Email this article ANALYSIS OF CLONAL NK CELL POPULATIONS USING SINGLE-CELL TRANSCRIPTOMICS DATA
- Authors: Ustiuzhanina M.O.1,2,3, Kovalenko E.I.1
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Affiliations:
- Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia
- Center of Life Sciences, Skolkovo Institute of Science and Technology, Moscow, Russia
- Pirogov Russian National Research Medical University, Moscow, Russia
- Section: Forum Sochi 2025
- Submitted: 28.04.2025
- Accepted: 22.06.2025
- URL: https://rusimmun.ru/jour/article/view/17245
- DOI: https://doi.org/10.46235/1028-7221-17245-AOC
- ID: 17245
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Abstract
Abstract
Background. Natural killer (NK) cells represent a critical component of antiviral immunity, demonstrating remarkable adaptability during infections such as human cytomegalovirus (hCMV) and COVID-19. Recent advances in single-cell transcriptomics have uncovered the existence of clonally expanded NK cell populations with distinct functional profiles, blurring the traditional boundaries between innate and adaptive immunity. However, the functional heterogeneity and immunological significance of these clones remain incompletely understood.
Aim. This study aimed to dissect clonal NK cell heterogeneity using published scRNA-seq datasets from hCMV-seropositive, seronegative, and COVID-19 patients, focusing on cluster-specific gene expression patterns.
Methods. Our computational pipeline employed Seurat-based integration and high-resolution clustering of datasets from hCMV-seropositive (n=5) and seronegative (n=2) donors, along with COVID-19 patients (n=2). We analyzed datasets using Seurat 5 in R. Quality-controlled data were normalized (SCTransform), integrated (batch-corrected), and clustered (UMAP). Differential gene expression (Wilcoxon test, log2FC > 0.25, p-adj < 0.05) and annotation were performed.
Results. In hCMV-seropositive individuals, we identified 12 transcriptionally distinct NK cell clusters exhibiting KLRC2 (NKG2C)-dependent organization, with specific clones showing either enhanced cytotoxic potential (marked by GZMB/GZMA upregulation) or unique inhibitory receptor profiles (variable KIR expression patterns). The hCMV-seronegative cohort displayed a simpler clonal structure with 9 clusters showing reduced KIR diversity but maintained distinct effector gene signatures. Analysis of COVID-19 patients revealed divergent clonal patterns: one patient showed reduced KLRC2 variability with prominent KLRC1 (NKG2A) expression, while another exhibited KIR heterogeneity without KLRC2 variation.
Conclusion. Our analysis reveals distinct clonal NK cell populations in hCMV and COVID-19 contexts, characterized by divergent expression of activating and inhibitory receptors. These findings demonstrate infection-specific dynamics of clonal NK cell populations, highlighting their adaptive potential through differential receptor expression in antiviral responses.
About the authors
Maria O. Ustiuzhanina
Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia;Center of Life Sciences, Skolkovo Institute of Science and Technology, Moscow, Russia;
Pirogov Russian National Research Medical University, Moscow, Russia
Email: mashaust1397@gmail.com
ORCID iD: 0000-0003-3378-6508
SPIN-code: 5451-6672
Scopus Author ID: 57224919329
ResearcherId: ABG-4443-2021
Аспирант, м.н.с. лаборатория методов иммуносеквенирования
Russian Federation, 117997, улица Миклухо-Маклая, дом 16/10; 121205, Большой бульвар д. 30, стр. 1; 117513, ул. Островитянова д. 1E. I. Kovalenko
Shemyakin & Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Moscow, Russia
Author for correspondence.
Email: lenkovalen@mail.ru
ORCID iD: 0000-0001-8119-8247
PhD, Senior Researcher of Laboratory of Cell Interactions
Russian Federation, 117997, 16/10 Miklukho-Maklaya str.References
- Béziat V, Liu LL, Malmberg JA, Ivarsson MA, Sohlberg E, Björklund AT, Retière C, Sverremark-Ekström E, Traherne J, Ljungman P, Schaffer M, Price DA, Trowsdale J, Michaëlsson J, Ljunggren HG, Malmberg KJ. 2013. NK cell responses to cytomegalovirus infection lead to stable imprints in the human KIR repertoire and involve activating KIRs. Blood. 121,2678–2688.
- Grassmann S, Pachmayr LO, Leube J, Mihatsch L, Andrae I, Flommersfeld S, Oduro J, Cicin-Sain L, Schiemann M, Flossdorf M, Buchholz VR. 2019. Distinct Surface Expression of Activating Receptor Ly49H Drives Differential Expansion of NK Cell Clones upon Murine Cytomegalovirus Infection. Immunity. 50,1391-1400.e4.
- Gumá M, Budt M, Sáez A, Brckalo T, Hengel H, Angulo A, López-Botet M. Expansion of CD94/NKG2C+ NK cells in response to human cytomegalovirus-infected fibroblasts. Blood. 2006 May 1;107(9):3624-31.
- Ludwig LS, Lareau CA, Ulirsch JC, Christian E, Muus C, Li LH, Pelka K, Ge W, Oren Y, Brack A, Law T, Rodman C, Chen JH, Boland GM, Hacohen N, Rozenblatt-Rosen O, Aryee MJ, Buenrostro JD, Regev A, Sankaran VG. 2019. Lineage Tracing in Humans Enabled by Mitochondrial Mutations and Single-Cell Genomics. Cell. 176,1325-1339.e22.
- Maucourant C, Filipovic I, Ponzetta A, Aleman S, Cornillet M, Hertwig L, Strunz B, Lentini A, Reinius B, Brownlie D, Cuapio A, Ask EH, Hull RM, Haroun-Izquierdo A, Schaffer M, Klingström J, Folkesson E, Buggert M, Sandberg JK, Eriksson LI, Rooyackers O, Ljunggren HG, Malmberg KJ, Michaëlsson J, Marquardt N, Hammer Q, Strålin K, Björkström NK. 2020. Natural killer cell immunotypes related to COVID-19 disease severity. Sci Immunol. 5,6832.
- Rückert T, Lareau CA, Mashreghi MF, Ludwig LS, Romagnani C. 2022. Clonal expansion and epigenetic inheritance of long-lasting NK cell memory. Nature Immunology 2022 23:11. 23,1551–1563.
- Ustiuzhanina MO, Streltsova MA, Timofeev ND, Kryukov MA, Chudakov DM, Kovalenko EI. 2024. Autologous T-Cell-Free Antigen Presentation System Unveils hCMV-Specific NK Cell Response. Cells 2024, Vol 13, Page 530. 13,530.
- Ustiuzhanina MO, Vavilova JD, Boyko AA, Streltsova MA, Kust SA, Kanevskiy LM, Sapozhnikov AM, Iskhakov RN, Gubernatorova EO, Drutskaya MS, Bychinin M V., Zhukova OA, Novikova ON, Sotnikova AG, Yusubalieva GM, Baklaushev VP, Kovalenko EI. 2023. Coordinated Loss and Acquisition of NK Cell Surface Markers Accompanied by Generalized Cytokine Dysregulation in COVID-19. International Journal of Molecular Sciences 2023, Vol 24, Page 1996. 24,1996.
- Ustiuzhanina MO, Boyko AA, Vavilova JD, Siniavin AE, Streltsova MA, Astrakhantseva I V., Drutskaya MS, Chudakov DM, Kovalenko EI. 2024. The Antigen-Specific Response of NK Cells to SARS-CoV-2 Correlates With KIR2DS4 Expression. J Med Virol. 96.
- Varchetta S, Mele D, Oliviero B, Mantovani S, Ludovisi S, Cerino A, Bruno R, Castelli A, Mosconi M, Vecchia M, Roda S, Sachs M, Klersy C, Mondelli MU. 2021. Unique immunological profile in patients with COVID-19. Cell Mol Immunol. 18,604.
- Witkowski M, Tizian C, Ferreira-Gomes M, Niemeyer D, Jones TC, Heinrich F, Frischbutter S, Angermair S, Hohnstein T, Mattiola I, Nawrath P, Mc Ewen S, Zocche S, Viviano E, Heinz GA, Maurer M, Kölsch U, Chua RL, Aschman T, Meisel C, Radke J, Sawitzki B, Roehmel J, Allers K, Moos V, Schneider T, Hanitsch L, Mall MA, Conrad C, Radbruch H, Duerr CU, Trapani JA, Marcenaro E, Kallinich T, Corman VM, Kurth F, Sander LE, Drosten C, Treskatsch S, Durek P, Kruglov A, Radbruch A, Mashreghi MF, Diefenbach A. 2021. Untimely TGFβ responses in COVID-19 limit antiviral functions of NK cells. Nature. 600,295–301.
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СМИ зарегистрировано Федеральной службой по надзору в сфере связи, информационных технологий и массовых коммуникаций (Роскомнадзор).