Integrated analyses of single-cell atlases reveal age, gender, and smoking status associations with cell type-specific expression of mediators of SARS-CoV-2 viral entry and highlights inflammatory programs in putative target cells

Christoph Muus; Malte D Luecken; Gokcen Eraslan; Avinash Waghray; Graham Heimberg; Lisa Sikkema; Yoshihiko Kobayashi; Eeshit Dhaval Vaishnav; Ayshwarya Subramanian; Christopher Smillie; Karthik Jagadeesh; Elizabeth Thu Duong; Evgenij Fiskin; Elena Torlai Triglia; Christophe Becavin; Meshal Ansari; Peiwen Cai; Brian Lin; Justin Buchanan; Sijia Chen; Jian Shu; Adam L Haber; Hattie Chung; Daniel T Montoro; Taylor Adams; Hananeh Aliee; Samuel J Allon; Zaneta Andrusivova; Ilias Angelidis; Orr Ashenberg; Kevin Bassler; Christophe Becavin; Inbal Benhar; Joseph Bergenstrahle; Ludvig Bergenstrahle; Liam Bolt; Emelie Braun; Linh T Bui; Mark Chaffin; Evgeny Chichelnitskiy; Joshua Chiou; Thomas M Conlon; Michael S Cuoco; Marie Deprez; David S Fischer; Astrid Gillich; Joshua Gould; Minzhe Guo; Austin J Gutierrez; Arun C Habermann; Tyler Harvey; Peng He; Xiaomeng Hou; Lijuan Hu; Alok Jaiswal; Peiyong Jiang; Theodoros Kapellos; Christin S Kuo; Ludvig Larsson; Michael A Leney-Greene; Kyungtae Lim; Monika Litvinukova; Ji Lu; Leif S Ludwig; Wendy Luo; Henrike Maatz; Elo Maddissoon; Lira Mamanova; Kasidet Manakongtreecheep; Charles-Hugo Marquette; Ian Mbano; Alexi M McAdams; Ross J Metzger; Ahmad N Nabhan; Sarah K Nyquist; Jose Ordovas-Montanes; Lolita Penland; Olivier B Poirion; Segio Poli; CanCan Qi; Daniel Reichart; Ivan Rosas; Jonas Schupp; Rahul Sinha; Rene V Sit; Kamil Slowikowski; Michal Slyper; Neal Smith; Alex Sountoulidis; Maximilian Strunz; Dawei Sun; Carlos Talavera-Lopez; Peng Tan; Jessica Tantivit; Kyle J Travaglini; Nathan R Tucker; Katherine Vernon; Marc H Wadsworth; Julia Waldman; Xiuting Wang; Wenjun Yan; Ali Onder Yildirim; William Zhao; Carly G K Ziegler; Aviv Regev

, sex, and smoking status associations with expression of ACE2, TMPRSS2, and CTSL across level 3 cell type annotations. Effect size (y axis) of the association as log fold changes (sex, smoking status) and slope of log expression with age. Bars that are colored in indicate associations with an FDR-corrected p-value of < 0.05 where the pseudo-bulk analysis shows a consistent effect direction. Error bars represent model uncertainties. Comparison of signature scores of cell programs between DP and DN cells for each cell type stratified by gene complexity bin. Cells were partitioned into 10 gene complexity bins for every cell type. (d,e) IL6 and its receptor's expression in specific cell types in lung and heart. (d) Significance (dot size) and fold change (dot color) of differential expression between DP and DN cells within different types (rows) for IL6 and its receptors IL6R and IL6ST (columns) across tissues. (e) Top: Significance (dot size) and fold change (dot color) of differential expression between DP and DN cells within different cell types in the heart (rows)for IL6 and its receptors IL6R and IL6ST (columns). Bottom: Significance (dot size) and effect size (dot color) from a mixed . CC-BY-ND 4.0 International license author/funder. It is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.04.19.049254 doi: bioRxiv preprint effects model of co-expression of IL6, IL6R, or IL6ST (columns) coexpression with ACE2. (f) Distribution of number of counts in peaks (y axis) in ACE2 + epithelial cells (having at least 1 fragment in the ACE2 gene locus) and ACE2cells. Figure 12. Co-expression of ACE2 and IL6,IL6R,IL6ST . Co-expression of ACE2 and IL6,IL6R,IL6ST in select single-cell datasets. P-values and significance (FDR 10%) derived from the logistic mixed-effects model. Figure 13 . Expression of Ace2, Tmprss2 and Ctsl in mouse placenta. UMAP embedding (as in Fig. 5k ) of scRNA-seq profiles of placenta cells collected at E.14.5 (top) or along a time course (bottom), colored by expression level ofAce2, Tmprss2, and Ctsl. Extended Data Figure 14 . Additional analyses to identify other proteases that may have a role in infection. (a) Multiple proteases are co-expressed with ACE2 in another human lung scRNA-seq ("aggregated lung"). Scatter plot of significance (y axis, -log10(adjusted p value)) and effect size (x axis) of co-expression of each protease gene (dot) with ACE within each indicated epithelial cell type (color). Dashed line: significance threshold. TMPRSS2 and PCSKs that significantly coexpressed with ACE2 are marked. (b) ACE2-protease co-expression with PCSKs, TMPRSS2 and CTSL across lung cell types ("aggregated lung"). Significance (dot size, -log10(adjusted p value)) and effect size (color) for co-expression of ACE2 with selected proteases (columns) across cell types (rows). (c-d) Predicted cleavage sites in the SARS-CoV-2 S-protein S1/S2 region. (c) Multiple amino acid sequence alignment of SARS-CoV-2 S-protein S1/S2 region with orthologous sequences from other betacoronaviruses (top) and polybasic cleavage sites of other human pathogenic viruses (bottom). The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.04.19.049254 doi: bioRxiv preprint Significance (dot size) and effect size (dot color) of co-expression of each protease (columns) with ACE2 in each cell subset (rows). Figure 16 . Variation in f