Available online 9 May 2023
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Antibodies produced by antibody-secreting plasma cells (ASCs) underlie multiple forms of long-lasting immunity. Here we examined the mechanisms regulating ASC turnover and persistence using a genetic reporter to time-stamp ASCs. This approach revealed ASC lifespans as heterogeneous and falling on a continuum, with only a small fraction surviving for >60days. ASC longevity past 60days was independent of isotype but correlated with a phenotype that developed progressively and ultimately associated with an underlying "long-lived" ASC (LL ASC)-enriched transcriptional program. While some of the differences between LL ASCs and other ASCs appeared to be acquired with age, other features were shared with some younger ASCs, such as high CD138 and CD93. Turnover was unaffected by altered ASC production, arguing against competition for niches as a major driver of turnover. Thus, ASC turnover is set by intrinsic lifespan limits, with steady-state population dynamics governed by niche vacancy rather than displacement.
Antibodies are tantamount to immunity in a variety of settings. Most licensed vaccines, for example, work by generating high titers of pathogen-specific antibodies.1 Within this framework, however, antibody specificities persist in serum above protective thresholds for varying periods of time ranging from months to decades,2,3 presumably reflecting the lifespans of the antibody-secreting plasma cell (ASC) populations generated during the immune responses and responsible for their secretion. Half-life variation with antibody specificity suggests in turn that there may be a deterministic component to ASC lifespan. For example, extrapolating from antibody amounts, human ASC generated against measles virus have the potential to survive≈3,000 years, whereas those against influenza vaccine survive some months in a typical response.2,4,5 Determinism in lifespan heterogeneity may also extend to ASC phenotype, with B220+ ASCs in mice turning over more rapidly than putative "long-lived" CD19lo and B220neg ASCs.6,7,8 In humans, evidence that CD19lo ASCs are long-lived includes representation of early-life immunization specificities in healthy adults and donor ASCs with this phenotype persisting in transplant recipients >1 year after engraftment.9,10,11 However, it is not clear if this reflects discrete states of what could be called "short-lived" and long-lived ASCs, or if, rather, these are selected examples of the possible lifespans ASCs may have. Understanding the basis of ASC lifespans is clearly relevant to understanding and potentially regulating the persistence of antibody specificities.
ASCs are thought to reside in specialized survival niches, a concept proposed by MacLennan and colleagues, who noted a density of ASCs within a narrow range, above which ASCs were not maintained over time,12 suggesting a limit to the density of ASCs that could be supported by a tissue. This led to the theory of competition for niches as the driving force for ASC turnover.13 To accommodate ASC turnover at homeostasis, it is proposed that existing and nascent ASCs compete directly for survival and localization stimuli, which will at some frequency result in displacement and thus the death of existing ASCs.13,14 A tenet of this argument is that, in an infection, only a small number of ASCs of any given specificity will be displaced, such that it will take many bouts of displacement and replacement to reduce the starting number of antigen-specific ASCs by any reasonable amount.13,15 In this model, all niches can be considered equivalent and sufficient to support ASC survival, while ASCs can be considered as equally displaceable to each other, rather than operating on a continuum of potential residence times. While helpful as a rationale to explain ASC homeostasis, this mechanism has yet to be formally tested, and in its current form does not explain differences in survival based on specificity or on niche variability.16
To investigate ASC lifespan in greater detail, we generated a time-stamp system in mice. ASC longevity past 60days was independent of isotype but correlated with a phenotype that developed progressively and ultimately associated with an underlying long-lived ASC (LL ASC)-enriched transcriptional program. Manipulating the production of ASCs up and down indicated that the dominant form of regulation of homeostatic ASC turnover was independent of competition, fitting with the idea that ASC turnover initiated with cell death rather than displacement. While some of the differences between LL ASCs and other ASCs appeared to be acquired with age, others (such as high CD138 and CD93) appeared to be inborn, giving insight into the regulation of ASC turnover and establishment of population structure. Collectively these findings have implications for tailoring ASC persistence for specific immunological outcomes including the maintenance of long-term immunity.
Resolution of time-stamped ASC in BLTcre.Mcl1fl/+ mice
To study ASC turnover, we generated the BLIMP1-TdTomato inducible-Cre (BLTcre) mouse in which TdTomato (TDT) was placed downstream of the Prdm1 locus, resulting in TDT expression in cells transcribing Prdm1.17 Prdm1 encodes Blimp1, a transcription factor required for ASC development.18,19 That TDTbright cells in BLTcre mice were ASCs was confirmed by their universal expression of CD98 and CD138 (Figure1A) and by high amounts of intracellular immunoglobulin (Ig) light chains, Igλ or Igκ (
Understanding the determinants of ASC persistence is important to developing the means of controlling antibody production in the critical areas of vaccination and autoimmune diseases. Previous analysis of ASC lifespan has focused on measuring antibody specificities in serum,2 characterizing subpopulations of ASCs in bone marrow,6,9,10,40 modulating factors that influence invitro or invivo ASC survival,41,42,43 and nonspecifically tracing turnover after immunization.30 These studies have
Key resources table
REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies anti-BrdU-Fitc (3D4) BD Pharmingen Cat# 560209; RRID: AB_1645615 hCD4-A700 (A161A1) Biolegend Cat#357418; RRID: AB_2616933 hCD4-biotin (OKT4) Walter& Eliza Hall Antibody Facility Cat#(hCD4-bt); RRID: CVCL_2666 CD98 (RL388) Biolegend Cat#128202; RRID: AB_1134242 CD20 (5D2) Genentech Cat#Genentech_5D2; RRID: AB_2936925 CD40L (MR1) Walter& Eliza Hall Antibody Facility Cat#MR1; RRID: CVCL+8964 Chemicals, peptides, and recombinant proteins NP-Osu Biosearch Technologies
We thank Genentech for providing the 5D2 antibody. We thank Rory Markovic, Debbie Ramsey, and the Monash Animal Research Platform for animal welfare, husbandry, and housing assistance. We thank Steve Rockman for support with RNA-seq lab work and Celine Pattaroni and the Monash Bioinformatics Platform for bioinformatics support, advice, and access to Limma, Voom, and Degust. We thank Carole Ford and BD Biosciences for assistance with single-cell sequencing and Alfred Research Alliance FlowCore
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APRIL is critical for plasmablast survival in the bone marrow and poorly expressed by early-life bone marrow stromal cells
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miR-148a controls metabolic programming and survival of mature CD19-negative plasma cells in mice
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IRF4 activity is required in established plasma cells to regulate gene transcription and mitochondrial homeostasis
Blimp-1 controls plasma cell function through the regulation of immunoglobulin secretion and the unfolded protein response
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Mcl-1 is essential for germinal center formation and B cell memory
Long-lived plasma cells accumulate in the bone marrow at a constant rate from early in an immune response
Mcl-1 is essential for the survival of plasma cells
Genetic timestamping of plasma cells invivo reveals tissue-specific homeostatic population turnover
Modelling cell lifespan and proliferation: is likelihood to die or to divide independent of age?
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These authors contributed equally
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