Expression of Sporophytic Storage Proteins in the Corm of the Quillwort (Isoetes echinospora Dur.).

Parenchyma cells from the corm tissue of the aquatic lycopod Isoetes echinospora Dur. were shown by electron microscopy to be packed with amyloplasts, lipid bodies, and protein bodies. The protein bodies are morphologically similar to those identified in seeds and certain vegetative tissues of higher plants. Globoid-containing protein bodies (1-10 [mu]m) isolated in a sucrose gradient possessed a buoyant density of 1.28 g/mL and contained globulin (salt-soluble) proteins. Sucrose gradient centrifugation of crude globulins revealed only two components with mean sedimentation coefficients of approximately 2S and 11S. The 2S component, designated VSP-IsA, was composed of a 15.7-kD polypeptide. The 11S component, designated VSP-IsB, had a molecular mass of 215 kD as estimated by gel filtration and was composed of 39- to 42-kD polypeptides. Two-dimensional gel electrophoresis showed constituent polypeptides distinguished by differences in net charge and molecular mass. Affinity-purified antibodies against VSP-IsA and VSP-IsB prepared and used as probes on immunoblots cross-react only with their specific antigens, suggesting that the proteins are not immunologically related. Indirect immunolocalization studies confirmed that VSP-IsB is deposited in protein bodies. These globulin proteins, like those from some seeds, form the principal storage reserves of the corm tissue.

Induction of an anti-Fab, anti-DNA and anti-RNA polymerase I autoantibody response network in rabbits immunized with SLE anti-DNA antibody.

Systemic lupus erythematosus (SLE)-like anti-IgG Fab autoantibodies (autoAb) were induced in rabbits by immunization with either human, mouse or rabbit anti-DNA Ab. In direct-binding radioimmunoassay (RIA), affinity-purified anti-normal rabbit (NR) Fab autoAb cross-reacted with normal mouse (NM) Fab, ssDNA (but not dsDNA), poly(dA,dT), and RNA polymerase I (RPI). Affinity-purified anti-NM IgG Ab isolated from the same antisera cross-reacted with NR Fab, ssDNA and RPI. In inhibition RIA, soluble NR Fab inhibited anti-NR Fab binding to NR Fab and ssDNA, but enhanced binding to RPI. In contrast, ssDNA or RPI inhibitors had no effect upon autoAb binding to NR Fab. Anti-DNA, anti-RPI and anti-RPI 190 kD subunit autoAb, induced by immunization with lupus mouse anti-DNA Ab, also reacted with NM Fab, but were idiotypically specific for lupus mouse anti-DNA Fab. Further, rabbit anti-DNA and anti-RPI IgG autoAb, induced by immunization with rabbit anti-DNA IgG, were each idiotypically specific for homologous and autologous rabbit anti-ssDNA Fab. Together, these data provide evidence that anti-DNA, anti-RPI and anti-Fab autoAb are linked in a complex, multiple-specific and perhaps regulatory, immune response idiotype network in SLE.

Anti-RNA polymerase I antibodies in the sera of MRL lupus mice at the initial stages of disease are directed primarily against phosphorylation-dependent epitopes.

Anti-RNA polymerase I (RPI) antibodies in the sera of MRL/Mp-lpr/lpr and MRL/Mp(-)+/+ mice, which develop an autoimmune disease similar to human systemic lupus erythematosus, were screened for reactivity with purified RPI or RPI which had been dephosphorylated. In every case (n = 10), dephosphorylation of RPI resulted in a significant decrease (33-95%) in antibody binding. The anti-RPI antibodies in the sera of the same mice approximately 6 weeks later also reacted better with untreated as compared to dephosphorylated RPI but, in every case, the decrease in antibody (0-30%) caused by dephosphorylation was substantially diminished. That the proportion of anti-RPI antibodies in the sera of MRL mice decreased with progression of lupus-like disease was confirmed by closely monitoring the antibodies over the course of disease. Anti-RPI antibodies produced at the earliest stages appeared to be directed almost exclusively against phosphorylation-dependent determinants since dephosphorylation of RPI essentially abolished antibody binding. Subsequently, the percentage of the total anti-RPI antibodies in the sera of these mice directed towards phosphorylation-independent epitopes increased linearly with time. The importance of phosphorylation-dependent epitopes on RPI for the development of the anti-RPI autoimmune response was supported by the observation that treatment of mice with alkaline phosphatase partially attenuated anti-RPI antibody production.

Rabbits produce SLE-like anti-RNA polymerase I and anti-DNA autoantibodies in responses to immunization with either human or murine SLE anti-DNA antibodies.

Anti-DNA and anti-DNA polymerase I (RPI) autoantibody responses are symptoms of systemic lupus erythematosus (SLE). To investigate the relationship between these antibodies (Ab), rabbits were immunized with one of the following preparations: human SLE anti-DNA Ab; human SLE anti-DNA IgG; normal human anti-DNA Ab; human Grave's disease anti-DNA Ab; murine SLE anti-DNA Ab or anti-DNA IgG Fab; various normal human, murine, or rabbit IgG preparations; or complete Freund's adjuvant (CFA), alone. All of the animals immunized with anti-DNA Ab (n = 14) generated Ab reactive in radioimmunoassay with: ssDNA, dsDNA, RPI, the soluble fraction of rabbit liver crude nuclear extract, and the immunogen. Induced rabbit anti-DNA Ab in turn induced these responses in a different rabbit: a rabbit immunized with rabbit anti-DNA IgG Ab which had been previously induced by immunization with human anti-DNA Ab, produced Ab reactive with ssDNA, dsDNA, RPI, and the soluble fraction of rabbit liver nuclear extract. Although an individual animal's antisera reacted consistently over the course of immunization with the same individual RPI subunit(s), antisera from different animals reacted with different subunits of the 9-subunit RPI complex in Western blot analyses: 190 kD (n = 6); 120 kD (n = 1); 62 kD (n = 4); 45 kD (n = 2); and, no reactivity (n = 2). In contrast, animals immunized with normal IgG or CFA produced responses only against the immunogen. Together, these data suggest that anti-DNA and anti-RPI responses are connected through an autoimmune network in SLE.

Structural comparison of the Autographa californica nuclear polyhedrosis virus-induced RNA polymerase and the three nuclear RNA polymerases from the host, Spodoptera frugiperda.

A partial subunit structure has been determined for the novel RNA polymerase that is induced in fall armyworm (Spodoptera frugiperda) cells upon infection with the Autographa californica nuclear polyhedrosis virus (AcNPV). The putative structure includes nine polypeptides; the complexity of this structure is in accord with the high sedimentation coefficient (15S) estimated for this enzyme. A comparison of the putative structure of the virus-induced polymerase with those of the three host nuclear RNA polymerases shows that the structure of the viral polymerase is apparently unlike any of the host nuclear polymerases. This conclusion is reinforced by immunoblot experiments that show no cross-reactivity between the virus-induced polymerase and an antiserum directed against Drosophila RNA polymerase II. The virus-induced RNA polymerase appears at the onset of the late phase of infection and still appears when viral DNA synthesis is blocked by aphidicolin. Thus, the virus-induced polymerase seems to be composed of early viral products.

Anti-RNA polymerase I antibodies in the urine of patients with systemic lupus erythematosus.

Urine samples from patients with systemic lupus erythematosus (SLE) (n = 80), patients with rheumatoid arthritis (RA) (n = 21), and healthy controls (n = 36) were analyzed by radio-immunoassay (RIA) for anti-RNA polymerase I (RPI) antibodies. Significant levels of anti-RPI antibodies were detected in the urine of 46% of the patients with SLE but in only 19% of the patients with RA and in no sample from healthy individuals. The presence of anti-RPI antibodies in the urine was confirmed by demonstrating that IgG purified from the urine of patients with SLE was capable of inhibiting the transcriptional activity of RPI in vitro. If the quantity of anti-RPI antibodies excreted is related to disease activity, analysis of urine for these antibodies may be a useful alternative for the purpose of monitoring the progression of disease in individuals with SLE because of the ease by which the sample can be collected.

Monoclonal antibody against the lupus antigen Sm cross-reacts with RNA polymerase I.

Monoclonal anti-Sm antibody, a specificity directed against a constituent of nuclear ribonucleoprotein and considered to be a marker for systemic lupus erythematosus (SLE), was tested for its ability to react with four other rheumatic disease antigens of known enzymatic activity. No binding of the antibody was observed in radioimmunoassays with immobilized protein kinase NII, poly(A) polymerase, or topoisomerase I. In contrast, anti-Sm antibody did react with RNA polymerase I. Under conditions of antibody excess, anti-Sm was determined to bind RNA polymerase I on an equimolar basis, indicating that the polymerase possesses a single epitope recognized by the anti-Sm antibody. Addition of the anti-Sm antibody to in vitro transcription reactions resulted in inhibition of RNA polymerase I activity but had no effect on the reaction catalyzed by RNA polymerase II. When the subunits of RNA polymerase I were separated by polyacrylamide gel electrophoresis under denaturing conditions and incorporated individually into the radioimmunoassay, anti-Sm antibody bound only to the sixth polymerase polypeptide (Mr, 21,000). These data establish an immunological relationship between two important rheumatic disease antigens and help explain the apparent diversity of the autoimmune response in murine and human SLE.

Biochemistry of Fern Spore Germination: Globulin Storage Proteins in Matteuccia struthiopteris L.

Two globulin storage proteins have been identified in spores of the ostrich fern, Matteuccia struthiopteris (L.) Todaro. The two proteins comprise a significant amount of the total spore protein, are predominantly salt-soluble, and can be extracted by other solvents to a limited extent. The large 11.3 Svedberg unit (S) globulin is composed of five polypeptides with molecular weights of 21,000, 22,000, 24,000, 28,000 and 30,000. Each polypeptide has several isoelectric point (pI) variants between pH 5 and 7. The small 2.2S storage protein has a pI > 10.5 and is composed of at least two major polypeptides of 6,000 and 14,000 M(r). The amino acid composition of both storage proteins reveals that the 11.3S protein is particularly rich in aspartic and glutamic acid, while the 2.2S protein has few acidic amino acids. During imbibition and germination the globulin fraction declines rapidly, with a corresponding degradation of individual polypeptides of each protein. Polyclonal antibodies against each of the two proteins were produced and used for immunolocalization to determine the site of storage protein deposition within the quiescent spore. The proteins were sequestered in protein bodies of 2 to 10 micrometers, that are morphologically similar to those found in the seeds of flowering plants. The results suggest that spore globulins are biochemically similar to seed globulins, especially those found in some cruciferous seeds.

Autoantibodies against RNA polymerase I in scleroderma and Sjögren's syndrome sera.

Anti-RNA polymerase I antibodies were detected by radioimmunoassay in the sera of 8 out of 9 Sjögren's syndrome patients, 11 out of 19 individuals with scleroderma, and 19 out of 19 systemic lupus erythematosus patients. The results of the radioimmunoassay were confirmed by demonstrating the ability of the patients' IgG to inhibit RNA polymerase I activity in vitro. Sera from four patients in each category were also tested for reaction with individual subunits of RNA polymerase I. All 4 SLE sera, 3 out of 4 scleroderma sera and 1 out of 4 Sjögren's syndrome sera contained antibodies against the 65 kDa (S3) subunit of RNA polymerase I in addition to antibodies against one other subunit. Sera from the remaining scleroderma and Sjögren's syndrome patients tested in this assay contained only anti-S3 antibodies. These results demonstrate that anti-RNA polymerase I antibodies are characteristic of a variety of rheumatic autoimmune sera.

Anti-RNA polymerase I antibodies: potential role in the induction and progression of murine lupus nephritis.

Antibodies against RNA polymerase I were detected in plasma and kidney eluates of NZB/W mice. Plasma concentrations of the antibodies were the highest in mice with incipient nephritis and the lowest in mice with progressive nephritis. Mice with attenuated nephritis due to immunosuppressive therapy had intermediate plasma concentrations of the antibodies. The specific concentrations (ng/microgram IgG) of anti-RNA polymerase I antibodies in kidney eluates were significantly (10- to 70-fold) greater than the corresponding plasma concentrations. These results indicated that the decreased plasma concentration of the antibodies in mice with more advanced disease was at least partially due to selective concentration of anti-RNA polymerase I antibodies in the kidneys. The degree of this selective concentration was directly proportional (R2 = 0.9962) to the severity of renal disease, as reflected by the concentration (microgram/g tissue) of IgG eluted from the kidneys. The concentration (microgram/g tissue) of anti-RNA polymerase I eluted from the kidneys also was increased in mice with more severe renal disease. Further, the extent of this increase was greater than that of total IgG, again suggesting that anti-RNA polymerase I antibodies had been selectively concentrated in the kidneys. These findings are strongly suggestive of an important role for the RNA polymerase I/anti-RNA polymerase I antibody system in the pathogenesis of murine lupus nephritis.

Antibodies against nuclear poly(A) polymerases in rheumatic autoimmune diseases.

Sera from 53 patients, 26 with systemic lupus erythematosus (SLE), 8 with rheumatoid arthritis (RA), 9 with Sjogren's syndrome (SS), and 10 with scleroderma (Scl), were screened for the presence of antibodies against liver-type poly(A) polymerase and tumor-type poly(A) polymerase. Sixty percent of the patients with the above four autoimmune diseases have antibodies directed against liver poly(A) polymerase, whereas sera from 74% of the patients contained anti-hepatoma poly(A) polymerase antibodies. About 25% of the patients produced antibodies exclusively against the tumor poly(A) polymerase. IgG containing anti-liver or anti-tumor poly(A) polymerase antibodies inhibited the activity of the respective enzyme. IgG containing antibodies against liver and tumor enzymes inhibited the activity of both enzymes, whereas IgG from sera that did not react with poly(A) polymerase had no effect on either enzyme. These data demonstrated the specificity of these autoantibodies and confirmed the results of the radioimmunoassay.

Anti-RNA polymerase I antibodies in sera of MRL lpr/lpr and MRL +/+ autoimmune mice. Correlation of antibody production with delayed onset of lupus-like disease in MRL +/+ mice.

Sera from individual MRL/lpr and MRL/++ mice, which develop an autoimmune disease similar to human systemic lupus erythematosus (SLE), were screened over a period of approximately 30 wk for the presence of anti-RNA polymerase I and anti-ssDNA antibodies. Even though onset of the disease is delayed in MRL/++ as compared to MRL/lpr mice, anti-ssDNA antibodies were present in comparable concentrations in the sera of all mice by the age of 6 wk. As observed in sera of human SLE patients, anti-RNA polymerase I antibodies were detected in the sera of all MRL mice. However, unlike the anti-ssDNA antibodies, anti-RNA polymerase I antibodies were detected much later in MRL/++ mice (mean age, 22.8 wk) as compared to MRL/lpr mice (mean age, 9.6 wk). The presence of anti-RNA polymerase I antibodies in sera of MRL mice was thus a much better indicator of disease status than the presence of anti-ssDNA antibodies. The appearance and increase in anti-RNA polymerase I antibodies in the sera of MRL/++ mice correlated (R2 = 0.964) with a precipitous decrease in anti-ssDNA antibodies, starting at about 20 wk of age. These results suggest a possible relationship between the RNA polymerase I and DNA autoimmune reactions.

Immunization of rabbits with purified RNA polymerase I induces a distinct population of antibodies against nucleic acids as well as anti-RNA polymerase I antibodies, both characteristic of systemic lupus erythematosus.

Rabbits were immunized with either RNA polymerase I or poly(A) polymerase that had been purified to apparent homogeneity and was devoid of nucleic acids. Sera from rabbits thus immunized were screened for antibodies against nucleic acids. All seven rabbits injected with RNA polymerase I but none of the four rabbits immunized with poly(A) polymerase produced anti-nucleic acid antibodies. Anti-RNA polymerase I antibodies were induced after a single injection of the enzyme. Anti-polynucleotide antibodies were not detectable until after the second immunization. Anti-RNA polymerase I antibodies could be detected with as little as 100 pg of purified RNA polymerase I in the radioimmunoassay. At least 50 ng of poly(A) or 200 ng of DNA was required to detect anti-nucleic acid antibodies. The immunoreactivity of anti-RNA polymerase I antisera was greater with synthetic polynucleotides than with DNA, particularly early in the immunization schedule. Alkaline phosphatase treatment of poly(A) to remove 5' phosphates nearly abolished its antigenicity with respect to the early sera and decreased antibody binding of later sera by 60%. These results indicate that the anti-nucleic acid antibodies produced early were primarily directed against determinants including the 5'-terminal phosphates while antibodies produced later were directed against other sites. The antinucleic acid antibodies and anti-RNA polymerase I antibodies formed two distinct populations that were not immunologically crossreactive. We suggest that after injection, RNA polymerase I becomes associated with the nucleic acids present in blood plasma which renders them immunogenic; thus, association of nucleic acids with autoimmunogenic RNA polymerase I may be one of the mechanisms by which anti-DNA antibodies are induced in systemic lupus erythematosus.

Comparison of cytosolic and nuclear poly(A) polymerases from rat liver and a hepatoma: structural and immunological properties and response to NI-type protein kinases.

Poly(A) polymerases were purified from the cytosol fraction of rat liver and Morris hepatoma 3924A and compared to previously purified nuclear poly(A) polymerases. Chromatographic fractionation of the hepatoma cytosol on a DEAE-Sephadex column yielded approximately 5 times as much poly(A) polymerase as was obtained from fractionation of the liver cytosol. Hepatoma cytosol contained a single poly(A) polymerase species [48 kilodaltons (kDa)] which was indistinguishable from the hepatoma nuclear enzyme (48 kDa) on the basis of CNBr cleavage maps. Liver cytosol contained two poly(A) polymerase species (40 and 48 kDa). The CNBr cleavage patterns of these two enzymes were distinct from each other. However, the cleavage pattern of the 40-kDa enzyme was similar to that of the major liver nuclear poly(A) polymerase (36 kDa), and approximately three-fourths of the peptide fragments derived from the 48-kDa species were identical with those from the hepatoma enzymes (48 kDa). NI-type protein kinases from liver or hepatoma stimulated hepatoma nuclear and cytosolic poly(A) polymerases 4-6-fold. In contrast, the liver cytosolic 40- and 48-kDa poly(A) polymerases were stimulated only slightly or inhibited by similar units of the protein kinases. Antibodies produced in rabbits against purified hepatoma nuclear poly(A) polymerase reacted equally well with hepatoma nuclear and cytosolic enzyme but only 80% as well with the liver cytosolic 48-kDa poly(A) polymerase and not at all with liver cytosolic 40-kDa or nuclear 36-kDa enzymes. Anti-poly(A) polymerase antibodies present in the serum of a hepatoma-bearing rat reacted with hepatoma nuclear and cytosolic poly(A) polymerases to the same extent but only 40% as well with the liver cytosolic 48-kDa enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

Structural and immunological identity of rat hepatoma and fetal liver nuclear poly(A) polymerase.

Poly(A) polymerase was partially purified from isolated nuclei of fetal rat liver. Antibodies produced in rabbits immunized with purified nuclear poly(A) polymerase from a rat hepatoma exhibited nearly identical affinity for the partially purified fetal liver and hepatoma enzymes. The extent of the antibody reaction with adult liver nuclear poly(A) polymerase partially purified in a similar manner was only 1.4% of that obtained with the hepatoma enzyme. Immune complex formation was observed between the antibodies and a major polypeptide in the fetal liver enzyme preparation which corresponded to the hepatoma enzyme (mol. wt. 48 000). No other polypeptide in the fetal liver enzyme preparation reacted with the antibodies. The 48-kDa fetal liver polypeptide produced a CNBr cleavage pattern identical to that of hepatoma poly(A) polymerase which is known to be different from the cleavage pattern of the adult liver major nuclear poly(A) polymerase. A fetal liver polypeptide corresponding to the adult liver enzyme (mol. wt. 38 000) was not evident. These results coupled with other data suggest that the hepatoma nuclear poly(A) polymerase is an oncofetal protein.

Purification and characterization of a nuclear protein kinase from rat liver and a hepatoma that is capable of activating poly(A) polymerase.

The cyclic nucleotide-independent protein kinase which is separated from poly(A) polymerase during its purification from nuclei of rat liver and Morris hepatoma 3924A was purified essentially to homogeneity. Liver nuclear poly(A) polymerase was dissociated from protein kinase by phosphocellulose column chromatography. In contrast, protein kinase copurified with the hepatoma poly(A) polymerase on the phosphocellulose column. Neither liver nor hepatoma kinase was stimulated by spermine or inhibited by heparin. These enzymes did not utilize GTP as phosphoryl donor, or histones or tyrosine-containing [Val5]-angiotensin II as phosphoryl acceptors. The apparent Km with respect to ATP was similar for the liver (4.7 microM) and hepatoma (11 microM) kinases, and the apparent Km with respect to casein was identical (0.6 microgram/microliter) for these enzymes. Both enzymes were capable of phosphorylating poly(A) polymerase and stimulating both tumor and liver poly(A) polymerase activity. However, in addition to their different chromatographic properties, the two kinases differed in molecular weight (liver, 37,000; hepatoma, 56,000), in their response to various divalent metal ions, and in their ability to phosphorylate hepatoma poly(A) polymerase (Km 7.9 and 30 microgram/microliter for liver and hepatoma enzymes, respectively). These latter characteristics distinguished the liver and hepatoma protein kinases from each other as well as from the previously described NI protein kinase.

Phosphorylation of RNA polymerase I augments its interaction with autoantibodies of systemic lupus erythematosus patients.

Purified RNA polymerase I was phosphorylated by the endogenous protein kinase or dephosphorylated by alkaline phosphatase and used as antigen in a radioimmunoassay with sera from systemic lupus erythematosus patients or serum from an immunized rabbit. Enzyme incubated in the absence of ATP or phosphatase served as control. Three to seven times more of the autoantibodies in the patients' sera reacted with phosphorylated RNA polymerase I than with control enzyme. The reactivity of the dephosphorylated enzyme with lupus autoantibodies was only 50-60% of that observed with control enzyme. Neither phosphorylation nor dephosphorylation of the enzyme had an effect on its reaction with the rabbit antibodies. The effect of phosphorylation on the reaction of each RNA polymerase I subunit (S1-S8; Mr = 190,000-17,000) with the patients' antibodies was determined by an immunoblot procedure following resolution of the subunits on polyacrylamide gels. Prior phosphorylation of the enzyme resulted in a dramatic increase in binding of each patient's antibodies to all polymerase subunits with the exception of S4. Anti-S4 antibody was not detected with either phosphorylated or control enzyme. Strikingly, antibodies in each patients' sera reacted with S6 only after its phosphorylation. Similarly, anti-S5 antibodies in the serum of one patient were only detected with phosphorylated RNA polymerase I. The present data suggest that at least a significant fraction of the anti-RNA polymerase I autoantibodies in the sera of systemic lupus erythematosus patients might be directed against phosphorylated sites on the enzyme and that phosphorylation may have a role in the production of this and other autoimmunogenic nuclear components which are hallmarks of this disease.

Induction of a distinct nuclear poly(A) polymerase and the production of anti-tumor poly(A) polymerase antibodies in hepatocarcinogenesis.

Primary hepatomas induced in rats by an azo dye contained a distinct nuclear poly(A) polymerase which was different from the corresponding normal liver enzyme with respect to molecular weight and peptide map. The two enzymes could also be distinguished by their immunological characteristics. Sera from the carcinogen-fed rats contained antibodies to the tumor poly(A) polymerase even before the appearance of neoplastic nodules.

Structurally and immunologically distinct poly(A) polymerases in rat liver. Occurrence of a tumor-type enzyme in normal liver.

Poly(A) polymerases purified from rat liver nuclei consisted of two distinct species, a predominant enzyme of Mr = 38,000 and a minor one of Mr = 48,000. Prior to extensive purification, the minor enzyme constituted approximately 1% of the total liver poly(A) polymerase. Poly(A) polymerase purified from a rat tumor, Morris hepatoma 3924A, was comprised of a single species of Mr = 48,000 which was identical to the minor liver enzyme with respect to chromatographic and immunological characteristics. Gel filtration on Sephacryl S-200 using 0.3 M NaCl for elution showed that the major liver poly(A) polymerase had a molecular weight of 156,000, which corresponded to a tetramer of the 38-kDa polypeptide, whereas the hepatoma and minor liver 48-kDa species existed as dimers with a molecular weight of 96,000. Fractionation by Sephacryl S-200 resulted in complete loss of both liver poly(A) polymerase activities which could be restored by exogenous N1-type protein kinase. Following CNBr cleavage, the 48-kDa poly(A) polymerase from liver and hepatoma exhibited nearly identical peptide maps which were distinct from that of the major liver enzyme (38 kDa). Antibodies raised against tumor poly(A) polymerase reacted with the 48-kDa polypeptide but not with the 38-kDa liver enzyme. Immune complex formation was observed between seven of the eight CNBr cleavage products derived from the 48-kDa polypeptide of both liver and hepatoma. It is concluded that distinct genes in rat liver code for two structurally and immunologically unique nuclear poly(A) polymerases, one of which is identical to the enzyme from the hepatoma.

Absence of inactivation or phosphorylation of ornithine decarboxylase by nuclear protein kinase NII and of immunological cross-reactivity between RNA polymerase I and ornithine decarboxylase.

Incubation with protein kinase NII did not result in phosphorylation or inactivation of mouse kidney ornithine decarboxylase. Partially purified ornithine decarboxylase preparations contained a protein kinase activity and stimulated the activity of RNA polymerase I. However, these properties were due to contaminating protein(s) since further purification reduced the kinase activity and removal of the ornithine decarboxylase with a specific antiserum did not abolish the ability to stimulate RNA polymerase I. Antibodies to RNA polymerase I did not interact with ornithine decarboxylase and antibodies to ornithine decarboxylase did not interact with RNA polymerase I. These results indicate that: a) mammalian ornithine decarboxylase activity is not regulated by phosphorylation by protein kinase NII or the contaminating kinase, and b) the ability of impure preparations of ornithine decarboxylase to stimulate RNA polymerase I is due to a contaminating unrelated protein.