Asymmetric Cell Division in T Lymphocyte Fate Diversification

Abstract

Immunological protection against microbial pathogens is dependent on robust generation of functionally diverse T lymphocyte subsets. Upon microbial infection, naïve CD4(+) or CD8(+) T lymphocytes can give rise to effector- and memory-fated progeny that together mediate a potent immune response. Recent advances in single-cell immunological and genomic profiling technologies have helped elucidate early and late diversification mechanisms that enable the generation of heterogeneity from single T lymphocytes. We discuss these findings here and argue that one such mechanism, asymmetric cell division, creates an early divergence in T lymphocyte fates by giving rise to daughter cells with a propensity towards the terminally differentiated effector or self-renewing memory lineages, with cell-intrinsic and -extrinsic cues from the microenvironment driving the final maturation steps.

Keywords: T lymphocyte differentiation; T lymphocyte fate determination; adaptive immunity; asymmetric division; single-cell analyses.

Figure 1

T lymphocyte differentiation into diverse fates. (A) Hypothetical model of CD8⁺ T lymphocyte fate specification. Asymmetric division of an activated naïve cell (white) gives rise to pre-effector (teal) and pre-memory (yellow) daughter cells, while environmental cues, such as cytokines and metabolites, may differentially affect progression into the terminal effector (blue) or T_EM (orange) and T_CM (light orange) cell fates, respectively. Dotted lines indicate the proposed ontogeny of long-lived effector (dark blue) and T_RM (red) cells. (B) Hypothetical model of CD4⁺ T lymphocyte fate specification. Differentiation into T_H1 effector (light green) and T_FH (gray) cells diverges early during an immune response to intracellular pathogens, as a possible result of asymmetric division. A subset of T_H1 effector cells survives to form the T_EM pool (dark green), while a subset of T_FH cells survive to form T_CM cells (black). It remains to be determined whether infections by extracellular pathogens induce divergent pathways of differentiation into T_H2 or T_H17 effector cells (blue) and T_FH/T_CM cells. (C) Potential mechanisms regulating establishment and maintenance of asymmetric T lymphocyte division. (i) An antigen-presenting cell provides an initial polarity cue (MHC/peptide and ICAM) to a T lymphocyte through interaction with the TCR and LFA-1 [5, 57], leading to (ii) formation of an immune synapse and establishment of asymmetry. (iii) Critical components of the immune synapse, including PDK1 [88], PI3K [89], and Cdc42 [90], which have been previously shown to be activators of aPKC in adipocytes and Drosophila and C. elegans [–93], may function similarly in T lymphocytes to transmit the initial polarity cue to aPKC. (iv) Activated aPKC, bound with PAR3 and PAR6 [58], can then phosphorylate, and thereby exclude, important fate determinants that enter the distal pole (red) of the T lymphocyte, thus (v) maintaining asymmetry through the completion of division. In Drosophila and C. elegans, the Scribble and PAR complexes have an antagonistic relationship (i.e. aPKC phosphorylates Lgl to induce polarization, whereas Lgl represses aPKC activity), which serves to restrict aPKC activity to one pole of a dividing cell and coordinate the maintenance of asymmetry [94]. It remains to be determined whether a similar antagonistic mechanism functions in T lymphocytes to facilitate (vi) the generation of effector-fated daughter cells, which express IL-2Ra, IFNγR, and T-bet (blue), and memory-fated daughter cells via asymmetric division. Abbreviations: T_EM, effector memory cell; T_CM, central memory cell; T_RM, tissue resident memory cell; T_H, helper T cell; T_FH, follicular helper T cell; TCR, T cell receptor; LFA, lymphocyte function-associated antigen; aPKC, atypical protein kinase C; PDK1, phosphoinositide-dependent kinase-1; PI3K, phosphoinositide-3 kinase; Cdc42, cell division control protein 42; PAR, partitioning defective; Lgl, lethal giant larvae; IL-2Rα, interleukin-2 receptor α; IFNγR, interferon-γ receptor.

Figure 2

Possible approach to study T lymphocyte differentiation combining in vivo fate mapping with single-cell technologies. Microbial infection triggers the adaptive immune response. T lymphocytes are activated and undergo multiple rounds of cell division for clonal expansion and differentiation into effector and memory subsets. (A) Asymmetric division at the first cell division of an activated naïve T cell (white) results in unequal inheritance of a hypothetical fate-associated molecule (small green or pink circles) into one of the two daughter cells (pre-memory cells (yellow); pre-effector cells (teal)). Fusion of the fate-determining molecule to a fluorescent protein could enable the tracking of progenies throughout the immune response, if expression of the fate-determining molecule is maintained and detectable in the progenies of only one daughter cell. (B) Single cells sorted on the basis of fusion fluorescent protein expression at early and late times post-infection, together with cell surface markers that distinguish terminal effector cells (dark blue) at days 7–8 post-infection and memory cell subsets at day 30 post-infection and beyond (long-lived effector (purple), T_CM (light orange), T_EM (orange), T_RM (red)) by flow cytometry. (C) Single-cell technologies enable the interrogation of individual responding T lymphocytes at multiple levels of regulation: RNA, gene expression, epigenetic, cellular metabolism, and protein expression. Combinatorial analyses of these approaches with computational modeling create a global overview of when and how T cell fates are acquired during an immune response. Abbreviations: T_CM, central memory cell; T_EM, effector memory cell; T_RM, tissue resident memory cell; miRNA, microRNA; MARS-Seq, Massively Parallel Single-Cell RNA-Seq; ATAC-Seq, Assay for Transposase-Accessible Chromatin with high-throughput sequencing; ChIP, chromatin immunoprecipitation; Seq, sequencing.