Iron has a fundamental role in many metabolic processes, including electron transport, deoxyribonucleotide synthesis, oxygen transport and many essential redox reactions involving haemoproteins and Fe-S cluster proteins. Defective iron homeostasis results in either iron deficiency or iron overload. Precise regulation of iron transport in mitochondria is essential for haem biosynthesis, haemoglobin production and Fe-S cluster protein assembly during red cell development. Here we describe a zebrafish mutant, frascati (frs), that shows profound hypochromic anaemia and erythroid maturation arrest owing to defects in mitochondrial iron uptake. Through positional cloning, we show that the gene mutated in the frs mutant is a member of the vertebrate mitochondrial solute carrier family (SLC25) that we call mitoferrin (mfrn). mfrn is highly expressed in fetal and adult haematopoietic tissues of zebrafish and mouse. Erythroblasts generated from murine embryonic stem cells null for Mfrn (also known as Slc25a37) show maturation arrest with severely impaired incorporation of 55Fe into haem. Disruption of the yeast mfrn orthologues, MRS3 and MRS4, causes defects in iron metabolism and mitochondrial Fe-S cluster biogenesis. Murine Mfrn rescues the defects in frs zebrafish, and zebrafish mfrn complements the yeast mutant, indicating that the function of the gene may be highly conserved. Our data show that mfrn functions as the principal mitochondrial iron importer essential for haem biosynthesis in vertebrate erythroblasts.

Hypoxia-inducible factor-1 (HIF-1) is a transcription factor activated by hypoxia. The HIF-1 activation transduction pathway is poorly understood. In this report, we investigated the activation of extracellular regulated kinases (ERK) in hypoxia and their involvement in HIF-1 activation. We demonstrated that in human microvascular endothelial cells-1 (HMEC-1), ERK kinases are activated during hypoxia. Using dominant negative mutants, we showed that ERK1 is needed for hypoxia-induced HIF-1 transactivation activity. Moreover, using a kinase assay and Western blot experiments, we showed that HIF-1K K is phosphorylated in hypoxia by an ERK-dependent pathway. These results evidence the role of mitogen-activated protein kinase in the transcriptional response to hypoxia.z 2000 Federation of European Biochemical Societies.

The protein chaperone heat shock protein 90 (Hsp90) is a major regulator of different transcription factors such as MyoD, a basic helix loop helix (bHLH) protein, and the bHLHPer-aryl hydrocarbon nuclear translocator (ARNT)-Sim (PAS) factors Sim and aryl hydrocarbon receptor (Ahr). The transcription factor hypoxia-inducible factor-1K K (HIF-1K K), involved in the response to hypoxia, also belongs to the bHLH-PAS family. This work was aimed to investigate the putative role of Hsp90 in HIF-1 activation by hypoxia. Using a EGFP-HIF-1K K fusion protein, co-immunoprecipitation experiments evidenced that the chimeric protein expressed in COS-7 cells interacts with Hsp90 in normoxia but not in hypoxia. We also demonstrated that Hsp90 interacts with the bHLH-PAS domain of HIF-1K K. Moreover, Hsp90 is not co-translocated with HIF-1K K into the nucleus. At last, we showed that Hsp90 activity is essential for HIF-1 activation in hypoxia since it is inhibited in the presence of geldanamycin. These results indicate that Hsp90 is a major regulator in HIF-1K K activation.z 1999 Federation of European Biochemical Societies.

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