Programmed Death Ligand 2 (PD-L2) is a co-stimulatory molecule predominantly expressed on antigen-presenting cells. PD-L2 interacts with membrane-bound receptor PD-1 on activated T-cells. This interaction transmits a negative signal to T-cells inducing anergy or apoptosis. Viruses such as HIV induce PD-L2, which enables viral persistence. In this study, regulation of PD-L2 gene expression in feline lymphocytes in homeostasis and lentiviral infection was investigated. The upstream region of PD-L2 was amplified and sequenced to identify putative transcription factor binding sites, including NF-κB, STAT6, Oct-1/2, GATA-3 and NFAT. Assays to assess transcriptional activation from incremental segments upstream of PD-L2 identified the first 2 kb as the possible location of the proximal promoter. Transfection of NF-κB or NFAT into T-cells increased PD-L2 transcription, but FIV infection did not. Results suggest that transcriptional activation of PD-L2 in homeostatic T-cells differs from macrophages and DCs; and that acute FIV infection does not affect PD-L2 expression.