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Haem homeostasis is regulated by the conserved and concerted functions of HRG-1 proteins

Nature volume 453, pages 1127–1131 (2008)Cite this article

Abstract

Haems are metalloporphyrins that serve as prosthetic groups for various biological processes including respiration, gas sensing, xenobiotic detoxification, cell differentiation, circadian clock control, metabolic reprogramming and microRNA processing1,2,3,4. With a few exceptions, haem is synthesized by a multistep biosynthetic pathway comprising defined intermediates that are highly conserved throughout evolution5. Despite our extensive knowledge of haem biosynthesis and degradation, the cellular pathways and molecules that mediate intracellular haem trafficking are unknown. The experimental setback in identifying haem trafficking pathways has been the inability to dissociate the highly regulated cellular synthesis and degradation of haem from intracellular trafficking events6. Caenorhabditis elegans and related helminths are natural haem auxotrophs that acquire environmental haem for incorporation into haemoproteins, which have vertebrate orthologues7. Here we show, by exploiting this auxotrophy to identify HRG-1 proteins in C. elegans, that these proteins are essential for haem homeostasis and normal development in worms and vertebrates. Depletion of hrg-1, or its paralogue hrg-4, in worms results in the disruption of organismal haem sensing and an abnormal response to haem analogues. HRG-1 and HRG-4 are previously unknown transmembrane proteins, which reside in distinct intracellular compartments. Transient knockdown of hrg-1 in zebrafish leads to hydrocephalus, yolk tube malformations and, most strikingly, profound defects in erythropoiesis—phenotypes that are fully rescued by worm HRG-1. Human and worm proteins localize together, and bind and transport haem, thus establishing an evolutionarily conserved function for HRG-1. These findings reveal conserved pathways for cellular haem trafficking in animals that define the model for eukaryotic haem transport. Thus, uncovering the mechanisms of haem transport in C. elegans may provide insights into human disorders of haem metabolism and reveal new drug targets for developing anthelminthics to combat worm infestations.

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Figure 1: Identification of hrg-1 and hrg-4 in C. elegans.
Figure 2: hrg-1 and hrg-4 are essential for haem homeostasis in C. elegans.
Figure 3: HRG-1 is essential for erythropoiesis and development in zebrafish.
Figure 4: Expression, localization and functional studies of worm and mammalian hrg-1.

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Gene Expression Omnibus

Data deposits

The microarray data have been deposited with the Gene Expression Omnibus at NCBI under accession number GSE8696.

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Acknowledgements

The Hamza laboratory thanks the DC-Baltimore WormClub for advice and criticism. We also thank P. Aplan, H. Dailey, I. Mather and S. Severance for critical discussions and reading of the manuscript; M. Cam and G. Poy for expertise with microarrays; J. Lippincott-Schwartz and D. Hailey for organelle markers; H.-F. Lin and R. Handin for the Tg(CD41–GFP) transgenic zebrafish line; J. Italiano for use of the Orca IIER charge-coupled device camera/Metamorph software; P. Krieg for the pT7TS Xenopus oocyte expression vector; P. Ponka and R. Eisenstein for the SIH iron chelation; D. Beckett for use of the fluorescent spectrophotometer; and M. Petris for the hZIP4 plasmid. Many of the worm strains were provided by the Caenorhabditis Genetics Center. This work was supported by funding from the National Institutes of Health (NIH) (I.H., M.D.F. and B.H.P.), the March of Dimes Birth Defects Foundation (I.H. and B.H.P.), the NIH/National Institute of Diabetes and Digestive and Kidney Diseases Intramural Research Program (M.K.), Council for Scientific and Industrial Research and Kanwal Rekhi Fellowships (S.K.U.), and a Howard Hughes Medical Institute Undergraduate Science Education Program grant (S.U.).

Author Contributions Experimental design and execution were as follows: worm experiments and microarrays, A.R., A.U.R., M.K. and I.H.; mammalian experiments, A.R., A.U.R., M.T., C.H., S.U., M.D.F. and I.H.; zebrafish experiments, J.A. and B.H.P.; Xenopus injections and measurements, S.K.U. and M.K.M. I.H. wrote the manuscript. All authors discussed the results and commented on the manuscript.

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Authors and Affiliations

  1. Department of Animal & Avian Sciences and Department of Cell Biology & Molecular Genetics, University of Maryland, College Park, Maryland 20742, USA,

    Abbhirami Rajagopal, Anita U. Rao, Caitlin Hall, Suji Uhm & Iqbal Hamza

  2. Division of Hematology, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA,

    Julio Amigo & Barry H. Paw

  3. Department of Pathology, Children’s Hospital Boston, Boston, Massachusetts 02115, USA,

    Meng Tian & Mark D. Fleming

  4. National Centre for Biological Sciences, Tata Institute of Fundamental Research, UAS-GKVK campus, Bangalore 560 065, India

    Sanjeev K. Upadhyay & M. K. Mathew

  5. Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA,

    Michael Krause

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Correspondence to Iqbal Hamza.

Supplementary information

Supplementary information

The file contains Supplementary Figures S1-S8 and Supplementary Table S1 with Legends, Supplementary Methods followed by Supplementary Notes. The anti-sense hHRG1 primer was corrected on 06 January 2009 (PDF 868 kb)

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Rajagopal, A., Rao, A., Amigo, J. et al. Haem homeostasis is regulated by the conserved and concerted functions of HRG-1 proteins. Nature 453, 1127–1131 (2008). https://doi.org/10.1038/nature06934

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