Prediction of Non-canonical Routes for SARS-CoV-2 Infection in Human Placenta Cells

Abstract

The SARS-CoV-2 is the causative agent of the COVID-19 pandemic. The data available about COVID-19 during pregnancy have demonstrated placental infection; however, the mechanisms associated with intrauterine transmission of SARS-CoV-2 is still debated. Intriguingly, while canonical SARS-CoV-2 cell entry mediators are expressed at low levels in placental cells, the receptors for viruses that cause congenital infections such as the cytomegalovirus and Zika virus are highly expressed in these cells. Here we analyzed the transcriptional profile (microarray and single-cell RNA-Seq) of proteins potentially interacting with coronaviruses to identify non- canonical mediators of SARS-CoV-2 infection and replication in the placenta. Despite low levels of the canonical cell entry mediators ACE2 and TMPRSS2, we show that cells of the syncytiotrophoblast, villous cytotrophoblast, and extravillous trophoblast co-express high levels of the potential non-canonical cell-entry mediators DPP4 and CTSL. We also found changes in the expression of DAAM1 and PAICS genes during pregnancy, which are translated into proteins also predicted to interact with coronaviruses proteins. These results provide new insight into the interaction between SARS-CoV-2 and host proteins that may act as non-canonical routes for SARS-CoV-2 infection and replication in the placenta cells.

Keywords: COVID-19; CTSL; DAAM1; DPP4; PAICS; SARS-CoV-2; placenta; virus entry mediator.

FIGURE 1

The expression landscape of human placenta proteins potentially interacting with SARS-CoV-2. (A). Heatmap illustrating gene expression (counts RMA normalized) of SARS-CoV-2 interacting proteins from placental samples in the first, second and third trimester of gestation (GSE9984). (B). Heat-scatter plot presenting ten differentially expressed genes in the placenta that encode proteins potentially interacting with SARS-CoV as predicted by the P-HIPSter (http://phipster.org/) database. The color and size of the circles correspond to the log2FC and -log10 transformed FDR adjusted p-value, respectively. Fold change represents the gene expression of placenta samples from the third trimester of gestation compared with the first trimester. Bolded genes represent the six candidates selected for the further scRNA-seq evaluations. (C). Tissue-specific gene network of placenta proteins predicted to interact with coronaviruses. The network was generated using the humanbase (https://hb.flatironinstitute.org/) online tool. (D). Dimensionality reduction demonstrates different cell populations identified in five first-trimester human placental samples in a Uniform Manifold Approximation and Projection (UMAP) plot, by using scRNA-Seq data from Vento-Tormo et al., 2018. The images were generated using the COVID-19 Cell Atlas (https://www.covid19cellatlas.org) functionalities. (E). UMAP plots representing the gene expression (log-transformed normalized counts) of ACE2, TMPRSS2, CTSL, DPP4, DAAM1, and PAICS in human placental cells from the syncytiotrophoblast, villous cytotrophoblast, and extravillous trophoblast. The images were generated using COVID-19 Cell Atlas (https://www.covid19cellatlas.org) functionalities. (F). Cell illustrations depicting the protein localization of ACE2, TMPRSS2, CTSL, DPP4, DAAM1, and PAICS in cell compartments, as described in the Human Protein Atlas database (http://www.proteinatlas.org). Illustrations were created using BioRender App (https://biorender.com/). (G). Heat-scatter plot presenting gene expression of ACE2, TMPRSS2, CTSL, DPP4, DAAM1, and PAICS in the placenta from COVID-19 pregnant women compared to healthy pregnant (GSE17995). The color and size of the circles correspond to the log2FC and -log10 transformed p-values, respectively. Fold change represents the gene expression of placenta samples from the third trimester of gestation of COVID-19 women compared with matched third trimester uninfected women. RMA: Robust Multichip Average; FC: Fold Change; FDR: false discovery rate; Padj: adjusted p-value; SCT, syncytiotrophoblast; VCT, villous cytotrophoblast; EVT, extravillous trophoblast; HB, Hofbauer cells; FB: fibroblasts; dM, decidual macrophages; Endo, endothelial cells; l, lymphatic; m, maternal; f, fetal.

FIGURE 2

Single-cell analysis of the human placenta indicates potential non-canonical routes of SARS-CoV-2. (A). Co-expression analysis of DAAM1, CTSL, DPP4, and PAICS with the canonical virus entry-associated genes ACE2 and TMPRSS2. (B). Co-expression analysis of DAAM1 and PAICS with the non-canonical virus entry genes DPP4 and CTLS. The images were generated using COVID-19 Cell Atlas (https://www.covid19cellatlas.org) functionalities.