key: cord-0799349-f4mj7i69 authors: Seethy, Ashikh A.; Singh, Sunil; Mukherjee, Indrani; Pethusamy, Karthikeyan; Purkayastha, Kakali; Sharma, Jai Bhagwan; Sharma, Radhey S.; Dhar, Ruby; Karmakar, Subhradip title: Potential SARS-CoV-2 interactions with proteins involved in trophoblast functions – An in-silico study date: 2020-10-22 journal: Placenta DOI: 10.1016/j.placenta.2020.10.027 sha: 5a5862643b3b0ddd76b292e1b02a0908a8b739bd doc_id: 799349 cord_uid: f4mj7i69 BACKGROUND: Though a large number of pregnant females have been affected by COVID-19, there is a dearth of information on the effects of SARS-CoV-2 infection on trophoblast function. We explored in silico, the potential interactions between SARS-CoV-2 proteins and proteins involved in the key functions of placenta. METHODS: Human proteins interacting with SARS-CoV-2 proteins were identified by Gordon et al. (2020). Genes that are upregulated in trophoblast sub-types and stages were obtained by gene-expression data from NCBI-GEO and by text-mining. Genes altered in pathological states like pre-eclampsia and gestational diabetes mellitus were also identified. Genes crucial in placental functions thus identified were compared to the SARS-CoV-2 interactome for overlaps. Proteins recurring across multiple study scenarios were analyzed using text mining and network analysis for their biological functions. RESULTS: The entry receptors for SARS-CoV-2 – ACE2 and TMPRSS2 are expressed in placenta. Other proteins that interact with SARS-CoV-2 like LOX, Fibulins-2 and 5, NUP98, GDF15, RBX1, CUL3, HMOX1, PLAT, MFGE8, and MRPs are vital in placental functions like trophoblast invasion and migration, syncytium formation, differentiation, and implantation. TLE3, expressed across first trimester placental tissues and cell lines, is involved in formation of placental vasculature, and is important in SARS-CoV (2003) budding and exit from the cells by COPI vesicles. CONCLUSION: SARS-CoV-2 can potentially interact with proteins having crucial roles in the placental function. Whether these potential interactions identified in silico have effects on trophoblast functions in biological settings needs to be addressed by further in vitro and clinical studies. COVID-19 patients, belong to the reproductive age group and about 47% of the affected 48 individuals are females. [3] Thus, a large number of females in the reproductive age group 49 have already been infected with SARS-CoV-2, and more are at potential risk of infection, 50 given the R0 of about 2.28 of COVID-19. [4] 51 A characterization of pregnant females who were hospitalized for delivery during the 52 COVID-19 outbreak showed that 15.4% of these women were positive for SARS-CoV-2, and 53 among these, 87.9% were asymptomatic.[5] The major consensus is that there is no vertical 54 transmission of SARS-CoV-2, like the predecessor coronaviruses causing SARS and MERS, [6-55 9] though certain systematic reviews and case reports do not completely rule out such a 56 possibility.[10-12] SARS-CoV-2 infection in the third trimester of pregnancy was found to be 57 associated with an increased risk of preterm delivery and intrauterine fetal distress and the 58 requirement for Caesarean sections. [13] During SARS of 2003, pregnant females who 59 presented with the disease during the first trimester had an adverse outcome, with 57% of 60 pregnancies ending in spontaneous miscarriage, and 4 out of 5 patients who presented with 61 the disease progressing into preterm delivery. [14] The reasons behind this adverse outcome 62 in pregnancy in females infected with SARS-CoV remains enigmatic. At present, there is 63 limited data on the effect of first trimester SARS-CoV-2 infection on the pregnancy outcome. 64 Initial reports suggest that COVID-19 does not increase the risk of spontaneous 65 abortions, [15] though placenta can be infected by SARS-CoV-2, as evident by in situ RNA 66 hybridisation, electron microscopy, [16] [17] [18] [19] [20] and detection of SARS-CoV-2 spike protein in 67 placental villi in COVID-19 positive pregnancies. [12, 21] 68 Since the placenta executes and orchestrates fetal growth-related pathways, placental 69 dysfunction has deleterious effects on the outcome of pregnancy. While certain reports 70 suggest that SARS-CoV-2 infection is not associated with specific histopathological Table 1) 90 91 Datasets from NCBI-GEO involving studies on placenta were analyzed for differential gene 92 expression using GEO-2R. The datasets used and the samples for differential expression are 93 summarised in Table 1 113 The complete approach to the discovery of the proteins involved in placental functions that 114 can bind to SARS-CoV-2 is given in Supplementary Figure 1 Analysis of two different datasets, GSE9984 and GSE28551, were performed to identify the 119 SARS-CoV-2 interacting proteins that are upregulated in first-trimester placenta and term 120 placenta. The data sets revealed that 3 proteins (GSE9984) and 14 proteins (GSE28551), 121 upregulated in first-trimester placenta interacting with SARS-CoV-2 in the two queried data 122 sets (Figures 1a and 1b) . We further observed 21 proteins (GSE9984) and 19 proteins 123 (GSE28551) that were upregulated in term placenta in these two data sets (Figures 1a and 124 1b) that could interact with SARS-CoV-2. We then identified the candidate proteins in the 125 first trimester (3 and 14) and term placenta (21 and 19) that were common in the two data 126 sets. Our results yielded LOX as a common candidate in the term placenta and SCARB1 and 127 PRKAR2B in the first trimester that could potentially interact with SARS-CoV-2 ( Figure 1c ). 128 LOX (lysyl oxidase) upregulated in term placenta can interact with SARS-CoV-2 orf8. 129 Downregulation of LOX has been implicated in impaired trophoblast migration and invasion. 130 [33] The interaction network of LOX is shown in Figure 1d . LOX is also highly expressed in interactome. Our analysis also identified many novel, unreported candidate proteins that 310 could be targeted during these infections. We also explored the differentially expressed 311 proteomes from trophoblast cell lines and found a significant number of overlaps with SARS- 312 CoV-2, indicating that these cells can be used as a model to study viral infection. 313 Since the SARS-CoV-2 interactome reported by Gordon, et Figures 1a and 1b show data from GSE9984 and GSE28551 , respectively. Figure 1c shows the overlapping proteins in term placenta (left) and first trimester placenta (right) from GSE9984 and GSE28551, that interact with SARS-CoV-2. Figure 1d shows the LOX interaction network. The expansions of all the protein names and the SARS-CoV-2 proteins interacting with them are given in Supplementary Table 2. (Figures 3a, 3b and 3c, respectively) . Figure 3d shows RBX1 CUL3 complex which is predominantly expressed in invasive EVT. Figures 4a and 4c show genes upregulated in HTR-8/SVneo and SGHPL5 cells, respectively, in comparison with JEG3 cells (Figures 4b and 4d ). The dataset used was GSE20510. Figure 4e shows combined analysis of datasets GSE20510 and GSE28551 involving genes that are upregulated in first trimester placenta and surrogates used in in vitro studies for first-trimester placenta -HTR8/SVneo and SGHPL-5 cell lines, and the human SARS-CoV-2 interactome. STRING (Figure 4f) and GeneMANIA (Figure 4g (Figures 5 a, b, and c) and Gestational Diabetes Mellitus (Figure 5d) . Figure 5e and 5f show GeneMANIA protein interaction networks of MFGE8 and PLAT, respectively. Figure 5g shows STRING protein interaction network of PVR (CD155). The expansions of all the gene names are given in Supplementary Table 2. 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