1.
Pottmeier, Philipp, 1987-, et al.
(författare)
Sex-biased Gene Expression During Neural Differentiation of Human Embryonic Stem Cells
Annan publikation (övrigt vetenskapligt/konstnärligt) abstract
Sex differences in the adult human brain are mainly attributed to hormonal influence. Recently however, genetic differences and their impact on the developing nervous system have attracted increasing attention. In order to understand the genetically driven sexual dimorphisms, we investigated genome wide gene expression in an in vitro differentiation model of male and female human embryonic stem cell lines. Four male and four female derived hESC lines were differentiated towards a population of mixed neurons over 37 days. Differential gene expression and gene set enrichment analyses were conducted based on bulk RNA sequencing data. While similar differentiation tendencies in all cell lines proved the robustness and reproducibility of our differentiation protocol, we found sex-biased gene expression already in the undifferentiated ESCs at day 0, but most profound after 37 days of differentiation. Male and female cell lines show sex-biased expression of genes involved in neurodevelopment, suggesting a sex difference in differentiation trajectory. Interestingly, the highest contribution was found to arise from the male transcriptome, involving both Y chromosome and autosomal genes. We propose 13 sex-biased candidate genes (10 upregulated in male cell lines and 3 in female lines) that could strongly affect neuronal development. In addition, we confirmed gene dosage compensation of X/Y homologs escaping X chromosome inactivation through their Y homolog and identified a significantly increased Y homolog overexpression of the gametologs TXLNG/Y and KDM6A/UTY after 37 days of neuronal differentiation. These results suggest sex differences in the trajectories of neuronal differentiation which could ultimately contribute to sex biases during human brain development.
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Ali, Muhammad Akhtar, et al.
(författare)
The transcriptional modulator ZBED6 regulates cell cycle and growth of human colorectal cancer cells
Annan publikation (övrigt vetenskapligt/konstnärligt) abstract
The transcription factor ZBED6 is a repressor of IGF2 whose action impacts development, cell proliferation and growth in placental mammals. In human colorectal cancers, IGF2 overexpression is mutually exclusive with somatic mutations in PI3K signaling components, providing genetic evidence for a role in the PI3K pathway. To understand the role of ZBED6 in tumorigenesis, we engineered and validated somatic cell ZBED6 knock-outs in the human colorectal cancer cell lines RKO and HCT116. Transcriptome analyses revealed enrichment of cell cycle-related processes among differentially expressed genes in both cell lines. Chromatin immunoprecipitation sequencing analyses displayed enrichment of ZBED6 binding at genes upregulated in ZBED6-/- knockout clones. Ten differentially expressed genes were identified as putative direct gene targets and their downregulation by ZBED6 was experimentally validated. Eight of these genes were linked to the Wnt, Hippo, TGF-b, EGFR or PI3K pathways, all involved in colorectal cancer development. Ablation of ZBED6 affected the cell cycle and led to increased growth rate of ZBED6-/- RKO cells. These observations support a role for transcriptional modulation by ZBED6 in cell cycle regulation and growth of colorectal cancers.
5.
Arvidsson, Malou, et al.
(författare)
An annotated high-content fluorescence microscopy dataset with Hoechst 33342-stained nuclei and manually labelled outlines : Dataset record
2022
Annan publikation (övrigt vetenskapligt/konstnärligt) abstract
Here we present a benchmarking dataset of fluorescence microscopy images with Hoechst 33342-stained nuclei together with annotations of nuclei, nuclear fragments and micronuclei. Images were randomly selected from an RNA interference screen with a modified U2OS osteosarcoma cell line, acquired on a Thermo Fischer CX7 high-content imaging system at 20x magnification. Labelling was performed by a single annotator and reviewed by a biomedical expert.The dataset contains 50 images showing over 2000 labelled nuclear objects in total, which is sufficiently large to train well-performing neural networks for instance or semantic segmentation. It is pre-split into training, development and test set, each in a zip file. The dataset should be referred to as Aitslab_bioimaging1. A preprint of a brief article describing the dataset is also available from zenodo (Arvidsson M, Kazemi Rashed S, Aits S. zenodo 2022, https://doi.org/10.1016/j.dib.2022.108769)
6.
Badhai, Jitendra, et al.
(författare)
Differential expression of RPS19 5’UTR variants implicated in Diamond-Blackfan anemia
2012
Annan publikation (övrigt vetenskapligt/konstnärligt) abstract
Heterozygous mutations in the ribosomal protein (RP) S19 gene RPS19 are found in about 25% of patients with the congenital erythroblastopenia Diamond-Blackfan anemia (DBA). The RPS19 gene encodes a single RPS19 isoform from three known transcriptional start sites (TSS) with different 5’ untranslated region (UTR). The regulation of RPS19 expression is poorly understood as well as the significance of different 5’UTRs. A few rare sequence variants within the 5’UTR have also been reported in patients with DBA. We determined the transcriptional start sites (TSS) and the tissue distribution of variant 5’UTRs of RPS19. Twenty-nine novel TSS in K562 cells and testis were identified. We then analyzed the relative proportion of three selected 5’UTRs of different length on a panel of primary tissues. The shorter 5’UTR were most abundant in all tissues but with large variations in relative levels of shorter versus longer transcripts. To clarify the effect of different RPS19 5’UTRs on translation we designed and expressed constructs using three 5’UTRs of different length. The short 5’UTR(+35nt.) translate 4-6 folds more efficiently than the two longer variants with 5’UTRs of 382nt. and 467nt., respectively We also introduced DBA associated insertion (c.-149_-148insGCCA, c.-149_-148insAGCC ) and deletion (c.-144_-141delTTTC) variants in the 5’UTR. . Interestingly, the DBA associated 5’UTR sequence variants showed a 20-30% reduction in RPS19 levels when compared to the corresponding w.t. constructs. Our results indicate that the RPS19 gene has a broad range of TSS with tissue specific variations. We also show that sequence variants in the 5’UTR in some DBA patients reduce RPS19 expression with implications for the pathophysiology of the disease.
7.
Bandaru, Manoj Kumar, 1987-, et al.
(författare)
Apoc2 mutant zebrafish: a model for hypertriglyceridemia and early-stage atherosclerosis
Annan publikation (övrigt vetenskapligt/konstnärligt) abstract
Zebrafish larvae in a hypertriglyceridemic background can be useful to identify and characterize causal genes for triglyceride metabolism. A previous, small-scale study suggested that apolipoprotein C-II (apoc2)-mutant zebrafish larvae can be used to model hypertriglyceridemia-induced atherosclerosis. We aimed to replicate these findings in a large-scale study and asses if APOC-II may represent a useful therapeutic target. We generated apoc2 mutant zebrafish using CRISPR-Cas9 and examined cardiometabolomic traits in their offspring (F1 generation). Systematic characterization of 384 larvae using our image and assay-based, high-throughput pipeline showed that compound heterozygous larvae for loss of function mutations in apoc2 (n=35) have higher whole-body levels of triglycerides (0.71±0.16 SD), HDL cholesterol (0.32±0.15 SD) and total cholesterol (0.37±0.18 SD), and a trend for lower whole-body glucose levels (0.23±0.14 SD) compared with larvae without mutations in apoc2 (n=174). Such larvae also tended to have more vascular lipid deposition, however this effect did not reach significance (P=0.12). Interestingly, the trends for lower whole-body glucose levels and more vascular lipid deposition in larvae with anticipated loss of functional apoc2 reached significance when larvae (n=3812) from other screens, in which apoc2 was not experimentally perturbed were included as additional wildtype controls. Thus, our large-scale study confirms the role of apoc2 in hypertriglyceridemia and early-stage atherosclerosis. While apoc2 mutant zebrafish model can be used as a genetic background to identify and characterize causal genes for triglyceride metabolism, independent and opposite effects on triglycerides and glucose suggest that APOC-II is likely not a suitable target for prevention and treatment of coronary artery disease.
8.
Bandaru, Manoj Kumar, 1987-, et al.
(författare)
Image-based, in vivo characterization of cardiometabolic consequences of mutations in pcsk9
Annan publikation (övrigt vetenskapligt/konstnärligt) abstract
Based on the association of loss-of-function mutations in proprotein convertase subtilisin/kexin type 9 (PCSK9) with low plasma LDL cholesterol levels, inhibition of the PCSK9 protein using monoclonal antibodies have emerged as an effective treatment option to lower LDL cholesterol levels and reduce the risk of coronary artery disease. Despite these beneficial effects, PCSK9 inhibitors may increase the risk of diabetes. In this study, we mimicked the mechanistic action of PCSK9 inhibitors in humans by inducing mutations in pcsk9 in zebrafish and examining their effects on dyslipidemia, early-stage atherosclerosis and diabetes-related traits in data from nearly 5000 zebrafish larvae. At 10 days of age, larvae with mutations in pcsk9 were characterized by lower whole-body LDL cholesterol levels (beta±SE -0.056±0.025 SD units) and protection against early-stage atherosclerosis, with less vascular lipid deposition (-0.133±0.035 SD) and less co-localization of macrophages with lipids (-0.086±0.032 SD). Mutant larvae also had fewer pancreatic β-cells (-0.153±0.055 SD). Thus, our findings in pcsk9 mutant larvae are in line with results from people carrying loss-of-function PCSK9 mutations, and are also in line with the effects of PCSK9 inhibitors in humans. Further, our results suggest that mutations in pcsk9 may increase the risk of diabetes through a direct effect on pancreatic β-cells.
9.
Brueffer, Christian, et al.
(författare)
The Mutational Landscape of the SCAN-B Real-World Primary Breast Cancer Transcriptome
2020
Annan publikation (övrigt vetenskapligt/konstnärligt) abstract
Breast cancer is a disease of genomic alterations, of which the complete panorama of somatic mutations and how these relate to molecular subtypes and therapy response is incompletely understood. Within the Sweden Cancerome Analysis Network–Breast project (SCAN-B; ClinicalTrials.govNCT02306096), an ongoing study elucidating the tumor transcriptomic profiles for thousands of breast cancers prospectively, we developed an optimized pipeline for detection of single nucleotide variants and small insertions and deletions from RNA sequencing (RNA-seq) data, and profiled a large real-world population-based cohort of 3,217 breast tumors. We use it to describe the mutational landscape of primary breast cancer viewed through the transcriptome of a large population-based cohort of patients, and relate it to patient overall survival. We demonstrate that RNA-seq can be used to call mutations in important breast cancer genes such asPIK3CA,TP53, andERBB2, as well as the status of key molecular pathways and tumor mutational burden, and identify potentially druggable genes in 86.8% percent of tumors. To make this rich and growing mutational portraiture of breast cancer available for the wider research community, we developed an open source web-based application, the SCAN-B MutationExplorer, accessible athttp://oncogenomics.bmc.lu.se/MutationExplorer. These results add another dimension to the use of RNA-seq as a potential clinical tool, where both gene expression-based and gene mutation-based biomarkers can be interrogated simultaneously and in real-time within one week of tumor sampling.
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