Mechanism of Iron Uptake and Efflux
The Tf-TfR pathway delivers transferrin iron into cells solely via clathrin- pit-mediated endocytosis. Distinct from this, GAPDH-mediated trafficking also involves two other mechanisms: lipid-raft endocytosis and micropinocytosis (Kumar et al. 2011). The affinity of interaction for Tf-TfR1 and Tf-TfR2 is extremely high (KD = 1.0 nM and 25 nM, respectively) as compared to the lower affinity for Tf-GAPDH (KD = 120 nM in vitro and 60 nM in vivo) (Kawabata et al. 1999; Raje et al. 2007). In addition, GAPDH has also been identified to play a role in transferrin-mediated efflux of iron from macrophages. In this role, a distinct form of GAPDH is re-localized to the surface of cells which captures apo- transferrin (KD 1.11 nM) in close proximity to the iron transporter ferroportin to remove excess iron from cells (Sheokand et al. 2014).
Role of Post-Translational Modifications
Numerous reports have indicated that GAPDH can switch between its alternate functions depending on the pattern of multiple post-translational modifications (Seidler 2013; Sirover 2014). Recent studies have indicated that post-translational modifications are responsible for altered membrane localization and transferrinbinding ability of GAPDH in response to cellular iron levels (Sheokand et al. 2014). The discovery of differential binding by GAPDH to either holo or apo forms of transferrin during conditions of iron depletion or excess, respectively, has shown that this switch in ligand specificity occurs due to the presence of different isoforms of GAPDH. An abundance of post-translational modifications (PTMs) including oxidation, dimethylation, acetylation, nitrosylation, and phosphorylation were evident in membrane GAPDH from iron-depleted cells as compared to GAPDH isolated from iron-loaded cells (Sheokand et al. 2014). These results indicate that, in addition to post-transcriptional regulation, posttranslational modifications are an important regulatory mechanism for proteins involved in iron homeostasis.