After antigen encounter, naive lymphocytes differentiate into populations of memory cells that share a common set of functions including faster response to antigen re-exposure and the ability to self-renew. However memory lymphocytes in different lymphocyte lineages are functionally and phenotypically diverse. It is not known whether discrete populations of T and B cells use similar transcriptional programs during differentiation into the memory state. We used cross-species genomic analysis to examine the pattern of genes upregulated during the differentiation of naive lymphocytes into memory cells in multiple populations of human CD4, CD8 and B cell lymphocytes as well as two mouse models of memory development. We identified and validated a signature of genes that was upregulated in memory cells compared to naive cells in both human and mouse CD8 memory differentiation, suggesting marked evolutionary conservation of this transcriptional program. Surprisingly, this conserved CD8 differentiation signature was also upregulated during memory differentiation in CD4 and B cell lineages. To validate the biologic significance of this signature we showed that alterations in this signature of genes could distinguish between functional and exhausted CD8 T cells from a mouse model of chronic viral infection. Finally, we generated genome-wide microarray data from tetramer-sorted human T cells and showed profound differences in this differentiation signature between T cells specific for HIV from those specific for influenza. Thus, our data suggest that in addition to lineage-specific differentiation programs, T and B lymphocytes employ a common transcriptional program during memory development that is disrupted in chronic viral infection.