Comment on: “Mapping Systemic Lupus Erythematosus heterogeneity at the single-cell level” Nehar-Belaid, D., et al. Nat Immunol Sept 2020. https://doi.org/10.1038/s41590-020-0743-0
Commented by: Marta E. Alarcón Riquelme, Head of Medical Genomics Coordinator of 3TR, GENYO. Centre for Genomics and Oncological Research: Pfizer / University of Granada/ Andalusian Regional Government, Spain
Technology has come to a point that we can assess the transcriptome of a single cell. In SLE, several studies have been published analyzing the transcriptome of single cells in tissues 1,2. These studies allow to define exactly in which cells, the abnormalities reside and possibly the unique cell mechanisms behind the abnormalities we observe in SLE. The paper by Nehar-Belaid and Hong (contributed equally)3 describes the analysis of peripheral blood mononuclear cells, and the results do give some surprises. The type I interferon signature is widely expressed, but interestingly, the interferon signature was observed in defined subpopulations of the major cell types, including plasma cells. These analyses require clustering of the cell populations into genes expressed in such a way that differentiate these, and sometimes identify what appears to be, new populations of cells. The groups or clusters hence identified, are then named somewhat arbitrarily.
The cells primarily expressing the interferon signature genes were: the major monocyte cluster and all seven minor clusters, a minor CD4+ T cell cluster; CD16+ monocytes, megakaryocytes, cDCs, plasma cells, pDCs, and an ISGhiGzK+acCD8+ T cells. Some genes allowed to group the patients into 9 groups called G1-G9: G1 was the group where type I interferon genes were most upregulated and were featured by ISG15 and IFI27; G2 by IFI35 and ADAR; G3 by IFI44L and PAPR9; G4 by TMEM140 mainly upregulated in megakaryocytes; the G5 genes were IFNGR1, CASP1 and FCGRIA, which were primarily expressed in monocytes, DCs ,and megakaryocytes; G6 genes were CCL4, CCL5 and IFNG and were upregulated in NK cells, GzH_acCD8+ T and ISGhi_GzK+_acCD8+ T cells; G7 had primarily SOCS1 upregulated and found in pDCs but also in subsets or clusters of T cells; G8 included IRF7 and IFNLR1 found in plasma cells in SLE patients, and G9 genes had the IFNAR genes found in PCs.
Many of the populations showed similar upregulation in patients and controls, therefore revealing how these genes are normally expressed in specific cell types, but others were clearly upregulated in SLE and cell subsets were expanded. The authors analyzed genes that co-expressed within cell populations with the major genes defining the clusters and found that monocytes with high interferon signature genes also co-expressed IL1B. Cells co-expressing IL1B still could be divided into three signatures of cells: one cell group co-expressing IFITM3 and IL1B, another ISG15 and IL1B and a third one, had both of them (10% of the cells).
Each of the major cell populations were finely characterized as well as signatures most associated with disease activity. For instance, ISG15 was overexpressed in CD14+ monocytes in patients with high disease activity. Interestingly, ISG15 expression was restricted to a B cell subpopulation that the authors named as B-SC5, which was the only cell type expressing TBX21 (the transcription factor T-bet). This with other genes expressed exclusively in this cell type suggested that these corresponded to the ones previously described as extrafollicular, autoreactive, also called DN2 B cells4. Those genes were ITGAX (CD11c), FGR, TFEC, FCRL2, FCRL3 and FCRL5, and IL10RA.
One type of pDC, named PDC-SC1 was expanded in SLE and contributed to the interferon signature. On the other hand, only one DC population was expanded in SLE and did not show any disease activity difference. The expanded cell cluster was named DC-SC2, which corresponded to the AXL+SIGLEC6+ DCs, the main DC subset expressing interferon signature genes. Finally, the T cell subsets mainly expressing interferon signature genes were the cytotoxic T cells, named T-SC4 cells. These cells were particularly expanded in SLE and included CD4+T cells that mapped to the ISGhiC11_CD4+T cluster, in turn expanded in SLE.
A specific cluster of NK cells, which was ISGhi was also specific for SLE. Two NK clusters, NK-SC2 and NK-SC3 were expanded in SLE and both had upregulated interferon and cytotoxicity-encoding genes with a particular increase of ISG15 and CD56 in the NK-SC2 subset.
Because the cohort used for the single cell study comprised children with SLE, the authors also analyzed the expression of genes mutated in SLE-related monogenic diseases, such as complement components C1Q, C1R or C2, TREX1 or DNASE1L3, among others. The authors found that most of these genes with the exception of the complement genes, were expressed primarily in pDCs and DCs, but also in plasma cells, the B-SC5 subset, and the NK-SC3 subset.
Finally, the authors used the single cell abundance to classify SLE patients into groups, validating the groups in a second adult cohort revealing similar expansions of cellular subpopulations that had association with disease severity.
- Arazi, A. et al. The immune cell landscape in kidneys of patients with lupus nephritis. Nat Immunol 20, 902-914 (2019).
- Der, E. et al. Tubular cell and keratinocyte single-cell transcriptomics applied to lupus nephritis reveal type I IFN and fibrosis relevant pathways. Nat Immunol 20, 915-927 (2019).
- Nehar-Belaid D, H.S., Marches R, Chen G, Bolisetty M, Baisch J, Walters L, Punaro M, Rossi RJ, Chung CH, Huynh RP, Singh P, Flynn WF, Tabanor-Gayle JA, Kuchipudi N, Mejias A, Collet MA, Lucido AL, Palucka K, Robson P, Lakshminarayanan S, Ramilo O, Wright T, Pascual V, Banchereau JF. Mapping systemic lupus erythematosus heterogeneity at the single-cell level. Nat Immunol 21, 1094-1106 (2020).
- Jenks, S.A. et al. Distinct Effector B Cells Induced by Unregulated Toll-like Receptor 7 Contribute to Pathogenic Responses in Systemic Lupus Erythematosus. Immunity 49, 725-739 e6 (2018).