Smoking Modulates Different Secretory Subpopulations Expressing SARS-CoV-2 Entry Genes in the Nasal and Bronchial Airways

Background: SARS-CoV-2 infection and disease severity are influenced by viral entry (VE) gene expression patterns in airway epithelium. The similarities and differences of VE gene expression (ACE2, TMPRSS2, and CTSL) across nasal and bronchial compartments has not been fully characterized using matched samples from large cohorts. Results: Gene expression data from 793 nasal and 1,673 bronchial brushes obtained from individuals participating in lung cancer screening or diagnostic workup revealed that smoking was the only clinical factor significantly and reproducibly associated with VE gene expression. ACE2 and TMPRSS2 expression were higher in smokers in the bronchus but not in the nose. scRNA-seq of nasal brushings indicated that ACE2 co-expressed genes were highly expressed in club and C15orf48+ secretory cells while TMPRSS2 co-expressed genes were highly expressed in keratinizing epithelial cells. In contrast, these ACE2 and TMPRSS2 modules were highly expressed in goblet cells in scRNA-seq from bronchial brushings. Cell-type deconvolution of the RNA-seq confirmed that smoking increased the abundance of several secretory cell populations in the bronchus, but only goblet cells in the nose. Conclusions: The association of ACE2 and TMPRSS2 with smoking in the bronchus is due to their high expression in goblet cells which increase in abundance in current smoker airways. In contrast, in the nose these genes are not predominantly expressed in cell populations modulated by smoking. Smoking-induced VE gene expression changes in the nose likely has minimal impact on SARS-CoV-2 infection, but in the bronchus, smoking may lead to higher viral loads and more severe disease.

Smoking modulates different secretory subpopulations expressing SARS-CoV-2 entry genes in the nasal and bronchial airways

Coronavirus Disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 SARS-CoV-2), which infects host cells with help from the Viral Entry (VE) proteins ACE2, TMPRSS2, and CTSL1–4. Proposed risk factors for viral infection, as well as the rate of disease progression, include age5,6, sex7, chronic obstructive pulmonary disease7,8, cancer9, and cigarette smoking10–13. To investigate whether the proposed risk factors increase viral infection by modulation of the VE genes, we examined gene expression profiles of 796 nasal and 1,673 bronchial samples across four lung cancer screening cohorts containing individuals without COVID-19. Smoking was the only clinical factor reproducibly associated with the expression of any VE gene across cohorts. ACE2 expression was significantly up-regulated with smoking in the bronchus but significantly down-regulated with smoking in the nose. Furthermore, expression of individual VE genes were not correlated between paired nasal and bronchial samples from the same patients. Single-cell RNA-seq of nasal brushings revealed that an ACE2 gene module was detected in a variety of nasal secretory cells with the highest expression in the C15orf48+ secretory cells, while a TMPRSS2 gene module was most highly expressed in nasal keratinizing epithelial cells. In contrast, single-cell RNA-seq of bronchial brushings revealed that ACE2 andTMPRSS2 gene modules were most enriched in MUC5AC+ bronchial goblet cells. The CTSL gene module was highly expressed in immune populations of both nasal and bronchial brushings. Deconvolution of bulk RNA-seq showed that the proportion of MUC5AC+ goblet cells was increased in current smokers in both the nose and bronchus but proportions of nasal keratinizing epithelial cells, C15orf48+ secretory cells, and immune cells were not associated with smoking status. The complex association between VE gene expression and smoking in the nasal and bronchial epithelium revealed by our results may partially explain conflicting reports on the association between smoking and SARS-CoV-2 infection.

Colonization of tomatoes by Verticillium dahliae: determinative phase II

To investigate the relationship between stem colonizing ability of Verticillium and wilt disease development, the colonization capacities of Verticillium dahliae, race 1 or race 2, were compared in near-isolines of the cultivar Craigella, with (Ve+) and without (Ve−) Ve-gene resistance, and in the cultivar IRAT-L3, which also lacks the Ve-gene. Only the Ve+/V.d.1 interaction was resistant to disease. Lateral and vertical invasiveness of the pathogen were assessed cytologically and fungal biomass was monitored by PCR-based diagnostics. The pathogen was most aggressive in the Ve−/V.d.1 interaction; colonization capacity was reduced in the other five combinations, but most severely, and equivalently, in Ve+/V.d.1, IRAT/V.d.1, and IRAT/V.d.2. Further study of the development and role of the vascular coating response in IRAT-L3 indicated a pattern of expression similar to that observed in other susceptible plants, and it was concluded that the reduced colonization of this cultivar by V. dahliae resulted from another unknown defensive mechanism. Stem colonizing ability that is compatible with resistance in one cultivar and race combination may promote symptom expression and even death in another. Keywords: tomato, Verticillium dahliae, vascular coating, resistance.

Vascular coating: a barrier to colonization by the pathogen in Verticillium wilt of tomato

Massive infusion of conidia of Verticillium albo-atrum Reinke & Berthier induced synchronous secretion of vascular coating in the petiolar xylem vessels of resistant and susceptible tomato near-isolines. More coating formed earlier in resistant than in susceptible plants. In the susceptible plants secretion was delayed in colonized trapping site vessels, but initiated in surrounding uncolonized ones. Controls were infused with water. Samples were quantified by light microscope assay techniques at 18, 48, and 120 h postinoculation for the following parameters: (i) delayed coating effect, (ii) overall coating capacity, and (iii) ability of fungus to "escape" laterally from trapping site vessels. The results showed that susceptibility to Verticillium was absolutely correlated with the presence of the delayed coating effect in the plant and increased ability of the fungus to spread laterally. Treatment of inoculated resistant plants with an inhibitor of coating secretion resulted in conversion to the susceptible phenotype. The progeny of a genetic backcross for the dominant (Ve) and recessive (ve) alleles at the Ve locus (Velve × velve) were assayed for the same parameters as well as for disease resistance based on symptom expression. The results confirmed the previous observations and suggested that in tomato the delayed phenotype is recessive. The data strongly supports two hypotheses: (i) coating forms a barrier against fungal penetration and (ii) the timing of the coating response in trapping site vessels results, directly or indirectly, from expression of the Ve gene.