Stochastic Expansions Maintain the Clonal Stability of CD8+ T Cell Populations Undergoing Memory Inflation Driven by Murine Cytomegalovirus

Abstract

CMV is an obligate and persistent intracellular pathogen that continually drives the production of highly differentiated virus-specific CD8+ T cells in an antigen-dependent manner, a phenomenon known as memory inflation. Extensive proliferation is required to generate and maintain inflationary CD8+ T cell populations, which are counterintuitively short-lived and typically exposed to limited amounts of antigen during the chronic phase of infection. An apparent discrepancy therefore exists between the magnitude of expansion and the requirement for ongoing immunogenic stimulation. To address this issue, we explored the clonal dynamics of memory inflation. First, we tracked congenically marked OT-I cell populations in recipient mice infected with murine CMV (MCMV) expressing the cognate antigen OVA. Irrespective of numerical dominance, stochastic expansions were observed in each population, such that dominant and subdominant OT-I cells were maintained at stable frequencies over time. Second, we characterized endogenous CD8+ T cell populations specific for two classic inflationary epitopes, M38 and IE3. Multiple clonotypes simultaneously underwent antigen-driven proliferation during latent infection with MCMV. In addition, the corresponding CD8+ T cell repertoires were stable over time and dominated by persistent clonotypes, many of which also occurred in more than one mouse. Collectively, these data suggest that stochastic encounters with antigen occur frequently enough to maintain oligoclonal populations of inflationary CD8+ T cells, despite intrinsic constraints on epitope display at individual sites of infection with MCMV.

The CMV-Specific CD8+ T Cell Response Is Dominated by Supra-Public Clonotypes with High Generation Probabilities

“…We observed that public clonotypes constituted the majority of expanded clones despite accounting only for a minority of unique CDR3 sequences, which is in line with previous reports investigating this aspect for IE3- and M38-specific T cell responses at late time points of infection [ 25 ]. Beyond mere classification into private vs. public, the degree of sharedness can be highly heterogenous.…”

“…Overall, here we show that generation probability is a major determinant of the T cell response to SIINFEKL-expressing CMV. The results are consistent with so far described T cell responses against other natural CMV epitopes [ 25 ], arguing against a special intrinsic feature of SIINFEKL to induce (supra-)public T cell responses. Instead, the fact that similarly public responses are observed towards natural CMV epitopes points into the direction that the public nature of the T cell response could be due to the mode of antigen presentation rather than the epitope itself.…”

“…Next, we investigated whether public clones are also preferentially recruited into the immune response. While evidence for this has been collected before, previous analyses were mostly confined to individual time points and investigated only private vs. public clones in general [ 15 , 16 , 25 ], thereby not incorporating the resolution of differential degrees of sharedness over the entire course of an immune response. Since the CDR3β repertoire is more diverse than the CDR3α repertoire and at the same time less prone to bias through second chains from non-excluded alleles, we focused our analyses from hereon on CDR3β sequences.…”

“…Further, it is unclear how dominant such public clonotypes are within CMV-specific immune responses. A recent study showed that public clonotypes are dominant in inflationary CMV-specific T cell populations, but this work only investigated very late time points of infection, did not take into account different degrees of sharedness and calculated generation probability merely by distance from germline [ 25 ]. In recent years, computation of generation probabilities became significantly more refined and can now also take into account biochemical features of CDR3 sequences, such as hydrophobicity of amino acids at certain positions or common motifs [ 26 , 27 , 28 ].…”

Revisiting CD8 T-cell ‘Memory Inflation’: New Insights with Implications for Cytomegaloviruses as Vaccine Vectors

“…As intraplantar infection is closer to a realistic application of CMV vector-based vaccines, we focused our interest on this infection route and studied the dynamics of the CD8 T-cell response in spleen ( Figure 2) and lungs ( Figure 3) in the long-term course, monitoring a large cohort of infected mice by sampling at 3-month intervals. Recent work by Smith and colleagues [47] in a model of systemic infection after intraperitoneal virus application has shown that MI is based on the expansion of CD8 T-cell clonotypes that differ between individual mice, not by statistical error but by biological variance. Specifically, some clonotypes persisted for the entire observation period in all mice tested individually and longitudinally, although with expanding and contracting clone sizes in the time course.…”