D-Dopachrome tautomerase (D-DT) : functional homologue or cross-regulator of macrophage migration inhibitory factor (MIF)?
Year: 2011
Type: dissertation
Abstract: Macrophage migration inhibitory factor (MIF) is a pleiotropic immunoregulatory and pro-inflammatory cytokine, which plays a pivotal role in the pathogenesis of various acute and chronic inflammatory diseases. For example, endotoxemia studies in mice reveal that MIF administration decreases the survival rate, whereas MIF neutralization or genetic deletion of Mif improve the chances for a beneficial outcome. Additionally, high MIF levels are measured in a variety of human tumors and functional studies reveal MIF involvement in multiple aspects of tumor progression including control of cell proliferation and prevention of apoptosis. Unlike other cytokines, MIF also is a phenylpyruvate tautomerase that converts two non-physiologic enzymatic substrates, D-dopachrome and p-hydroxyphenylpyruvate (HPP). To date, possible relationships between the tautomerase and inflammatory or tumor-supportive activities of MIF remain unclear. In 1993, D-dopachrome tautomerase (D-DT) was discovered during an investigation of melanogenesis. The tertiary structure of D-DT is remarkably similar to MIF, identifying D-DT as the only known eukaryotic MIF homologue discovered to date. D-DT shares MIF’s tautomerase activity and, thus, both proteins are members of the MIF tautomerase superfamily. Despite D-DT’s intriguing similarities to the very well studied MIF, there have been very few reports on the biological functions and mechanisms of action of D-DT. This thesis aimed at the functional characterization of D-DT in biological and pathophysiologic conditions. Prior to apoptotic studies, D-DT cDNA was cloned into a mammalian expression vector, cells were transfected with the newly generated plasmid and protein overexpression confirmed. Successive data demonstrated that D-DT did not obtain anti-apoptotic effects upon nitric oxide-induced apoptosis in macrophages. For the first time, this suggested contrary effects of D-DT in comparison to MIF. In a subsequent cell survival study of D-DT it was illustrated that D-DT enhanced macrophage viability to a more pronounced extent than MIF. Interestingly, it was further shown that simultaneous treatment of cells with MIF and D-DT increased macrophage viability even more, which suggested additive effects of both proteins. Additionally, D-DT was characterized as a protein that tautomerizes the substrate HPP and, thus, possessed analog enzymatic activity to MIF. However, quantitative differences in the tautomerization rate became evident between different mammalian homologues of D-DT. The covalent MIF inhibitor 4-IPP also inhibited the tautomerase activity of D-DT, whereas D-DT was not susceptible to ISO-1. This thesis also demonstrated first insight into D-DT’s in vivo role in a murine model of endotoxic shock. Neutralization of D-DT had a protective effect in the LPS-shock model. This finding accorded with results reported for MIF and, therefore, suggested analog functions of D-DT in this context. Taken together, this thesis reports on mostly similar, but also contrary biologic functions of MIF and D-DT. As this study is one of the first approaches to determine D-DT’s role in disease, continuative investigations need to be conducted to further validate these observations. Recent studies showed that the deletion of the MIF receptor, CD74, abrogates downstream MIF effects to a greater extend than MIF deficiency alone. My work also indicates the importance of further studies to determine D-DT’s potency as a MIF receptor ligand.
Source: RWTH Publications (RWTH Aachen)
Authors: Wibke Schulte, Jürgen Bernhagen
Language: English
Subfield: Immunology
Field: Immunology and Microbiology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W1005473919