Non-neural expression of SARS-CoV-2 entry genes in the olfactory epithelium suggests mechanisms underlying anosmia in COVID-19 patients

David Brann; Tatsuya Tsukahara; Caleb Weinreb; Darren W. Logan; Sandeep Robert Datta

The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.03.25.009084 doi: bioRxiv preprint all cells with the normalized expression of CoV-2 related genes ACE2 and TMPRSS2, as well as markers of several cell types. ACE2 and TMPRSS2 are expressed in respiratory and olfactory cell types, but not in OSNs. ACE2 and TMPRSS2 are detected in HBC (KRT5) and sustentacular (CYP2A13) cells, as well as other respiratory epithelial cell types, including respiratory ciliated (FOXJ1) cells. (E) Cell type markers and various CoV related genes including ACE2 and TMPRSS2, are expressed in respiratory and olfactory cell types, but not in OSNs. Gene expression for ACE2 and TMPRSS2 as well as marker genes for olfactory and respiratory epithelial subtypes are shown normalized by their maximum expression across cell types. MHV, mouse hepatitis virus. Given the potential for the RE and OE in the nasal cavity to be directly infected with CoV-2, we assessed the expression of Ace2 and other CoV entry genes in the mouse olfactory bulb (OB), which is directly connected to the OSNs in OE via cranial nerve I (CN I); in principle, alterations in bulb function could cause anosmia independently of functional changes in the OE. To do so, we performed scSeq (using DropSeq, see Methods) on the mouse OB, and merged these data with a previously published OB scSeq analysis, yielding a dataset with nearly 50,000 single cells (see Methods) (45) . This analysis revealed that Ace2 expression was absent from OB neurons and instead was observed only in vascular pericytes (Figures 4A-D and S5-6), which are involved in blood pressure regulation, maintenance of the blood-brain barrier, and inflammatory responses (46) . Although other potential CoV proteases were expressed in the OB, Tmprss2 was not expressed in the olfactory bulb. We also performed Smart-seq2-based deep sequencing of single OB dopaminergic neurons ( Figures 4E and S7 , see Methods), which confirmed the absence of Ace2 and Tmprss2 expression in that cell type. In addition, re-analysis of 10 deeply sequenced datasets from different regions of the nervous system demonstrated that Ace2 expression is absent from neurons, consistent with prior immunostaining results ( Figure S8 ) (47) . Together these findings suggest that OB neurons are likely not a primary site of infection, but that vascular pericytes may be sensitive to CoV-2 infection in the OB and brain.