We recently reported that Inosine Monophosphate Dehydrogenase (IMPDH), a rate-limiting enzyme in de novo guanine nucleotide biosynthesis, clustered into macrostructures in response to decreased nucleotide levels and that there were differences between the IMPDH isoforms, IMPDH1 and IMPDH2. We hypothesised that the Bateman domains, which are present in both isoforms and serve as energy-sensing/allosteric modules in unrelated proteins, would contribute to isoform-specific differences and that mutations situated in and around this domain in IMPDH1 which give rise to retinitis pigmentosa (RP) would compromise regulation. We employed immuno-electron microscopy to investigate the ultrastructure of IMPDH macrostructures and live-cell imaging to follow clustering of an IMPDH2-GFP chimera in real-time. Using a series of IMPDH1/IMPDH2 chimera we demonstrated that the propensity to cluster was conferred by the N-terminal 244 amino acids, which includes the Bateman domain. A protease protection assay suggested isoform-specific purine nucleotide binding characteristics, with ATP protecting IMPDH1 and AMP protecting IMPDH2, via a mechanism involving conformational changes upon nucleotide binding to the Bateman domain without affecting IMPDH catalytic activity. ATP binding to IMPDH1 was confirmed in a nucleotide binding assay. The RP-causing mutation, R224P, abolished ATP binding and nucleotide protection and this correlated with an altered propensity to cluster. Collectively these data demonstrate that (i) the isoforms are differentially regulated by AMP and ATP by a mechanism involving the Bateman domain, (ii) communication occurs between the Bateman and catalytic domains and (iii) the RP-causing mutations compromise such regulation. These findings support the idea that the IMPDH isoforms are subject to distinct regulation and that regulatory defects contribute to human disease.
References
[1]
Collart FR, Huberman E (1988) Cloning and sequence analysis of the human and Chinese hamster inosine-5′-monophosphate dehydrogenase cDNAs. J Biol Chem 263: 15769–15772.
[2]
Natsumeda Y, Ohno S, Kawasaki H, Konno Y, Weber G, et al. (1990) Two distinct cDNAs for human IMP dehydrogenase. J Biol Chem 265: 5292–5295.
[3]
Carr SF, Papp E, Wu JC, Natsumeda Y (1993) Characterization of human type I and type II IMP dehydrogenases. J Biol Chem 268: 27286–27290.
[4]
Hager PW, Collart FR, Huberman E, Mitchell BS (1995) Recombinant human inosine monophosphate dehydrogenase type I and type II proteins. Purification and characterization of inhibitor binding. Biochem Pharmacol 49: 1323–1329.
[5]
Jackson RC, Weber G, Morris HP (1975) IMP dehydrogenase, an enzyme linked with proliferation and malignancy. Nature 256: 331–333.
[6]
Senda M, Natsumeda Y (1994) Tissue-differential expression of two distinct genes for human IMP dehydrogenase (E.C.1.1.1.205). Life Sci 54: 1917–1926.
[7]
Bowne SJ, Sullivan LS, Blanton SH, Cepko CL, Blackshaw S, et al. (2002) Mutations in the inosine monophosphate dehydrogenase 1 gene (IMPDH1) cause the RP10 form of autosomal dominant retinitis pigmentosa. Hum Mol Genet 11: 559–568.
[8]
Kennan A, Aherne A, Palfi A, Humphries M, McKee A, et al. (2002) Identification of an IMPDH1 mutation in autosomal dominant retinitis pigmentosa (RP10) revealed following comparative microarray analysis of transcripts derived from retinas of wild-type and Rho(-/-) mice. Hum Mol Genet 11: 547–557.
[9]
Gunter JH, Thomas EC, Lengefeld N, Kruger SJ, Worton L, et al. (2008) Characterisation of inosine monophosphate dehydrogenase expression during retinal development: Differences between variants and isoforms. Int J Biochem Cell Biol 40: 1716–1728.
[10]
Bowne SJ, Sullivan LS, Mortimer SE, Hedstrom L, Zhu J, et al. (2006) Spectrum and Frequency of Mutations in IMPDH1 Associated with Autosomal Dominant Retinitis Pigmentosa and Leber Congenital Amaurosis. Invest Ophthalmol Vis Sci 47: 34–42.
[11]
Aherne A, Kennan A, Kenna PF, McNally N, Lloyd DG, et al. (2004) On the molecular pathology of neurodegeneration in IMPDH1-based retinitis pigmentosa. Hum Mol Genet 13: 641–650.
[12]
Mortimer SE, Hedstrom L (2005) Autosomal dominant retinitis pigmentosa mutations in inosine monophosphate dehydrogenase type I disrupt nucleic acid binding. Biochem J 390: 41–47.
[13]
Bateman A (1997) The structure of a domain common to archaebacteria and the homocystinuria disease protein. Trends in Biochemical Sciences 22: 12–13.
[14]
Kemp BE (2004) Bateman domains and adenosine derivatives form a binding contract. J Clin Invest 113: 182–184.
[15]
Oakhill JS, Scott JW, Kemp BE (2012) AMPK functions as an adenylate charge-regulated protein kinase. Trends Endocrinol Metab 23: 125–132.
[16]
Ignoul S, Eggermont J (2005) CBS domains: structure, function, and pathology in human proteins. Am J Physiol Cell Physiol 289: C1369–1378.
[17]
Nimmesgern E, Black J, Futer O, Fulghum JR, Chambers SP, et al. (1999) Biochemical analysis of the modular enzyme inosine 5′-monophosphate dehydrogenase. Protein Expr Purif 17: 282–289.
[18]
Zhou X, Cahoon M, Rosa P, Hedstrom L (1997) Expression, purification, and characterization of inosine 5′-monophosphate dehydrogenase from Borrelia burgdorferi. J Biol Chem 272: 21977–21981.
[19]
Zhang R, Evans G, Rotella FJ, Westbrook EM, Beno D, et al. (1999) Characteristics and crystal structure of bacterial inosine-5′-monophosphate dehydrogenase. Biochemistry 38: 4691–4700.
[20]
Sintchak MD, Fleming MA, Futer O, Raybuck SA, Chambers SP, et al. (1996) Structure and mechanism of inosine monophosphate dehydrogenase in complex with the immunosuppressant mycophenolic acid. Cell 85: 921–930.
[21]
Pimkin M, Markham GD (2008) The CBS subdomain of inosine 5′-monophosphate dehydrogenase regulates purine nucleotide turnover. Mol Microbiol 68: 342–359.
[22]
Scott JW, Hawley SA, Green KA, Anis M, Stewart G, et al. (2004) CBS domains form energy-sensing modules whose binding of adenosine ligands is disrupted by disease mutations. J Clin Invest 113: 274–284.
[23]
Ji Y, Gu J, Makhov AM, Griffith JD, Mitchell BS (2006) Regulation of the interaction of inosine monophosphate dehydrogenase with mycophenolic acid by GTP. J Biol Chem 281: 206–212.
[24]
Colby TD, Vanderveen K, Strickler MD, Markham GD, Goldstein BM (1999) Crystal structure of human type II inosine monophosphate dehydrogenase: implications for ligand binding and drug design. Proc Natl Acad Sci U S A 96: 3531–3536.
[25]
Risal D, Strickler MD, Goldstein BM (2003) Crystal Structure of the Human Type I Inosine Monophosphate Dehydrogenase and Implications for Isoform Specificity To be published.
[26]
Nimmesgern E, Fox T, Fleming MA, Thomson JA (1996) Conformational changes and stabilization of inosine 5′-monophosphate dehydrogenase associated with ligand binding and inhibition by mycophenolic acid. J Biol Chem 271: 19421–19427.
[27]
Link JO, Straub K (1996) Trapping of an IMP Dehydrogenase - Substrate Covalent Intermediate by Mycophenolic Acid. Journal of the American Chemical Society 118: 2091–2092.
[28]
Okada M, Shimura K, Shiraki H, Nakagawa H (1983) IMP dehydrogenase. II. Purification and properties of the enzyme from Yoshida sarcoma ascites tumor cells. J Biochem (Tokyo) 94: 1605–1613.
[29]
Holmes EW, Pehlke DM, Kelley WN (1974) Human IMP dehydrogenase. Kinetics and regulatory properties. Biochim Biophys Acta 364: 209–217.
[30]
An S, Kumar R, Sheets ED, Benkovic SJ (2008) Reversible compartmentalization of de novo purine biosynthetic complexes in living cells. Science 320: 103–106.
[31]
An S, Deng Y, Tomsho JW, Kyoung M, Benkovic SJ (2010) Microtubule-assisted mechanism for functional metabolic macromolecular complex formation. Proc Natl Acad Sci U S A 107: 12872–12876.
[32]
Narayanaswamy R, Levy M, Tsechansky M, Stovall GM, O'Connell JD, et al. (2009) Widespread reorganization of metabolic enzymes into reversible assemblies upon nutrient starvation. Proc Natl Acad Sci U S A 106: 10147–10152.
[33]
Noree C, Sato BK, Broyer RM, Wilhelm JE (2010) Identification of novel filament-forming proteins in Saccharomyces cerevisiae and Drosophila melanogaster. J Cell Biol 190: 541–551.
[34]
Carcamo WC, Satoh M, Kasahara H, Terada N, Hamazaki T, et al. (2011) Induction of cytoplasmic rods and rings structures by inhibition of the CTP and GTP synthetic pathway in mammalian cells. PLoS One 6: e29690.
[35]
Whitehead JP, Simpson F, Hill MM, Thomas EC, Connolly LM, et al. (2004) Insulin and oleate promote translocation of inosine-5′ monophosphate dehydrogenase to lipid bodies. Traffic 5: 739–749.
[36]
Lucas M, Encinar JA, Arribas EA, Oyenarte I, Garcia IG, et al. (2010) Binding of S-methyl-5′-thioadenosine and S-adenosyl-L-methionine to protein MJ0100 triggers an open-to-closed conformational change in its CBS motif pair. J Mol Biol 396: 800–820.
[37]
Hnizda A, Spiwok V, Jurga V, Kozich V, Kodicek M, et al. (2010) Cross-talk between the catalytic core and the regulatory domain in cystathionine beta-synthase: study by differential covalent labeling and computational modeling. Biochemistry 49: 10526–10534.
[38]
Oakhill JS, Steel R, Chen ZP, Scott JW, Ling N, et al. (2011) AMPK is a direct adenylate charge-regulated protein kinase. Science 332: 1433–1435.
[39]
Xiao B, Heath R, Saiu P, Leiper FC, Leone P, et al. (2007) Structural basis for AMP binding to mammalian AMP-activated protein kinase. Nature 449: 496–500.
[40]
Day P, Sharff A, Parra L, Cleasby A, Williams M, et al. (2007) Structure of a CBS-domain pair from the regulatory gamma1 subunit of human AMPK in complex with AMP and ZMP. Acta Crystallogr D Biol Crystallogr 63: 587–596.
[41]
Meyer S, Savaresi S, Forster IC, Dutzler R (2007) Nucleotide recognition by the cytoplasmic domain of the human chloride transporter ClC-5. Nat Struct Mol Biol 14: 60–67.
[42]
Bennetts B, Rychkov GY, Ng HL, Morton CJ, Stapleton D, et al. (2005) Cytoplasmic ATP-sensing domains regulate gating of skeletal muscle ClC-1 chloride channels. J Biol Chem 280: 32452–32458.
[43]
Xiao B, Sanders MJ, Underwood E, Heath R, Mayer FV, et al. (2011) Structure of mammalian AMPK and its regulation by ADP. Nature 472: 230–233.
[44]
McLean JE, Hamaguchi N, Belenky P, Mortimer SE, Stanton M, et al. (2004) Inosine 5′-monophosphate dehydrogenase binds nucleic acids in vitro and in vivo. Biochem J 379: 243–251.
[45]
Mortimer SE, Xu D, McGrew D, Hamaguchi N, Lim HC, et al. (2008) IMP dehydrogenase type 1 associates with polyribosomes translating rhodopsin mRNA. J Biol Chem 283: 36354–36360.
[46]
Xu D, Cobb GC, Spellicy CJ, Bowne SJ, Daiger SP, et al. (2008) Retinal isoforms of inosine 5′-monophosphate dehydrogenase type 1 are poor nucleic acid binding proteins. Arch Biochem Biophys 472: 100–104.
[47]
Pimkin M, Pimkina J, Markham GD (2009) A regulatory role of the Bateman domain of IMP dehydrogenase in adenylate nucleotide biosynthesis. J Biol Chem 284: 7960–7969.
[48]
Su H, Gunter JH, de Vries M, Connor T, Wanyonyi S, et al. (2009) Inhibition of inosine monophosphate dehydrogenase reduces adipogenesis and diet-induced obesity. Biochem Biophys Res Commun 386: 351–355.
[49]
Kozhevnikova EN, van der Knaap JA, Pindyurin AV, Ozgur Z, van Ijcken WF, et al. (2012) Metabolic Enzyme IMPDH Is Also a Transcription Factor Regulated by Cellular State. Mol Cell 47: 133–139.
[50]
Olsen JV, Blagoev B, Gnad F, Macek B, Kumar C, et al. (2006) Global, In Vivo, and Site-Specific Phosphorylation Dynamics in Signaling Networks. Cell 127: 635–648.
[51]
Dephoure N, Zhou C, Villen J, Beausoleil SA, Bakalarski CE, et al. (2008) A quantitative atlas of mitotic phosphorylation. Proc Natl Acad Sci U S A 105: 10762–10767.
[52]
Glesne DA, Collart FR, Huberman E (1991) Regulation of IMP dehydrogenase gene expression by its end products, guanine nucleotides. Mol Cell Biol 11: 5417–5425.
[53]
Nixon SJ, Webb RI, Floetenmeyer M, Schieber N, Lo HP, et al. (2009) A single method for cryofixation and correlative light, electron microscopy and tomography of zebrafish embryos. Traffic 10: 131–136.
[54]
Martin S, Driessen K, Nixon SJ, Zerial M, Parton RG (2005) Regulated localization of Rab18 to lipid droplets: effects of lipolytic stimulation and inhibition of lipid droplet catabolism. J Biol Chem 280: 42325–42335.
[55]
Schieber NL, Nixon SJ, Webb RI, Oorschot VM, Parton RG (2010) Modern approaches for ultrastructural analysis of the zebrafish embryo. Methods Cell Biol 96: 425–442.
[56]
Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, et al. (2004) UCSF Chimera–a visualization system for exploratory research and analysis. J Comput Chem 25: 1605–1612.
[57]
Janosik M, Kery V, Gaustadnes M, Maclean KN, Kraus JP (2001) Regulation of human cystathionine beta-synthase by S-adenosyl-L-methionine: evidence for two catalytically active conformations involving an autoinhibitory domain in the C-terminal region. Biochemistry 40: 10625–10633.
