COMMENTARY

Protein : A critical posttranslation modification in plants and animals

Kevin L. Moore1 Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, and Departments of Cell Biology and Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104

rotein tyrosine sulfation is a posttranslational modification restricted to proteins that transit the secretory pathway that was Pfirst described by Bettelheim in bovine fibrinopeptide B in 1954 (1). Subsequent pioneering work by Wieland Huttner’s group (2, 3) and others characterized the enzyme activity responsible for the reaction, called tyrosylprotein sulfotrans- ferase (TPST), that catalyzes the trans- fer of sulfate from 3Ј-phosphoadenosine 5Ј-phosphosulfate to the hydroxyl group of peptidyl tyrosine residues to form a tyrosine O4-sulfate ester. The enzymes’ subcellular localization in the trans- Golgi network and its widespread tissue Fig. 1. Domain structure of TPSTs. Human TPST-1 and TPST-2 are type II transmembrane proteins of and cellular distribution have been well similar size (370–377 residues). Each has a short N-terminal cytoplasmic domain, a single transmembrane documented in animals, and several domain (TM), and a putative Ϸ 40-residue stem region, followed by a luminal catalytic domain. Arabo- dozen tyrosine-sulfated proteins, mostly dopsis TPST is a 500-residue type I transmembrane proteins with a 24-residue N-terminal signal peptide (SP), a 446-residue luminal catalytic domain, followed by a single TM and a short cytoplasmic domain. of animal origin, have been described, Potential N- sites are depicted as lancets. HS6ST2, heparan sulfate 6-O-sulfotransferase 2. many of which play important roles in inflammation, hemostasis, immunity, and other processes (2–4). Finally, the internal peptide sequence was obtained 6-O-sulfotransferase 2. In light of the general importance of protein tyrosine by in-gel tryptic digestion and mass major difference in the primary struc- sulfation in protein–protein interactions spectrometry from which a full-length ture of TPSTs in Arabidopsis and animal has become widely accepted. cDNA was isolated that encodes a 500- species, it will be interesting to assess In 1998, TPST-1 was purified from rat residue polypeptide precursor (Fig. 1). whether the tertiary structures and cata- liver, and mouse and human TPST-1 The work by Komori et al. (9) identi- lytic mechanisms of the enzyme in cDNAs were identified (5). A second fies several significant differences be- plants and animals are similar. A third isoenzyme called TPST-2 was reported tween the TPST enzyme system in Ara- difference of note is that only a single later that year (6, 7). Importantly, TPST bidopsis (and likely other plant species) TPST gene is apparent in the Arabidop- orthologs from numerous vertebrate and and animals. The first notable differ- sis genome and other plant species. In invertebrate species are readily identifi- ence is that the Arabidopsis TPST is a contrast, animal genomes have two able based on sequence homology with type I transmembrane protein, in con- TPST genes, with the exception of Dro- the mouse or human enzymes. However, trast to all TPSTs of animal origin that sophila melanogaster. One can only spec- despite the presence of tyrosine-sulfated have type II transmembrane topology ulate on the reason for this. Perhaps peptides in plants first reported in 1996 (4). The precursor includes an N-termi- animal species have far more TPST sub- (8) and the sequencing of several plant nal 24-residue signal peptide and the strates than plants and thus evolved genomes, no plant TPST orthologs have mature polypeptide is predicted to have more than one enzyme to effectively been identified. These observations a 446-residue luminal catalytic domain sulfate all of them. However, it is still strongly indicated that TPSTs in plants with six potential N-glycosylation sites formally possible that Arabidopsis has a have very limited sequence homology followed by a single transmembrane he- second distantly related Tpst gene. It with those in the animal kingdom and lix and a short cytoplasmic domain. The would be of interest to directly address identification of plant TPSTs would re- second difference is that the Arabidopsis this question by determining whether quire some heavy lifting. TPST has no significant sequence ho- the loss-of-function mutant of the Arabi- The heavy lifting has been provided mology with TPSTs of animal origin, dopsis TPST lacks the ability to synthe- by Komori et al. and is documented in a which includes the lack of recognizable size protein-tyrosine sulfate. article published in this issue of PNAS 5Ј-PSB and 3ЈPB motifs. These two mo- Komori et al. (9) also show that the (9). They performed some elegant, old- tifs are highly conserved in all known Arabidopsis TPST enzyme system shares fashioned protein purification work and cytosolic and membrane sulfotrans- several general features with animals. accomplished a Ϸ 9,100-fold enrichment ferases and are involved in binding of As in the mouse and human systems, of TPST activity by affinity chromatog- the 5Ј and 3Ј phosphate groups of the raphy on a peptide substrate column reaction product 3Ј,5Ј-ADP, respec- followed by chromatography on hy- tively (10). The only significant homol- Author contributions: K.L.M. wrote the paper. droxyapatite. The putative TPST was ogy of note is that residues 371–447 of The author declares no conflict of interest. then identified by photoaffinity cross- Arabidopsis TPST align with a region See companion article on page 15067. linking as a 62-kDa polypeptide, and near the C terminus of heparan sulfate 1E-mail: [email protected].

www.pnas.org͞cgi͞doi͞10.1073͞pnas.0908376106 PNAS ͉ September 1, 2009 ͉ vol. 106 ͉ no. 35 ͉ 14741–14742 Downloaded by guest on September 25, 2021 the Arabidopsis enzyme is widely ex- cence, and other abnormalities. These phenotypes observed in mice with defi- pressed in plant tissues and is localized findings indicate that protein-tyrosine ciencies of Tpst1 and/or Tpst2. Thus, it is in the Golgi. Likewise, the sites of ty- sulfation is of crucial importance to nor- almost certain that many TPST sub- rosine sulfation in plant proteins are mal growth and development in plants. strates await description in both plants highly acidic in nature like most ty- The overall severity of the phenotype is and animals. Komori et al. deserve con- rosine-sulfation sites in animal proteins. similar to that observed in Tpst1;Tpst2 gratulations for their important contri- However, at present, PSK and PSY1 are double knockout mice that have severely bution to the field, but much work re- the only tyrosine-sulfated proteins that impaired postnatal survival (11). Most mains to be done. have been described in Arabidopsis.In importantly, the phenotype of the Arabi- addition, a loss-of-function mutant of dopsis TPST loss-of-function mutant ACKNOWLEDGMENTS. My work is supported by the Arabidopsis TPST displayed a mark- cannot be fully explained based on the Oklahoma Center for the Advancement of Sci- edly abnormal phenotype that includes current understanding of the TPST sub- ence and Technology Grant HR07–068 and Na- severely stunted growth, early senes- strate repertoire, as is the case for the tional Institutes of Health Grant HD056022.

1. Bettelheim FR (1954) Tyrosine O-sulfate in a peptide 6. Ouyang YB, Moore KL (1998) Molecular cloning and 9. Komori R, Amano Y, Ogawa-Ohnishi M, Matsubayashi from fibrinogen. J Am Chem Soc 76:2838–2839. expression of human and mouse tyrosylprotein sulfo- Y (2009) Identification of tyrosylprotein sulfotrans- 2. Huttner WB (1982) Sulphation of tyrosine residues: A wide- transferase-2 and a tyrosylprotein sulfotransferase ho- ferase in Arabidopsis. Proc Natl Acad Sci USA spread modification of proteins. Nature 299:273–276. mologue in Caenorhabditis elegans. J Biol Chem 106:15067–15072. 3. Huttner WB, Baeuerle PA (1988) Protein sulfation on 273:24770–24774. 10. Kakuta Y, Pedersen LG, Pedersen LC, Negishi M (1998) tyrosine. Mod Cell Biol 6:97–140. 7. Beisswanger R, et al. (1998) Existence of distinct tyrosyl- Conserved structural motifs in the sulfotransferase 4. Moore KL (2003) The biology and enzymology of protein protein sulfotransferase genes: Molecular character- family. Trends Biochem Sci 23:129–130. tyrosine O-sulfation. J Biol Chem 278:24243–24246. ization of tyrosylprotein sulfotransferase-2. Proc Natl 5. Ouyang YB, Lane WS, Moore KL (1998) Tyrosylprotein Acad Sci USA 95:11134–11139. 11. Westmuckett AD, Hoffhines AJ, Borghei A, Moore KL sulfotransferase: Purification and molecular cloning of 8. Matsubayashi Y, Sakagami Y (1996) Phytosulfokine, (2008) Early postnatal pulmonary failure and primary an enzyme that catalyzes tyrosine O-sulfation, a com- sulfated peptides that induce the proliferation of sin- hypothyroidism in mice with combined TPST-1 and mon posttranslational modification of eukaryotic pro- gle mesophyll cells of Asparagus officinalis L. Proc Natl TPST-2 deficiency. Gen Comp Endocrinol 156:145– teins. Proc Natl Acad Sci USA 95:2896–2901. Acad Sci USA 93:7623–7627. 153.

14742 ͉ www.pnas.org͞cgi͞doi͞10.1073͞pnas.0908376106 Moore Downloaded by guest on September 25, 2021