Dataset Information

Global transcriptomic analyses of pluripotent stem cell-derived type II alveolar epithelial-like cells at 1 and 4 days after infection with SARS-CoV-2

ABSTRACT: We performed transcriptomic profiling of cells derived from human pluripotent stem cells (PSCs) using our previously described distal lung directed differentiation protocol to generate alveolar type II epithelial-like cells (iAT2s). We used the SPC2 human iPSC line containing a SFTPC-tdTomato knock-in reporter. SFTPC-tdTomato+ iAT2s were sorted on day 41 and again on day 69 of differentiation. iAT2s were single-cell passaged in self-renewing 3D alveolosphere culture approximately every 2 weeks through day 194 of differentiation. On day 208, iAT2s were single-cell passaged onto Transwell inserts. Apical media was removed on day 210 to initiate air-liquid interface culture. On day 218, 6 replicate wells of iAT2s were exposed to SARS-CoV-2 in an apical inoculum and 3 replicate wells were exposed to mock. On day 219, three mock and three post-infection samples (1 dpi) were collected. Three additional post-infection samples (4 dpi) were collected on day 222. We find that SARS-CoV-2 infected iAT2s express a rapid global transcriptomic change characterized by a shift to an inflammatory phenotype, time-dependent epithelial interferon responses, and rapid loss of the mature lung alveolar epithelial program.

ORGANISM(S): Homo sapiens

PROVIDER: GSE153277 | GEO | 2020/09/21

REPOSITORIES: GEO

ACCESS DATA

Similar Datasets

Project description:We performed transcriptomic profiling of freshly isolated primary adult human alveolar epithelial type 2 cells (AEC2s), their isogenic cultured progeny, and human induced pluripotent stem cell-derived AEC2s (iAEC2s). Distal lung preparations from adult donor lung explants from two individuals were cryopreserved using methods we have previously described. Primary AEC2s were purified using fluorescence activated cell sorting (FACS) to isolate cells co-expressing EPCAM and HTII-280. To establish cultures of primary AEC2s, these cells were combined with MRC5 fibroblasts at an 1:10 ratio on cell culture inserts in CK+DCI medium. iAEC2s were derived using our previously published lung directed differentiation protocol. We used the SPC2 human iPSC line and specifically the SPC2-ST-B2 (SFTPC tdT/WT ) clone containing a SFTPC tdTomato knock-in reporter. SFTPC tdTomato+ cells were sorted on day 41 of differentiation. iAEC2s were single-cell passaged in self-renewing feeder-free 3D cultures approximately every 2 weeks. We used the 10X Chromium System to generate single-cell transcriptomic data from 5 samples prepared in parallel and sequenced on the same day. We profiled (a) preculture primary AEC2s, (b) cultured primary AEC2s, (c) 3D feeder-free iAEC2s, (d) feeder-free iAEC2 cultured on cell culture inserts, and (e) iAEC2s cultured on cell inserts with MRC5 fibroblasts. The cultured primary AEC2 samples and the iAEC2 sample co-cultured with MRC5s were sorted for live EPCAM+ cells prior to encapsulation. All other samples were sorted for live cells. We find that although each population (fresh, primary cultured, and iPSC-derived AEC2s) occupies a distinct transcriptomic space, both cultured primary AEC2s and cultured feeder-free iAEC2s closely resemble their primary pre-cultured counterparts. We also find a continuum of progressively more proliferative states (from fresh primary to cultured primary to cultured iAEC2s) inversely associated with progressively less mature AEC2 states.

Project description:In susceptible plant hosts, co-evolution has favoured viral strategies to evade host defenses and utilize resources to their own benefit. The degree of manipulation of host gene expression is dependent on host-virus specificity and certain abiotic factors. In order to gain insight into global transcriptomic changes for a geminivirus pathosystem, South African cassava mosaic virus [ZA:99] (SACMV-ZA:99]) and Arabidopsis thaliana, 4 x 44K Agilent microarrays were adopted. After normalization, a 2-fold change filtering of data (p<0.05) identified 1,820 differentially expressed genes in apical leaf tissue. A significant increase in differential gene expression over time (451 genes at 14 dpi, 742 genes at 24 dpi, and 1011 genes at 36 dpi) was observed. This increase in expression, correlated with an increase in SACMV accumulation as virus copies were 5-fold higher at 24 dpi and 6-fold higher at 36 dpi than at 14 dpi (1.1x104 virus copies present at 14 dpi, 5.7x104 copies at 24 dpi, and 6.3x104 copies at 36 dpi). Many 2-fold genes were primarily involved in stress and defense responses, phytohormone signalling pathways, cellular transport, cell-cycle regulation, transcription, oxidation-reduction, and other metabolic processes. Forty-one genes (2.3%) were shown to be continuously expressed across the infection period, indicating that the majority of genes were transient and unique to a particular time point. Plant signalling networks were disrupted and manipulated by SACMV-[ZA:99] in order to affect homeostasis and antagonize hostM-bM-^@M-^Ys defense responses. At the same time, an adaptive response was initiated to reprogramme metabolism and divert energy from growth-related processes to defense, all leading to disruption of normal biological host processes. Comparisons between SACMV-[ZA:99] with plant-infecting RNA and DNA viruses revealed similarities and differences in expression patterns among viruses, showing either general defense or virus-specific responses. Within the Geminiviridae family in particular, similarities in cell-cycle regulation and gene expression patterns correlated between SACMV-[ZA:99] and Cabbage leaf curl virus (CaLCuV) but differences were also evident. For instance, CaLCuV showed antagonistic interactions between Salicyclic Acid (SA) and Jasmonic Acid (JA) pathways, whereas SACMV displayed synergism. Differences in gene induction, repression and outcome between the two geminiviruses clearly demonstrated host-specific interactions with SACMV-[ZA:99] leading to infection. To our knowledge this is the first geminivirus study identifying differentially expressed transcripts across 3 time points A three time-point (14, 24, and 36 dpi) study was carried out to identify differentially expressed genes in SACMV-[ZA:99] infected Arabidopsis leaf cells using a direct comparison design against mock-inoculated controls. Three biological replicates and 1 technical replicate for both SACMV-[ZA:99]-infected and mock-inoculated controls were conducted at each time point.

Dataset Information

Global transcriptomic analyses of pluripotent stem cell-derived type II alveolar epithelial-like cells at 1 and 4 days after infection with SARS-CoV-2

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure