the thymus and bone marrow. Regulation of its expression occurs via multiple pathways. These include protein-protein interactions, like those with TdIF1. TdIF1 is another protein that interacts with TdT to inhibit its function by masking the DNA binding region of the TdT polymerase. The regulation of TdT expression also exists at the transcriptional level, with regulation influenced by stage-specific factors, and occurs in a developmentally restrictive manner. Although expression is typically found to be in the primary lymphoid organs, recent work has suggested that stimulation via antigen can result in secondary TdT expression along with other enzymes needed for gene rearrangement outside of the thymus for T-cells. Patients with acute lymphoblastic leukemia greatly over-produce TdT. Cell lines derived from these patients served as one of the first sources of pure TdT and lead to the discovery that differences in activity exist between human and bovine isoforms. Mechanism Similar to many polymerases, the catalytic site of TdT has two divalent cations in its palm domain that assist in nucleotide binding, help lower the pKa of the 3'-OH group and ultimately facilitate the departure of the resultant pyrophosphate by-product. Isoform Variation Several isoforms of TdT have been observed in mice, bovines, and humans. To date, two variants have been identified in mice while three have been identified in humans. The two splice variants identified in mice are named according to their respective lengths: TdTS consists of 509 amino acids while TdTL, the longer variant, consists of 529 amino acids. The differences between TdTS and TdTL occur outside regions that bind DNA and nucleotides. That the 20 amino acid difference affects enzymatic activity is controversial, with some arguing that TdTL's modifications bestow exonuclease activity while others argue that TdTL and TdTS have nearly identical in vitro activity. Additionally, TdTL reportedly can modulate the catalytic activity of TdTS in vivo through an unknown mechanism. It is suggested that this aids in the regulation of TdT's role in V(D)J recombination. Human TdT isoforms have three variants TdTL1, TdTL2, and TdTS. TdTL1 is broadly expressed in lymphoid cell lines while TdTL2 is predominantly expressed in normal small lymphocytes. Both localize in the nucleus when expressed and both possess 3'->5' exonuclease activity. In contrast, TdTS isoforms do not possess exonuclease activity and perform the necessary elongation during V(D)J recombination. Since a similar exonuclease activity hypothesized in murine TdTL is found in human and bovine TdTL, some postulate