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R Richman

Publications and source records attributed to R Richman.

At least 37 records · Page 2Linked to original sources
PubMed1986

hv1 is an evolutionarily conserved H2A variant that is preferentially associated with active genes.

Polyclonal antibodies to the Tetrahymena macronuclear-specific histone variant hv1 cross-react with histone-like molecules from yeast, wheat, and mouse. A novel purification scheme has allowed isolation of sufficient hv1 to enable determination of the sequence of 61 amino-terminal residues as well as 27 additional internal residues. These data clearly demonstrate that hv1 shares a number of conserved sequence elements with the H2A family of histones. Comparison of hv1 with H2A.F (= H2A.Z = M1), another evolutionarily conserved H2A variant whose sequence is known, reveals that they share an unblocked amino-terminal alanine (instead of acetylserine) and a distinctive structure in a "variant box" region that distinguishes them from major H2As. In addition, 10 residues have been identified which are identical (or highly similar) in hv1 and H2A.F, but are different from residues conserved in the major H2As. Therefore, in many ways hv1 resembles chick H2A.F more than the major Tetrahymena H2A. The sites of acetylation of hv1 also differ from those of the major Tetrahymena H2As. In spite of their similarities, hv1 and H2A.Z differ significantly in their amino termini, and antibodies against hv1 do not react with H2A.Z. Interestingly, the nucleolar staining pattern reported with anti-hv1 serum is similar to that reported for an antiserum to another H2A variant, mouse testes-enriched H2A.X. Since both H2A.Z and hv1 appear to be enriched in transcriptionally active chromatin, these results suggest that there may be a number of different, functionally distinct, nonallelic variants in the H2A family of histones and that hv1 is a hybrid H2A variant with properties of both vertebrate H2A.Z and H2A.X.

Acetylation↗
PubMed1986

Nonrandom utilization of acetylation sites in histones isolated from Tetrahymena. Evidence for functionally distinct H4 acetylation sites.

Macro- and micronuclei of the ciliated protozoan Tetrahymena thermophila afford a unique opportunity to study histone acetylation under conditions where postsynthetic "transcription"-related acetylation and synthetic "deposition"-related acetylation are nonoverlapping. Recent studies have demonstrated that at least two general systems of acetylation operate in Tetrahymena. One is postsynthetic, macronuclear specific, and may be related to gene expression in that nucleus (Vavra, K. J., Allis, C. D., and Gorovsky, M. A. (1982) J. Biol. Chem. 257, 2591-2598). The other is synthetic, common to macro- and micronuclei, and is likely related to histone deposition during replication (Allis, C. D., Chicoine, L. G., Richman, R., and Schulman, I. G. (1985a) Proc. Natl. Acad. Sci. U. S. A., 82, 8048-8052). A unique feature of H3 and H4 in Tetrahymena is that neither are blocked at their amino termini. We have exploited this fact as well as the resolving capability of acid-urea gel electrophoresis and current microsequencing techniques to examine whether utilization of different NH2-terminal acetylation sites is random or nonrandom during the progression toward high acetylation states. Of the four acetylation sites which have been identified in H4 (in Tetrahymena these are lysines at positions 4, 7, 11, and 15), we find that lysine 7 is the exclusive site of postsynthetic acetylation in populations of monoacetylated H4 isolated from macronuclei. This site is retained in populations of diacetylated H4, which are acetylated exclusively at lysines 4 and 7. Our data also suggest that there is some preference for using lysine 11 (as compared to 15) as the third site of acetylation in triacetylated molecules. The data demonstrate that the postsynthetic acetylation-deacetylation process is surprisingly nonrandom for H4 in Tetrahymena macronuclei. We have also investigated the same question with macronuclear H3 (which contains acetylation sites at lysines 9, 14, 18, and 23). Our data demonstrate that unlike H4, lysines at position 9 or 14 are likely to be utilized as sites of acetylation within a population of monoacetylated H3. Both of these acetylation sites are retained in diacetylated H3 which suggests that if site 9 is used initially as the site of monoacetylation, 14 is used secondarily (and vice versa). Our data show, moreover, that there is a preference for utilizing lysine 18 as the third acetylation site (in triacetylated H3). Thus, these data show that H3 is also acetylated in a nonrandom fashion in macronuclei. Finally, we have determined which acetylation sites are utilized in macro- or micronuclear H4 when it is undergoing active synthesis and deposition.(ABSTRACT TRUNCATED AT 400 WORDS)

Acetylation↗
PubMed1986

An intervening sequence in an unusual histone H1 gene of Tetrahymena thermophila.

An intervening sequence of 254 base pairs interrupts the coding region of the single gene for macronuclear histone H1 of the ciliated protozoan, Tetrahymena thermophila. The intervening sequence has splice junctions similar to those found in RNA polymerase II genes of other organisms. No obvious similarities are observed between this intron and the self-splicing intervening sequence of the Tetrahymena ribosomal gene. The derived amino acid sequence describes a small extremely basic H1 protein missing most of the central hydrophobic domain that is conserved in all other H1 proteins. Macronuclei divide amitotically, without chromosome condensation, suggesting the conserved globular domain of H1 plays a role in higher-order chromatin structure.

Amino Acid Sequence↗
PubMed1985

Modulation of linker histones during development in Tetrahymena: selective elimination of linker histone during the differentiation of new macronuclei.

Macronuclei of Tetrahymena thermophila contain a typical H1 which has been shown to be missing from micronuclei. Instead, micronuclei contain three unique polypeptides, alpha, beta, and gamma, which are associated with linker regions of micronuclear chromatin. In this report polyclonal antibodies raised against macronuclear H1 are shown to react with alpha, beta, and gamma by immunoblotting analyses. This result suggests that these polypeptides share some common structural feature(s). Also consistent with this result is the finding that both macro- and micronuclei in growing and mating cells stain positively with H1 antibodies by in situ indirect immunofluorescence. However, these analyses demonstrate that the level of linker histone is greatly reduced in the micronucleus of starved cells and in young macronuclear anlagen. These results are in agreement with earlier biochemical studies and together provide strong evidence that dramatic changes in linker histone accompany nuclear differentiation (and dedifferentiation) in Tetrahymena.

Animals↗
PubMed1985

Deposition-related histone acetylation in micronuclei of conjugating Tetrahymena.

Macro- and micronuclei of the ciliated protozoan, Tetrahymena thermophila, afford a unique opportunity to study histone acetylation under conditions where acetylation associated with the regulation of transcription and acetylation associated with the deposition of histones on the DNA are separable. In this study we demonstrate that histone H3 and histone H4 synthesized in young (5 hr) conjugating Tetrahymena are deposited into micronuclei in acetylated forms. Most of the newly synthesized histone H3 migrates as a monoacetylated form while essentially all of the new histone H4 is deposited as a diacetylated species. Since micronuclei replicate rapidly during this stage of the life cycle, but are transcriptionally inactive, these data suggest that histone acetylation is related functionally to histone deposition and chromatin assembly. Pulse-chase experiments show that micronuclei also contain a butyrate-sensitive deacetylase activity(ies) which operates to remove the deposition-related acetate groups from newly synthesized and deposited H3 and H4. This enzymatic activity probably contributes to the steady state level of micronuclear histone acetylation that is low or nonexistent. Thus, evidence is emerging for at least two independent systems of histone acetylation in Tetrahymena. The first system is specific to macronuclei and may be related to gene expression. The second system is common to macro- or micronuclear histones (H3 and H4) and may be related to histone deposition during DNA replication.

Acetylation↗
PubMed1984

The effect of induction of hemoglobin synthesis in cultured Friend cells on the number of initiation sites for replication and transcription.

It was found that differentiation of murine erythroleukemia cells can be induced by 5-fluorodeoxyuridine (FudR), amethopterin and alpha-aminoisobutyrate. Each of these compounds is believed to delay the onset of DNA synthesis. Since relief of the FudR block to DNA synthesis by addition of thymidine can increase the number of initiation sites for replication (Taylor, 1977), the effect of various inducers and inhibitors of differentiation of Friend cells upon the relative number of initiation sites for replication and transcription was investigated. Very efficient inducers of hemoglobin synthesis, hexamethylene bisacetamide (HMBA) and dimethylsulfoxide (DMSO), increase the number of initiation sites for transcription and HMBA also increases the number of functional initiation sites for replication. Two other compounds that induce differentiation of Friend cells, low levels of actinomycin D and FudR, did not increase the number of initiation sites for transcription. Compounds that prevent induction of hemoglobin synthesis by HMBA and DMSO include 5-bromodeoxyuridine (BrdU) and novobiocin. Both of these compounds were found to decrease the number of functional initiation sites for transcription and it is known that both compounds reduce the number of initiation sites for replication. The relation between initiation of replication and transcription, and its effect upon differentiation of erythroleukemia cells is discussed.

Acetamides↗
PubMed1984

Abnormalities of thyroid function in infants with Down syndrome.

We describe 12 of 1130 infants with Down syndrome in whom various degrees of thyroid dysfunction were detected by neonatal screening. These aberrations were confirmed subsequently in 11 patients. In eight of 11 children, persistent primary hypothyroidism, was diagnosed, whereas in the remaining three patients transient thyroid abnormalities were noted. The twelfth patient died and could not be retested. We found an incidence of persistent primary congenital hypothyroidism in infants with Down syndrome of 1:141, or about 28 times more than in the general population. The cause of thyroid aberrations in these infants remains unclear; none of the studied patients had agenesis or ectopia of the thyroid gland. On initial screening most infants with Down syndrome had only mild biochemical abnormalities, with gradual decompensation occurring thereafter. Infants with Down syndrome are therefore at high risk for congenital hypothyroidism and should have careful follow-up to prevent further deterioration of their mental development or growth.

Child, Preschool↗
PubMed1984

Proteolytic processing of h1-like histones in chromatin: a physiologically and developmentally regulated event in Tetrahymena micronuclei.

Micronuclei isolated from growing cells of Tetrahymena thermophila contain three H1-like polypeptides alpha, beta, and gamma. Micronuclei isolated from young conjugating cells (3-7 h) also contain a larger molecular weight polypeptide, X, which is being actively synthesized and deposited into these nuclei (Allis, C. D., and J. C. Wiggins, 1984, Dev. Biol., 101:282-294). Pulse-chase experiments (with growing and conjugating cells) suggested that X is a precursor to alpha and that alpha is further processed to gamma and a previously undescribed and relatively minor species, delta. These precursor-product relationships were supported by cross-reactivity with polyclonal antibodies raised against alpha and peptide mapping. While beta consistently became labeled under chase conditions (both in growing and mating cells), it was not clear whether it is part of the vivo processing event(s) which interrelates X, alpha, gamma, and delta. Beta was not recognized by alpha antibodies. Despite this uncertainty, these results suggest that proteolytic processing serves to generate significant changes in the complement of H1-like histones present in this nucleus.

Animals↗
PubMed1982

Urinary tract injury during cesarean section.

Lower urinary tract injury at the time of cesarean section is an uncommon complication. During a 5-year period, the incidence of bladder and ureter injuries at the Los Angeles County/University of Southern California Medical Center was .31 and .09%, respectively. When bladder injury occurs, it usually is due to surgical difficulty encountered while developing the bladder flap over the lower uterine segment. The difficulty is caused usually by scar tissue from previous surgery. Ureteral injury is a rare complication of cesarean section. It is attributable most often to ureteral transection or ligation associated with uterine incision extensions in the lower uterine segment or the vagina, and to attempts to achieve hemostasis. The data presented in this report indicate that cystotomy, when adequately repaired, is not associated with any complications. Furthermore, diagnostic cystotomy with intravenous injection of indigo carmine is a rapid, safe method of evaluating ureteral patency.

Cesarean Section↗
PubMed1981

The effect of added H1 histone and polylysine on DNA synthesis and cell division of cultured mammalian cells.

Addition of H1 histone or polylysine (10 microgram/ml) to cultured Friend erythroleukemia cells or to two mouse lymphoma cell lines (el-4 and S-49) increased levels of cell division in these cultures. There is a stimulation of incorporation of labeled thymidine into DNA in cultures containing H1 histone and polylysine. DNA fiber autoradiographic experiments revealed that replicon size is decreased in the cells cultured with H1 histone and polylysine at later periods of culture.

Animals↗
PubMed1976

Altered thyroidal responsivity to thyrotropin induced by circulating thyroid hormones. A "short-loop" regulatory mechanism?

We studied the effect of thyroid hormone administration on responsivity of murine thyroid to exogenous thyrotropin (TSH) in order to explore the possibility that the thyroid gland might be directly inhibited by its own hormones. In the rat both L-thyroxine (T4) and 3,5,3'-L-triiodothyronine (T3) pretreatment inhibited TSH-induced thyroidal ornithine decarboxylase (ODC) activity in vivo in a dose-related manner (half-maximal inhibition, 1.7 mug/rat and 0.6 mug/rat, respectively). Other structurally related compounds exhibited the following inhibitory potencies compared to T4: T3, 283%; triiodothyroacetic acid, 40%; D-T4, 18%; 3,5-L-diiodothyronine, 9%. Monoiodotyrosine, diiodotyrosine, and iodide were not inhibitory. The full inhibitory effect of T4 or T3 was observed when thyroid hormone was administered from 96 to 12 h before TSH and was also seen in hypophysectomized animals. Pretreatment with T4 or T3 in divided doses over 2 1/2 days inhibited TSH-induced increase in [1-14C]glucose oxidation to 14C02 and [3H] leucine incorporation into protein in rat thyroid. In the mouse T4 or T3 pretreatment (0.25-25 mug daily) caused dose-related inhibition of both thyroidal ODC activity and 131I release induced by TSH in vivo. In mice on a low-iodine diet (LID) but not in animals on a regular diet (RD) NaI pretreatment also blunted TSH-induced thyroidal ODC activation and 131I release. When LID or RD mice were pretreated with 12.5-125 mug of T4 or T3 over 2 1/2 days, TSH-induced in vitro stimulation of thyroid cyclic 3',5'-adenosine monophosphate formation was inhibited in a dose-related manner; NaI pretreatment was inhibitory in the LID mouse only. Prior administration of exogenous TSH blunted the activation of thyroid ODC and thyroid hormone release induced by subsequent TSH administration in rat and mouse. These studies indicate altered thyroid responsivity to TSH under the influence of circulating thyroid hormones and suggest the existence of a "short-loop" negative feedback regulating thyroid function.

Animals↗
PubMed1976

Fine needle cholangiography in the diagnosis of persistent jaundice. A case report.

The fine needle technic utilizes a sleeveless, thin, flexible needle for transhepatic cholangiography. This needle can enter small or normal bile ducts in a high percentage of patients. The value of fine needle cholangiography is illustrated in a patient with a stenosis of the common hepatic duct and a left hepatodochalduodenal fistula which eluded diagnosis by all other methods. The use of this technic is described as an alternative procedure to endoscopic retrograde cholangiography when the latter is technically impossible or as an adjunct to intraoperative extrahepatic cholangiography.

Adult↗
PubMed1975

Regulation of thyroid ornithine ornithine decarboxylase (ODC) by thyrotropin. I. The rat.

We studied the effects of TSH on rat thyroid ornithine decarboxylase (ODC) activity. After 1 day of goitrogen treatment, there was an abrupt fall in serum triiodothyronine (T3) a rise in circulating TSH, and a dramatic increase in thyroid ODC activity. Despite the continued rise in TSH and progressive increase in thyroid gland size with further treatment, thyroid ODC activity declined on the third day and remained at submaximal levels. Thyroid ODC activity was also stimulated in a dose-related manner by administration of exogenous TSH. Little TSH effect was noted before 3 h. Maximal ODC activity occurred between 4 and 5 h. The TSH stimulation of ODC could be inhibited by pretreatment with actinomycin D or cycloheximide, suggesting that the increase in ODC activity requires new RNA and protein synthesis. Although pretreatment with agents that alter microtubule structure (e.g., colchicine and vinblastine) prevent stimulation of ODC activity by TSH, additional data suggest, but do not confirm, that hrmone secretion and ODC activation may be dissociable. Further studies were undertaken to determine whether cyclic AMP (cAMP) or prostaglandins played any role in the regulation of thyroidal ODC activity. Dibutyryl cAMP, alone, or together with aminophylline, did not stimulate thyroidal ODC activity in dosages which concomitantly stimulated adrenal enzyme activity. Likewise, prostaglandin E2 (PGE2) did not stimulate thyroidal ODC activity, but did stimulate adrenal enzyme activity in a dose-related manner. However, pre-treatment of rats with inhibitors of prostaglandin synthesis prevented the activation of thyroidal ODC BY TSH. One inhibitor, indomethacin, attenuated the TSH stimulation of enzyme activity in a dose-related manner. Indomethacin pretreatment also resulted in approximately a 10-fold decrease in thyroidal prostaglandin levels. Exogenous PGE9, in dosages as high as 500 pg, did not overcome the inhibitory effect of indomethacin on ODC activation. Although the precise role for endogenous prostaglandins remains to be defined, it does appear that a reduction in thyroidal prostaglandins prevents activation of the enzyme by TSH.

Animals↗
PubMed1973

Regulation of adrenal ornithine decarboxylase by adrenocorticotropic hormone and cyclic AMP.

Adrenal ornithine decarboxylase activity was stimulated in a dose-related manner after administration of ACTH or dibutyryl ((6)N-2'-O-dibutyryl) cyclic AMP to hypophysectomized rats. Little effect was observed for 2 h, but striking increases in enzyme activity were observed 4 h after administration of these substances. Effects of ACTH and dibutyryl cyclic AMP were not secondary to stimulation of steroidogenesis, since hydrocortisone had no effect on adrenal ornithine decarboxylase although it did stimulate activity of the enzyme in the liver and kidney.ACTH, given subcutaneously to hypophysectomized rats, induced striking increases in adrenal cyclic AMP levels within 15-30 min with a fall towards the base line in 1 h. Increases in ornithine decarboxylase activity lag several hours after this endogenous cyclic AMP peak, in contrast to the stimulatin of steroidogenesis by the nucleotide that requires only 2-3 min. After graded doses of ACTH, increases in adrenal cyclic AMP levels at 30 min were paralleled by proportional increases in adrenal ornithine decarboxylase activity 4 h after hormone treatment. Whereas maximal levels of adrenal steroidogenesis have been observed at tissue cyclic AMP levels of 6 nmol/g. ACTH is capable of inducing increases in nucleotide levels up to 200 nmol/g or more. These high tissue levels of cyclic AMP, although unneccessary for maximal steroidogenesis, appear to stimulate adrenal ornithine decarboxylase activity. Several results in addition to the time lag in the stimulation of ornithine decarboxylase activity suggest a mechanism involving accumulation of the enzyme or some factor needed for its activity rather than direct activation of the enzyme by cyclic AMP. Thus, the addition of cyclic AMP directly to the ornithine decarboxylase assay mixture in vitro was without stimulatory effect. In addition, actinomycin D or cycloheximide in doses sufficient to block adrenal RNA and protein synthesis, respectively inhibited the stimulation of ornithine decarboxylase activity by ACTH in vivo. An adrenocortical cancer was found to maintain ornithine decarboxylase activity at very high levels, but did so at much lower cyclic AMP levels than those of ACTH-stimulated adrenals. It is concluded that ACTH stimulates adrenal ornithine decarboxylase activity and that this effect may be mediated by cyclic AMP. However, cyclic AMP be mediated by appear to be a determinant of the high level of enzyme activity found in adrenocortical cancer.

Adrenal Gland Neoplasms↗