The heme enzyme indoleamine 2,3-dioxygenase 1 (IDO1) catalyses the rate-limiting step of L-tryptophan catabolism along the kynurenine pathway. Expression of IDO1 in antigen-presenting cells (APCs) represents an important immunoregulatory mechanism signaling for immunosuppression during inflammation, infections and cancer [1]. The incorporation of the heme prosthetic group is essential for IDO1 enzyme activity. We examined the role of de novo heme biosynthesis and heme oxygenase-1 (HO-1)-catalysed heme degradation in controlling IDO1 in primary lipopolysaccharide (LPS)-stimulated human monocyte-derived macrophages (MDMs). Increasing intra-cellular heme through the addition of the heme precursor aminolevulinic acid [2] or inhibition of heme biosynthesis with succinylacetone [2], stimulated or inhibited IDO1 activity in LPS-stimulated MDMs, respectively, independent of changes to IDO1 protein levels. Human MDMs constitutively expressed HO-1, which was upregulated by the addition of exogenous hemin. Co-treatment of MDMs with hemin and LPS to simultaneously upregulate HO-1 and IDO1 expression increased IDO1 enzyme activity independent of a role for HO-1. However, pre-treatment with hemin to upregulate HO-1 levels prior to LPS-stimulation significantly decreased IDO1 mRNA and protein expression levels and enzyme activity in MDMs. Hemin pre-treatment partially inhibited LPS-stimulated phosphorylation of Signal transducer and activator of transcription 1 (STAT1) and nuclear factor-κB (NFκB), which are critical for IDO1 transcription. Inhibiting HO-1 expression with small interfering RNA (siRNA) significantly reversed the capacity of hemin pre-treatment to inhibit IDO1 expression and activity in MDMs. This study shows that intracellular heme biosynthesis and HO-1-mediated heme degradation are functionally linked to the control of IDO1 in LPS-stimulated MDMs. Regulatory crosstalk between the heme metabolism and IDO1 has important implications for the immune regulatory actions of APCs during inflammation and disease [1].