Arginine-Specific ADP-Ribosyltransferase and Its Acceptor Protein p33 in Chicken Polymorphonuclear Cells: Co-Localization in the Cell Granules, Partial Characterization, and In Situ Mono(ADP-Ribosyl)ation1

Abstract

We have reported the purification and characterization of arginine-specific ADP-ribosyltransferase from hen liver nuclei [Tanigawa, Y. et al. (1984) J. Biol. Chem. 259, 2022-2029] and the DNA-dependent mono(ADP-ribosyl)ation of p33, an acceptor protein in the nuclei [Mishima, K. et al. (1989) Eur. J. Biochem. 179, 267-273]. In the present study, we obtained evidence that among various tissues and cells from chicken, polymorphonuclear cells, so-called heterophils, possess both the ADP-ribosyltransferase and p33 at high levels. Percoll density gradient centrifugation of the postnuclear fraction of the heterophils revealed the co-localization of ADP-ribosyltransferase with p33 in the granule fraction. The enzyme and p33 were purified approximately 219- and 3.77-fold, respectively, from postnuclear pellet fraction to apparent homogeneity. The properties of heterophil ADP-ribosyltransferase and p33 were compared with those of the liver enzyme and p33. The molecular mass of the heterophil enzyme was estimated by SDS-polyacrylamide gel electrophoresis to be 27.5 kDa. The enzyme activity was stimulated by a sulfhydryl agent and inhibited by lysolecithin, NaCl, and inorganic phosphate. The mono(ADP-ribosyl)ation of p33 was markedly enhanced by polyanion, such as DNA, RNA, or poly(L-glutamate). SDS-polyacrylamide gel electrophoretic analysis after limited trypsin proteolysis of p33s, purified from chicken heterophils and liver, showed much the same pattern. Thus, it appears that ADP-ribosyltransferase and p33 present in heterophils are identical to those in the liver, respectively. p33 is considered to be an in situ substrate for ADP-ribosyltransferase, since it was specifically mono(ADP-ribosyl)ated in permeabilized heterophils.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of Glycosylphosphatidylinositiol-Anchored, Secreted, and Intracellular Vertebrate Mono-Adp-Ribosyltransferases

“…arginine methyl ester, agmatine) as ADP-ribose acceptors. The chicken transferase, unlike turkey transferase A, was inhibited by NaCl and lysophosphatidylcholine and stimulated by sulfhydryl reagents such as β-mercaptoethanol and by polyanions such as double-stranded DNA, RNA, or poly(L-glutamate) (50). Triton X-100 and CHAPS had no effect on enzyme activity.…”

“…A chicken ADP-ribosyltransferase (∼28 kDa) was isolated from hen liver and subsequently from chicken heterophil granules (50,94); a second 28-kDa isoform from heterophils was separated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (105). The ∼28-kDa transferase utilized histones, casein, protamine, and simple guanidino compounds (e.g.…”

“…Likewise, ADP-ribosylation of critical arginines in L-type pyruvate kinase (94) and in the α and β subunits of phosphorylase kinase (94) prevented cAMPdependent phosphorylation and subsequent modulation of kinase activities. The 33-kDa heterophil granule protein (p33), similar in amino acid sequence to the myb-induced myeloid protein-1 mim-1, was another substrate of the heterophil transferase (50,104). Effects of ADP-ribosylation on p33 or on granule function, however, have not been determined.…”

Myb induced myeloid protein 1 (Mim‐1) is an acetyltransferase

“…This highly purified Mim‐1 was incapable of acetylating histones in this assay (data not shown). The Mim‐1 protein was purified using the S column exploiting the net basic charge of the protein as previously described [22,23]. However, during the purification procedure we noticed that a small proportion (∼5%) of Mim‐1, as identified by sequencing, was retained by the Q column (see purification scheme in Fig.…”

Differentiation of HL‐60 cells is promoted by H‐toxin of Clostridium septicum

“…To clarify this phenomena, we investigated the effect of H-toxin on the endogenous mono-ADP-ribosylation of membrane proteins; because we thought that the first interaction of the toxin to HL-60 cells occurred in membranes and mono-ADP-ribosylation is an important posttranslational modification of proteins. Recently, the existence of ADP-ribosyltransferases in eukaryote cells was reported and some of them were purified [3][4][5][6][7]. We have studied the endogenous ADP-ribosylation in HL-60 cells [8].…”