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• W2014993721 abstract "Nuclear respiratory factor 1 (NRF-1) is a transcriptional activator that acts on a diverse set of nuclear genes required for mitochondrial respiratory function in mammalian cells. These genes encode respiratory proteins as well as components of the mitochondrial transcription, replication, and heme biosynthetic machinery. Here, we establish that NRF-1 is a phosphoprotein in vivo. Phosphorylation occurs on serine residues within a concise NH2-terminal domain with the major sites of phosphate incorporation at serines 39, 44, 46, 47, and 52. The in vivo phosphorylation pattern can be approximated in vitro by phosphorylating recombinant NRF-1 with purified casein kinase II. Phosphate incorporation at the sites utilized in vivo results in a marked stimulation of DNA binding activity which is not observed in mutated proteins lacking these sites. Pairwise expression of the wild-type protein with each of a series of truncated derivatives in transfected cells results in the formation of a dimer between wild-type and mutant forms demonstrating that a homodimer is the active binding species. Although NRF-1 can dimerize in the absence of DNA, phosphorylation does not enhance the formation of these dimers. These findings suggest that phosphorylation results in an intrinsic change in the NRF-1 dimer enhancing its ability to bind DNA. Nuclear respiratory factor 1 (NRF-1) is a transcriptional activator that acts on a diverse set of nuclear genes required for mitochondrial respiratory function in mammalian cells. These genes encode respiratory proteins as well as components of the mitochondrial transcription, replication, and heme biosynthetic machinery. Here, we establish that NRF-1 is a phosphoprotein in vivo. Phosphorylation occurs on serine residues within a concise NH2-terminal domain with the major sites of phosphate incorporation at serines 39, 44, 46, 47, and 52. The in vivo phosphorylation pattern can be approximated in vitro by phosphorylating recombinant NRF-1 with purified casein kinase II. Phosphate incorporation at the sites utilized in vivo results in a marked stimulation of DNA binding activity which is not observed in mutated proteins lacking these sites. Pairwise expression of the wild-type protein with each of a series of truncated derivatives in transfected cells results in the formation of a dimer between wild-type and mutant forms demonstrating that a homodimer is the active binding species. Although NRF-1 can dimerize in the absence of DNA, phosphorylation does not enhance the formation of these dimers. These findings suggest that phosphorylation results in an intrinsic change in the NRF-1 dimer enhancing its ability to bind DNA. Mitochondrial respiratory function requires the expression of essential gene products from both nuclear and mitochondrial genetic systems. Because of its compact structure and limited coding capacity, the mitochondrial genome encodes only 13 proteins along with the tRNAs and rRNAs required for their translation (for review, see Ref. 2Attardi G. Schatz G. Annu. Rev. Cell Biol. 1988; 4: 289-333Crossref PubMed Scopus (1054) Google Scholar). All of these proteins are subunits of the inner membrane respiratory complexes. Thus, nuclear genes must specify the majority of respiratory subunits and all of the proteins required for the expression, maintenance, and replication of mitochondrial DNA (for review, see Ref.8Clayton D.A. Annu. Rev. Cell Biol. 1991; 7: 453-478Crossref PubMed Scopus (524) Google Scholar). One approach to understanding nucleo-mitochondrial interactions in mammalian cells is to identify the nuclear transcription factors that govern the expression of these genes.NRF-1 1The abbreviations used are: NRF-1, nuclear respiratory factor 1; HA, hemagglutinin; CKII, casein kinase II; SRF, serum response factor; MEF2C, myocyte enhancer factor 2C; PAGE, polyacrylamide gel electrophoresis. 1The abbreviations used are: NRF-1, nuclear respiratory factor 1; HA, hemagglutinin; CKII, casein kinase II; SRF, serum response factor; MEF2C, myocyte enhancer factor 2C; PAGE, polyacrylamide gel electrophoresis. was originally identified as a nuclear transcription factor that acts on mammalian genes encoding cytochromec and a number of other respiratory proteins (4Evans M.J. Scarpulla R.C. J. Biol. Chem. 1989; 264: 14361-14368Abstract Full Text PDF PubMed Google Scholar, 6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 10Chau C.A. Evans M.J. Scarpulla R.C. J. Biol. Chem. 1992; 267: 6999-7006Abstract Full Text PDF PubMed Google Scholar). A possible role for the factor in intergenomic communication is supported by the discovery of functional NRF-1 recognition sites in nuclear genes specifying the rate-limiting heme biosynthetic enzyme, 5-aminolevulinate synthase (14Braidotti G. Borthwick I.A. May B.K. J. Biol. Chem. 1993; 268: 1109-1117Abstract Full Text PDF PubMed Google Scholar), and components of the mitochondrial transcription and replication machinery (6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 18Virbasius J.V. Scarpulla R.C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 1309-1313Crossref PubMed Scopus (596) Google Scholar, 22Scarpulla R.C. Trends Cardiovasc. Med. 1996; 6: 39-45Crossref PubMed Scopus (61) Google Scholar). The latter include the RNA subunit of mitochondrial RNA processing endonuclease, an enzyme implicated in the formation of mtDNA replication primers and mtTFA, an activator of mitochondrial transcription (for review, see Refs. 8Clayton D.A. Annu. Rev. Cell Biol. 1991; 7: 453-478Crossref PubMed Scopus (524) Google Scholar and16Shadel G.S. Clayton D.A. J. Biol. Chem. 1993; 268: 16083-16086Abstract Full Text PDF PubMed Google Scholar). These findings led to a model whereby NRF-1, along with other transcription factors, helps coordinate the synthesis and function of respiratory proteins from both genomes (6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 18Virbasius J.V. Scarpulla R.C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 1309-1313Crossref PubMed Scopus (596) Google Scholar, 22Scarpulla R.C. Trends Cardiovasc. Med. 1996; 6: 39-45Crossref PubMed Scopus (61) Google Scholar). In addition, it has been postulated that NRF-1 may play a role in other developmental and growth regulatory processes (10Chau C.A. Evans M.J. Scarpulla R.C. J. Biol. Chem. 1992; 267: 6999-7006Abstract Full Text PDF PubMed Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar). Most notably, the chicken homolog of NRF-1 has recently been associated with the expression of the histone H5 gene during erythrocyte development (21Gomez-Cuadrado A. Martin M. Noel M. Ruiz-Carrillo A. Mol. Cell. Biol. 1995; 15: 6670-6685Crossref PubMed Google Scholar).NRF-1 has been purified and a cDNA clone isolated and characterized (10Chau C.A. Evans M.J. Scarpulla R.C. J. Biol. Chem. 1992; 267: 6999-7006Abstract Full Text PDF PubMed Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar). The protein is related through a novel DNA binding domain to developmental regulatory proteins from sea urchins (12Calzone F.J. Hoog C. Teplow D.B. Cutting A.E. Zeller R.W. Britten R.J. Davidson E.H. Development. 1991; 112: 335-350Crossref PubMed Google Scholar) andDrosophila (13Desimone S.M. White K. Mol. Cell. Biol. 1993; 13: 3641-3949Crossref PubMed Scopus (83) Google Scholar). This highly conserved DNA binding domain is in the NH2-terminal half of the molecule, and the conservation extends beyond its NH2-terminal boundary through a region containing a complex nuclear localization signal (17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar,23Gugneja S. Virbasius C.A. Scarpulla R.C. Mol. Cell. Biol. 1996; 16: 5708-5716Crossref PubMed Scopus (64) Google Scholar). A large COOH-terminal domain in NRF-1 is required for transcriptional activation and is not conserved in the lower eukaryotic proteins (23Gugneja S. Virbasius C.A. Scarpulla R.C. Mol. Cell. Biol. 1996; 16: 5708-5716Crossref PubMed Scopus (64) Google Scholar). Here, we establish that a concise NH2-terminal domain in NRF-1 serves as a target for phosphorylation in vivo and in vitro. The phosphorylation occurs on multiple serine residues and enhances DNA binding activity. Although the phosphodomain overlaps a region required for NRF-1 homodimerization, phosphorylation does not alter the monomer-dimer equilibrium in the absence of DNA. Rather, phosphorylation appears to cause an intrinsic change in the NRF-1 dimer thereby enhancing its DNA binding.DISCUSSIONThe preponderance of evidence points to a role for NRF-1 in nucleo-mitochondrial interactions in mammalian cells (6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar, 18Virbasius J.V. Scarpulla R.C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 1309-1313Crossref PubMed Scopus (596) Google Scholar). Functional NRF-1 sites are present in the majority of nuclear genes encoding respiratory subunits and also in several genes required for mitochondrial transcription, replication, and heme biosynthesis (22Scarpulla R.C. Trends Cardiovasc. Med. 1996; 6: 39-45Crossref PubMed Scopus (61) Google Scholar). In addition, NRF-1 may serve a more integrative role in coordinate gene expression by acting on target genes encoding key components in the pathways of cellular metabolism, signal transduction, and chromosome maintenance (10Chau C.A. Evans M.J. Scarpulla R.C. J. Biol. Chem. 1992; 267: 6999-7006Abstract Full Text PDF PubMed Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar). Several structural domains in NRF-1 which are required for its biological function have been resolved. A DNA binding domain within the NH2-terminal two-thirds of the molecule is highly conserved in a small family of eukaryotic regulatory proteins which includes the sea urchin P3A2 factor and the Drosophilaerect wing gene product (17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar). These proteins are required for proper neuromuscular development in their respective organisms. The COOH-terminal domain is not conserved among family members. In NRF-1, this domain contains clusters of hydrophobic residues that are required for transcriptional activation (23Gugneja S. Virbasius C.A. Scarpulla R.C. Mol. Cell. Biol. 1996; 16: 5708-5716Crossref PubMed Scopus (64) Google Scholar). In addition to the DNA binding domain, significant sequence conservation with P3A2 is present between residues 37 and 116. Part of the sequence conservation in this region (residues 88–116) has been ascribed to a complex nuclear localization signal located just upstream from the DNA binding domain (21Gomez-Cuadrado A. Martin M. Noel M. Ruiz-Carrillo A. Mol. Cell. Biol. 1995; 15: 6670-6685Crossref PubMed Google Scholar, 23Gugneja S. Virbasius C.A. Scarpulla R.C. Mol. Cell. Biol. 1996; 16: 5708-5716Crossref PubMed Scopus (64) Google Scholar). No other functions had been assigned to the remainder of this conserved region.Here, we establish that NRF-1 is a phosphoprotein and that in vivo phosphorylation occurs within a concise NH2-terminal domain on serine residues 39, 44, 46, 47, and 52. Four of the five serines (39, 44, 46, and 47) are conserved in P3A2, but it is unknown whether they are phosphorylated in sea urchins. All five serines conform to the consensus for phosphorylation by CKII (35Songyang Z. Lu K.P. Kwon Y.T. Tsai L.-H. Filhol O. Cochet C. Brickey D.A. Soderling T.R. Bartleson C. Graves D.J. DeMaggio A.J. Hoekstra M.F. Blenis J. Hunter T. Cantley L.C. Mol. Cell. Biol. 1996; 16: 6486-6493Crossref PubMed Scopus (486) Google Scholar) (S/T-X-X-acidic, where the acidic residue can be glutamate, aspartate, phosphoserine, or phosphotyrosine), and the purified enzyme phosphorylates these sites in vitro. All five are surrounded by acidic amino acids that are prevalent in known CKII sites, and all but serine 44 have the requisite acidic residue at the +3 position. In the serine 44 site it is likely that the negative charge at this position is contributed by phosphoserine 47.Although CKII is the most likely candidate for the in vivoNRF-1 kinase, the in vivo and in vitrophosphorylation patterns are not identical. The major difference is that the region between residues 48 and 77 is more intensely phosphorylated in vitro than it is in vivo. The stoichiometry of phosphorylation suggests that this region accepts two phosphates on a molar basis. In addition to serine 52, which is weakly phosphorylated in vivo, threonines 51 and 62 conform to the consensus. These residues can account for the in vitrothreonine phosphorylation detected in wild-type and mutated proteins. Thus, it is likely that these residues are more accessible to phosphorylation in the in vitro CKII reaction. It is important to note, however, that threonine phosphorylation by itself does not contribute to enhanced DNA binding. Only that majority of sites which are phosphorylated both in vivo and in vitro can account for the stimulatory effect of phosphorylation on NRF-1 DNA binding activity.The formation of NRF-1 heterodimers in transfected cells expressing both wild-type and deleted proteins constitutes compelling evidence that the protein binds DNA as a dimer. This is consistent with the dyad symmetry in the pallidromic NRF-1 recognition site (6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar). Although glutaraldehyde cross-linking results in two species migrating as monomer and dimer on denaturing gels (not shown), attempts to form heterodimers in vitro by mixing native proteins were unsuccessful. This has also been observed with the chicken homolog of NRF-1 and has been attributed to a low equilibrium dissociation constant for the homodimer (21Gomez-Cuadrado A. Martin M. Noel M. Ruiz-Carrillo A. Mol. Cell. Biol. 1995; 15: 6670-6685Crossref PubMed Google Scholar). Synthesis of wild-type and deleted proteins in the same cellular compartment in transfected cells allows free association among subunits and results in the detection of heterodimers.Here, we demonstrate that the stability of the NRF-1 dimer in the absence of DNA is disrupted upon progressive NH2-terminal deletion to residue 78. Upon deletion to this residue, the predominant species detected on native gels is clearly monomer. This indicates that a strong determinant of dimerization is located in the NH2-terminal region of the protein apart from the DNA binding domain. However, a protein with the 1–78 deletion binds DNA as a homodimer, can heterodimerize with wild-type on DNA (Fig. 6), and can be cross-linked to a dimer with glutaraldehyde (not shown). We note that the Mut 3 protein does appear to bind DNA less strongly than the wild-type, which is consistent with removal of a dimerization domain. This is in agreement with the results obtained with chicken NRF-1 which suggest an overlap between DNA binding and dimerization functions between residues 79 and 172 (21Gomez-Cuadrado A. Martin M. Noel M. Ruiz-Carrillo A. Mol. Cell. Biol. 1995; 15: 6670-6685Crossref PubMed Google Scholar). We conclude that the NH2-terminal domain identified here is essential for stable subunit interactions in the absence of DNA. In addition, determinants within the DNA binding domain contribute a level of stability sufficient for DNA binding and for detection with cross-linking agents. However, we find no evidence that phosphorylation stimulates DNA binding by promoting dimer stability. Complete elimination of all NH2-terminal phosphorylation sites by site-directed mutagenesis does not result in any detectable dissociation to monomer on native gels. This suggests that phosphorylation affects DNA binding at the level of DNA-protein interaction.One might expect that the phosphorylation-dependent enhancement of DNA binding would result in an increased ability of wild-type NRF-1 to activate transcription compared with Mut 3. We have previously compared wild-type NRF-1 and Mut 3 for their ability totrans-activate a luciferase reporter whose expression was driven by four copies of the NRF-1 binding site from the rat cytochromec gene cloned upstream of a minimal promoter in transient cotransfection experiments in COS-1 cells (23Gugneja S. Virbasius C.A. Scarpulla R.C. Mol. Cell. Biol. 1996; 16: 5708-5716Crossref PubMed Scopus (64) Google Scholar). In those experiments, both proteins activated transcription by about 7–8-fold over that of an empty vector. The absence of a difference between these constructs under conditions of transient transfection most likely results from the fact that the transgene expresses NRF-1 or Mut 3 at very high levels. Since Mut 3 can still bind DNA and has an intact activation domain, overexpression may compensate for its reduced DNA binding activity relative to that of the wild-type phosphorylated protein.Several transcription factors are now known to be phosphorylated at CKII consensus sites both in vivo and in vitro. In particular, NRF-1 shares a number of interesting similarities with two such factors, SRF and MEF2C. Both SRF (30Manak J.R. Prywes R. Oncogene. 1993; 8: 703-711PubMed Google Scholar, 31Janknecht R. Hipskind R.A. Houthaeve T. Nordheim A. Stunnenberg H.G. EMBO J. 1992; 11: 1045-1054Crossref PubMed Scopus (108) Google Scholar, 32Marais R.M. Hsuan J.J. McGuigan C. Wynne J. Treisman R. EMBO J. 1992; 11: 97-105Crossref PubMed Scopus (128) Google Scholar, 36Manak J.R. de Bisschop N. Kris R.M. Prywes R. Genes Dev. 1990; 4: 955-967Crossref PubMed Scopus (118) Google Scholar) and MEF2C (27Molkentin J.D. Li L. Olson E.N. J. Biol. Chem. 1996; 271: 17199-17204Abstract Full Text Full Text PDF PubMed Scopus (90) Google Scholar) resemble NRF-1 in that phosphorylation of a restricted number of serine residues adjacent to the region of the molecule which contacts DNA accounts for enhanced factor binding to their cognate recognition sites. In SRF, dephosphorylation of specific serine residues resulted in decreases in both its association and dissociation rates leading to the conclusion that phosphorylation does not alter the equilibrium binding affinity for DNA (31Janknecht R. Hipskind R.A. Houthaeve T. Nordheim A. Stunnenberg H.G. EMBO J. 1992; 11: 1045-1054Crossref PubMed Scopus (108) Google Scholar, 32Marais R.M. Hsuan J.J. McGuigan C. Wynne J. Treisman R. EMBO J. 1992; 11: 97-105Crossref PubMed Scopus (128) Google Scholar). We also note that phosphorylation results in a marked increase in the rate of dissociation of NRF-1 from DNA, 2S. Gugneja and R. C. Scarpulla, unpublished observations. suggesting that its exchange rate on DNA may be similarly affected. Conversion of the essential serines to acidic amino acids in both SRF and MEF2C had the same stimulatory effect on DNA binding as phosphorylation. In addition, phosphorylation does not alter the ability of SRF to dimerize (33Manak J.R. Prywes R. Mol. Cell. Biol. 1991; 11: 3652-3659Crossref PubMed Scopus (72) Google Scholar) or of MEF2C to interact with other myogenic transcription factors (27Molkentin J.D. Li L. Olson E.N. J. Biol. Chem. 1996; 271: 17199-17204Abstract Full Text Full Text PDF PubMed Scopus (90) Google Scholar). In both cases it has been postulated that introduction of a negative charge may induce a conformational change that affects the DNA binding domain (27Molkentin J.D. Li L. Olson E.N. J. Biol. Chem. 1996; 271: 17199-17204Abstract Full Text Full Text PDF PubMed Scopus (90) Google Scholar,33Manak J.R. Prywes R. Mol. Cell. Biol. 1991; 11: 3652-3659Crossref PubMed Scopus (72) Google Scholar). Such a mechanism is consistent with the finding that NRF-1 phosphorylation also stimulates DNA binding without altering the association between homologous subunits. These similarities suggest that SRF, MEF2C, and NRF-1 may be targets for the same or very similar kinase-mediated signaling pathway.Control of NRF-1 function by phosphorylation is an attractive mechanism given the proposed role for the factor in nucleomitochondrial interactions and the integration of other growth regulatory pathways. Although CKII activity is stimulated by serum growth factors (for review, see Ref. 34Litchfield D.W. Luscher B. Mol. Cell. Biochem. 1993; 127/128: 187-199Crossref Scopus (160) Google Scholar) there is conflicting evidence that this stimulation leads to the activation of gene expression through the modification of specific transcription factors. No change in the phosphorylated state of SRF or the relative amounts of SRF isoforms has been found in response to growth factor treatment. Stable expression of a mutant SRF that binds its recognition site at constitutively high levels had no effect on c-fos expression (30Manak J.R. Prywes R. Oncogene. 1993; 8: 703-711PubMed Google Scholar). By contrast, phosphorylation of the CKII sites in the upstream binding factor markedly elevates the trans-activation of their ribosomal RNA target genes. In addition, the phosphorylated state of upstream binding factor but not its level of expression, is stimulated in proliferating relative to serum arrested cells, and this correlates with enhanced ribosomal RNA transcription (28Voit R. Schnapp A. Kuhn A. Rosenbauer H. Hirschmann P. Stunnenberg H.G. Grummt I. EMBO J. 1992; 11: 2211-2218Crossref PubMed Scopus (157) Google Scholar, 29O'Mahony D.J. Xie W. Smith S.D. Singer H.A. Rothblum L.I. J. Biol. Chem. 1992; 267: 35-38Abstract Full Text PDF PubMed Google Scholar). One unexplored possibility is that phosphorylation of transcription factors such as NRF-1 by CKII or related kinases occurs in response to intracellular signals that communicate the respiratory state of the mitochondria to the nuclear transcriptional apparatus. Such a mechanism may link the expression of nuclear genes to the availability of respiratory energy. The results presented here represent an essential first step in understanding the potential role of phosphorylation in NRF-1-dependent gene expression. Mitochondrial respiratory function requires the expression of essential gene products from both nuclear and mitochondrial genetic systems. Because of its compact structure and limited coding capacity, the mitochondrial genome encodes only 13 proteins along with the tRNAs and rRNAs required for their translation (for review, see Ref. 2Attardi G. Schatz G. Annu. Rev. Cell Biol. 1988; 4: 289-333Crossref PubMed Scopus (1054) Google Scholar). All of these proteins are subunits of the inner membrane respiratory complexes. Thus, nuclear genes must specify the majority of respiratory subunits and all of the proteins required for the expression, maintenance, and replication of mitochondrial DNA (for review, see Ref.8Clayton D.A. Annu. Rev. Cell Biol. 1991; 7: 453-478Crossref PubMed Scopus (524) Google Scholar). One approach to understanding nucleo-mitochondrial interactions in mammalian cells is to identify the nuclear transcription factors that govern the expression of these genes. NRF-1 1The abbreviations used are: NRF-1, nuclear respiratory factor 1; HA, hemagglutinin; CKII, casein kinase II; SRF, serum response factor; MEF2C, myocyte enhancer factor 2C; PAGE, polyacrylamide gel electrophoresis. 1The abbreviations used are: NRF-1, nuclear respiratory factor 1; HA, hemagglutinin; CKII, casein kinase II; SRF, serum response factor; MEF2C, myocyte enhancer factor 2C; PAGE, polyacrylamide gel electrophoresis. was originally identified as a nuclear transcription factor that acts on mammalian genes encoding cytochromec and a number of other respiratory proteins (4Evans M.J. Scarpulla R.C. J. Biol. Chem. 1989; 264: 14361-14368Abstract Full Text PDF PubMed Google Scholar, 6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 10Chau C.A. Evans M.J. Scarpulla R.C. J. Biol. Chem. 1992; 267: 6999-7006Abstract Full Text PDF PubMed Google Scholar). A possible role for the factor in intergenomic communication is supported by the discovery of functional NRF-1 recognition sites in nuclear genes specifying the rate-limiting heme biosynthetic enzyme, 5-aminolevulinate synthase (14Braidotti G. Borthwick I.A. May B.K. J. Biol. Chem. 1993; 268: 1109-1117Abstract Full Text PDF PubMed Google Scholar), and components of the mitochondrial transcription and replication machinery (6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 18Virbasius J.V. Scarpulla R.C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 1309-1313Crossref PubMed Scopus (596) Google Scholar, 22Scarpulla R.C. Trends Cardiovasc. Med. 1996; 6: 39-45Crossref PubMed Scopus (61) Google Scholar). The latter include the RNA subunit of mitochondrial RNA processing endonuclease, an enzyme implicated in the formation of mtDNA replication primers and mtTFA, an activator of mitochondrial transcription (for review, see Refs. 8Clayton D.A. Annu. Rev. Cell Biol. 1991; 7: 453-478Crossref PubMed Scopus (524) Google Scholar and16Shadel G.S. Clayton D.A. J. Biol. Chem. 1993; 268: 16083-16086Abstract Full Text PDF PubMed Google Scholar). These findings led to a model whereby NRF-1, along with other transcription factors, helps coordinate the synthesis and function of respiratory proteins from both genomes (6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 18Virbasius J.V. Scarpulla R.C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 1309-1313Crossref PubMed Scopus (596) Google Scholar, 22Scarpulla R.C. Trends Cardiovasc. Med. 1996; 6: 39-45Crossref PubMed Scopus (61) Google Scholar). In addition, it has been postulated that NRF-1 may play a role in other developmental and growth regulatory processes (10Chau C.A. Evans M.J. Scarpulla R.C. J. Biol. Chem. 1992; 267: 6999-7006Abstract Full Text PDF PubMed Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar). Most notably, the chicken homolog of NRF-1 has recently been associated with the expression of the histone H5 gene during erythrocyte development (21Gomez-Cuadrado A. Martin M. Noel M. Ruiz-Carrillo A. Mol. Cell. Biol. 1995; 15: 6670-6685Crossref PubMed Google Scholar). NRF-1 has been purified and a cDNA clone isolated and characterized (10Chau C.A. Evans M.J. Scarpulla R.C. J. Biol. Chem. 1992; 267: 6999-7006Abstract Full Text PDF PubMed Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar). The protein is related through a novel DNA binding domain to developmental regulatory proteins from sea urchins (12Calzone F.J. Hoog C. Teplow D.B. Cutting A.E. Zeller R.W. Britten R.J. Davidson E.H. Development. 1991; 112: 335-350Crossref PubMed Google Scholar) andDrosophila (13Desimone S.M. White K. Mol. Cell. Biol. 1993; 13: 3641-3949Crossref PubMed Scopus (83) Google Scholar). This highly conserved DNA binding domain is in the NH2-terminal half of the molecule, and the conservation extends beyond its NH2-terminal boundary through a region containing a complex nuclear localization signal (17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar,23Gugneja S. Virbasius C.A. Scarpulla R.C. Mol. Cell. Biol. 1996; 16: 5708-5716Crossref PubMed Scopus (64) Google Scholar). A large COOH-terminal domain in NRF-1 is required for transcriptional activation and is not conserved in the lower eukaryotic proteins (23Gugneja S. Virbasius C.A. Scarpulla R.C. Mol. Cell. Biol. 1996; 16: 5708-5716Crossref PubMed Scopus (64) Google Scholar). Here, we establish that a concise NH2-terminal domain in NRF-1 serves as a target for phosphorylation in vivo and in vitro. The phosphorylation occurs on multiple serine residues and enhances DNA binding activity. Although the phosphodomain overlaps a region required for NRF-1 homodimerization, phosphorylation does not alter the monomer-dimer equilibrium in the absence of DNA. Rather, phosphorylation appears to cause an intrinsic change in the NRF-1 dimer thereby enhancing its DNA binding. DISCUSSIONThe preponderance of evidence points to a role for NRF-1 in nucleo-mitochondrial interactions in mammalian cells (6Evans M.J. Scarpulla R.C. Genes Dev. 1990; 4: 1023-1034Crossref PubMed Scopus (325) Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar, 18Virbasius J.V. Scarpulla R.C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 1309-1313Crossref PubMed Scopus (596) Google Scholar). Functional NRF-1 sites are present in the majority of nuclear genes encoding respiratory subunits and also in several genes required for mitochondrial transcription, replication, and heme biosynthesis (22Scarpulla R.C. Trends Cardiovasc. Med. 1996; 6: 39-45Crossref PubMed Scopus (61) Google Scholar). In addition, NRF-1 may serve a more integrative role in coordinate gene expression by acting on target genes encoding key components in the pathways of cellular metabolism, signal transduction, and chromosome maintenance (10Chau C.A. Evans M.J. Scarpulla R.C. J. Biol. Chem. 1992; 267: 6999-7006Abstract Full Text PDF PubMed Google Scholar, 17Virbasius C.A. Virbasius J.V. Scarpulla R.C. Genes Dev. 1993; 7: 2431-2445Crossref PubMed Scopus (275) Google Scholar)." @default.
• W2014993721 created "2016-06-24" @default.
• W2014993721 creator A5021947650 @default.
• W2014993721 creator A5085943861 @default.
• W2014993721 date "1997-07-01" @default.
• W2014993721 modified "2023-10-14" @default.
• W2014993721 title "Serine Phosphorylation within a Concise Amino-terminal Domain in Nuclear Respiratory Factor 1 Enhances DNA Binding" @default.
• W2014993721 cites W1483361087 @default.
• W2014993721 cites W1488283633 @default.
• W2014993721 cites W1490733930 @default.
• W2014993721 cites W1494925286 @default.
• W2014993721 cites W1540990864 @default.
• W2014993721 cites W1557098204 @default.
• W2014993721 cites W1594358926 @default.
• W2014993721 cites W1616609814 @default.
• W2014993721 cites W1873199501 @default.
• W2014993721 cites W1950159988 @default.
• W2014993721 cites W1965372133 @default.
• W2014993721 cites W1987779112 @default.
• W2014993721 cites W1996877995 @default.
• W2014993721 cites W2001109729 @default.
• W2014993721 cites W2002662437 @default.
• W2014993721 cites W2011147958 @default.
• W2014993721 cites W2036931150 @default.
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