient provision of both metabolites may be supported by a by-pass in a reaction sequence involving phosphoenolpyruvate carboxylase, malate dehydrogenase, and malic enzyme.