A two-point linkage analysis was performed between blood group (14), allotype (8), polymorphic protein (11), DNA type I (2), and microsatellite (2) loci in Wild Boar x Pietrain and Meishan x Pietrain three-generation families. The following new pairwise linkages were detected: LPR-EAN (Zmax = 60.68, theta = 0.055), EAD-GH1 (Zmax = 17.43, theta = 0.246), EAO-P3 (Zmax = 15.81, theta = 0.239), and P3-S0003 (Zmax = 5.43, theta = 0.312). This study and published mapping data enabled the localization of LPR (LPR allotype) to chromosome 9, EAD (erythrocyte antigen D) to chromosome 12, and EAO (erythrocyte antigen O) and P3 (P3 allotype) to the q arm of chromosome 6 with gene order S0003-P3-EAO, EAO being the most distal.

Basal and stimulated activity of the c-fos promoter is reduced by triiodothyronine (T3) and retinoic acid (RA) in GH1 cells. We examined the influence of these ligands on the activity of reporter constructs containing the AP-1 site, the serum response element (SRE) and the cyclic AMP responsive element (CRE) of the c-fos promoter under control of an heterologous promoter. T3 and RA decreased the response of AP-1 and SRE sequences to phorbol esters, forskolin or serum but they did not reduce basal or forskolin-stimulated activity mediated by the CRE. Therefore, repression of c-fos gene expression by T3 and RA receptors appears to be exerted through transcriptional interference with the SRE and the AP-1 binding site of the promoter.

Growth hormone, GH1, or growth hormone and prolactin, GH3, secreting tumors were induced in 1 week old female Wistar-Furth rats and 17 week old female Wistar-Furth rats. At 11 or 30 weeks of age the parametrial fat pad (PFP) was weighed and a portion fixed with osmium tetroxide for cell quantitation by a Coulter counter technique. Tumor induction increased the body weight of both young and mature rats by 50 and 100% respectively when compared to age matched controls. Serum GH was elevated about 100 fold in both young and old treated rats when compared to controls. Tumor induction decreased the weight of the PFP in both young and old rats when compared to controls but the effect was only significant in the GH3 tumor bearing rats weights. The PFP made up a much smaller percentage of the body weight in all the treated groups when compared to respective age matched controls. Mean PFP adipocyte number was not affected by treatment or age. Tumor induction decreased the mean diameter and mean volume of adipocytes in both young and old rats of both treatment groups compared to age matched controls. The distribution of adipocytes by cell diameter was shifted toward smaller cells in the tumor-bearing rats. In conclusion, induction of GH1 or GH3 tumors elevated serum GH concentrations, increased body weight and decreased relative fat pad weight but did not affect adipocyte cell number. Hence, the proposed adipogenic function of GH does not appear to function in this model in vivo.

The group A streptococcus Streptococcus pyogenes is the causative agent of a wide spectrum of invasive infections, including necrotizing fasciitis, scarlet fever and toxic shock syndrome. In the context of its carbohydrate chemistry, it is interesting that S. pyogenes (in this work strain M1 GAS SF370) displays a spectrum of oligosaccharide-processing enzymes that are located in close proximity on the genome but that the in vivo function of these proteins remains unknown. These proteins include different sugar transporters (SPy1593 and SPy1595), both GH1GH1 `putative β-glucosidase'. Here, the solution of the three-dimensional structure of SPy1599 in a number of crystal forms complicated by unusual crystallographic twinning is reported. The structure is a classical (β/α)(8)-barrel, consistent with CAZy family GH1 and other members of the GH-A clan. SPy1599 has been annotated in sequence depositions as a β-glucosidase (EC 3.2.1.21), but no such activity could be found; instead, three-dimensional structural overlaps with other enzymes of known function suggested that SPy1599 contains a phosphate-binding pocket in the active site and has possible 6-phospho-β-glycosidase activity. Subsequent kinetic analysis indeed showed that SPy1599 has 6-phospho-β-glucosidase (EC 3.2.1.86) activity. These data suggest that SPy1599 is involved in the intracellular degradation of 6-phosphoglycosides, which are likely to originate from import through one of the organism's many phosphoenolpyruvate phosphotransfer systems (PEP-PTSs).

BACKGROUND

Mutations in the GH1 promoter are a rare cause of isolated growth hormone deficiency (IGHD).

OBJECTIVE

To identify the molecular aetiology of a family with IGHD.

METHODS

DNA sequencing, electromobility shift (EMSA) and luciferase reporter assays.

METHODS

University Hospital.

METHODS

Three siblings (2M) born to consanguineous parents presented with IGHD with normal pituitary on MRI.

METHODS

The GH1 proximal promoter, locus control region, five exons and four introns as well as GHRHR gene were sequenced in genomic DNA by Sanger method. DNA-protein interaction was evaluated by EMSA in nuclear extracts of GH3 pituitary cells. Dual-luciferase reporter assays were performed in cells transiently transfected with plasmids containing four different combinations of GH1 allelic variants (AV).

RESULTS

The patients harboured two homozygous variants (c.-185T>C and c.-223C>T) in the GH1 promoter within a highly conserved region and predicted binding sites for POU1F1/SP1 and SP1 respectively. The parents and brother were carriers and these variants were absent in 100 controls. EMSA demonstrated absent binding of GH3 nuclear extract to the c.-223C>T variant and normal binding of both POU1F1 protein and GH3 nuclear extract to the c.-185T>C variant. In contrast to GH1 promoter with AV only at c.-185, the GH1 promoter containing the AV only at c.-223 and at both positions drove significantly less expression of luciferase compared with the promoter containing either positions wild type in luciferase reporter assays.

CONCLUSIONS

To our knowledge, c.-223C>T is the first homozygous point mutation in the GH1 promoter that leads to short stature due to IGHD.

Lactose is an inevitable byproduct of the dairy industry. In addition to cheese manufacturing, the growing Greek yogurt industry generates excess acid whey, which contains lactose. Therefore, rapid and efficient conversion of lactose to fuels and chemicals would be useful for recycling the otherwise harmful acid whey. Saccharomyces cerevisiae, a popular metabolic engineering host, cannot natively utilize lactose. However, we discovered that an engineered S. cerevisiae strain (EJ2) capable of fermenting cellobiose can also ferment lactose. This finding suggests that a cellobiose transporter (CDT-1) can transport lactose and a β-glucosidase (GH1-1) can hydrolyze lactose by acting as a β-galactosidase. While the lactose fermentation by the EJ2 strain was much slower than the cellobiose fermentation, a faster lactose-fermenting strain (EJ2e8) was obtained through serial subcultures on lactose. The EJ2e8 strain fermented lactose with a consumption rate of 2.16g/Lh. The improved lactose fermentation by the EJ2e8 strain was due to the increased copy number of cdt-1 and gh1-1 genes. Looking ahead, the EJ2e8 strain could be exploited for the production of other non-ethanol fuels and chemicals from lactose through further metabolic engineering.

Gallic acid production in a batch bioreactor was evaluated using as catalytic material the mouldy polyurethane solids (MPS) obtained from a solid-state fermentation (SSF) bioprocess carried out for tannase production by Aspergillus niger GH1 on polyurethane foam powder (PUF) with 5 % (v/w) of tannic acid as inducer. Fungal biomass, tannic acid consumption and tannase production were kinetically monitored. SSF was stopped when tannase activity reached its maximum level. Effects of washing with distilled water and drying on the tannase activity of MPS were determined. Better results were obtained with dried and washed MPS retaining 84 % of the tannase activity. Maximum tannase activity produced through SSF after 24 h of incubation was equivalent to 130 U/gS with a specific activity of 36 U/mg. The methylgallate was hydrolysed (45 %) in an easy, cheap and fast bioprocess (30 min). Kinetic parameters of tannase self-immobilized on polyurethane particles were calculated to be 5 mM and 04.1 × 10(-2) mM/min for K M and V max, respectively. Results demonstrated that the MPS, with tannase activity, can be successfully used for the production of the antioxidant gallic acid from methyl-gallate substrate. Direct use of PMS to produce gallic acid can be advantageous as no previous extraction of enzyme is required, thus reducing production costs.

'Tar bush' and 'creosote bush' were substrates of fungal cultivation for tannase production and gallic acid and pyrocatechol accumulation. Aspergillus niger GH1 grew similarly on both plant materials under solid state culture conditions, reaching maximal levels after 4 d. Fungal strain degraded all tannin content of creosote bush after 4 d of fermentation and >75 % of tar bush after 5 d. Higher level of tannase activity was detected in tar bush fermentation. Biotransformation of tannins to gallic acid was high (93 % in creosote bush and 89 % in tar bush). Pyrocatechol was released poorly. Kinetic parameters of tannin conversion were calculated.

The cellular actions of the thyroid hormones L-thyroxine and L-triiodothyronine are mediated by the association of hormone with a chromatin-associated receptor. In cultured GH1 cells, a hormone-responsive rat pituitary cell line, thyroid hormone decreases the concentration of its receptor at early incubation times by reducing the accumulation of newly synthesized receptor. In this study, we demonstrate that cholera toxin also reduces the amount of nuclear receptor in GH1 cells in a time- and dose-dependent fashion, without altering the affinity of the receptor for hormone. The reduction of receptor mediated by cholera toxin is not secondary to a generalized inhibition of cell protein synthesis or cell replication rates and this effect can be abolished by pretreatment of the cholera toxin with soluble ganglioside II3-alpha-N- acetylneuraminosylgangliotetraosylceramide . This effect requires an intact cholera toxin molecule and does not occur at similar concentrations of the membrane-binding B subunit of cholera toxin. In order to study the influence of cholera toxin on thyroid hormone receptor turnover, we have used a dense amino acid-labeling technique. The results indicate that cholera toxin does not change the half-life of receptor, but decreases the rate of appearance of newly synthesized receptor. This decreased rate completely accounts for the lowered steady state receptor levels. The extent of cAMP stimulation by cholera toxin does not correlate with the extent of receptor reduction and forskolin, which stimulates cAMP 25- to 500-fold, does not decrease thyroid hormone receptor abundance. These studies suggest that cholera toxin modulates receptor levels by a mechanism(s) that is not mediated by cAMP in GH1 cells.

A 66-kDa thermostable family 1 Glycosyl Hydrolase (GH1) enzyme with β-glucosidase and β-galactosidase activities was purified to homogeneity from the seeds of Putranjiva roxburghii belonging to Euphorbiaceae family. N-terminal and partial internal amino acid sequences showed significant resemblance to plant GH1 enzymes. Kinetic studies showed that enzyme hydrolyzed p-nitrophenyl β-D: -glucopyranoside (pNP-Glc) with higher efficiency (K (cat)/K (m) = 2.27 x 10(4) M(-1) s(-1)) as compared to p-nitrophenyl β-D: -galactopyranoside (pNP-Gal; K (cat)/K (m) = 1.15 x 10(4) M(-1) s(-1)). The optimum pH for β-galactosidase activity was 4.8 and 4.4 in citrate phosphate and acetate buffers respectively, while for β-glucosidase it was 4.6 in both buffers. The activation energy was found to be 10.6 kcal/mol in the temperature range 30-65 °C. The enzyme showed maximum activity at 65 °C with half life of ~40 min and first-order rate constant of 0.0172 min(-1). Far-UV CD spectra of enzyme exhibited α, β pattern at room temperature at pH 8.0. This thermostable enzyme with dual specificity and higher catalytic efficiency can be utilized for different commercial applications.

With the perspective of bone marrow purging in autologous transplantation, we investigated the cytotoxicity of the anti-T cell immunotoxin (IT) WT1-ricin A (anti-CD7) to malignant T cells obtained from patients with T cell acute lymphoblastic leukaemia or lymphoma. The cytotoxic efficacy of IT was based on the extent of protein synthesis inhibition. Cytotoxicity of IT to malignant T cells showed a dependency on antigen density comparable to the T cell lines GH1, CEM, Jurkat, HSB-2 and HPB-ALL and was enhanced considerably in the presence of 6 mM ammonium chloride. The ultimate proof of cell kill can only be obtained from clonogenic assays; however, culturing of malignant T cells was not feasible. Therefore these assays were performed with the cell line CEM that expresses comparable amounts of CD7 antigen as malignant T cells of most patients. More than 6-logs of CEM appeared to be eliminated after incubation with 10(-8) M WT1-ricin A. Immunotoxins are only effective after entering the target cell. The pattern of internalization of the IT was determined by means of 125I-WT1. After internalization the CD7 antigen was re-expressed on the cell membrane. This enables a long incubation period resulting in an increased elimination of malignant T cells. Even after 16 h the IT was still accumulated intracellularly. This pattern of continuous uptake of IT was reflected in a gradually increasing cytotoxicity with incubation time. Effective bone marrow purging can be carried out without adverse effects on progenitor cells with 10(-8) M WT1-ricin A. At that concentration the antibody binding capacity was saturated. We showed that the protein synthesis inhibition in malignant T cells by WT1-ricin A is comparable to the inhibition in T cell lines and that high amounts of CEM cells can be killed. These data suggest that cell lines can be used to test the efficacy of IT to malignant T cells. WT1-ricin A appears to be very potent for the purging of autologous bone marrow from patients with T cell malignancies.

Ellagic acid is one of the most bioactive antioxidants with important applications in pharmaceutical, cosmetic, and food industries. However, there are few biotechnological processes developed for its production, because it requires precursors (ellagitannins) and the corresponding biocatalyst (ellagitannase). The aim of this study was to optimize the culture conditions for ellagitannase production by Aspergillus niger in solid-state fermentation (SSF). The bioprocess was carried out into a column bioreactor packed with polyurethane foam impregnated with an ellagitannins solution as carbon source. Four strains of Aspergillus niger (PSH, GH1, HT4, and HC2) were evaluated for ellagitannase production. The study was performed in two experimental steps. A Plackett-Burman design was used to determine the influencing parameters on ellagitannase production. Ellagitannins concentration, KCl, and MgSO4 were determined to be the most significant parameters. Box-Behnken design was used to define the interaction of the selected parameters. The highest enzyme value was obtained by A. niger PSH at concentrations of 7.5 g/L ellagitannins, 3.04 g/L KCl, and 0.76 g/L MgSO4. The methodology followed here allowed increasing the ellagitannase activity 10 times over other researcher results (938.8 U/g ellagitannins). These results are significantly higher than those reported previously and represent an important contribution for the establishment of a new bioprocess for ellagic acid and ellagitannase production.

The bioconversion of lignocellulose in various industrial processes, such as biofuel production, requires the degradation of cellulose. Actinomadura amylolytica YIM 77502^T is an aerobic, Gram-positive actinomycete that can efficiently degrade crystalline cellulose by extracellular cellulases. Genomic analysis of A. amylolytica identified 9 cellulase and 11 β-glucosidase genes that could potentially encode proteins that digest cellulose. Extracellular proteome characterization of A. amylolytica cell-free culture supernatant by liquid chromatography tandem mass spectrometry analysis revealed that 4 of these cellulases and 2 of these β-glucosidases functioned during cellulose hydrolysis. Thin-layer chromatography analysis revealed extracellular β-glucosidases play a major role in carboxyl methyl cellulose (CMC) degradation of products in culture supernatants. In this study, 2 of the identified secreted β-glucosidases, AaBGL1 and AaBGL2, were functionally expressed in Escherichia coli and found to have β-glucosidase activity with wide substrate specificities, including for p-nitrophenyl β-D-glucopyranoside (pNPG), p-nitrophenyl-beta-D-cellobioside (pNPC), and cellobiose. Moreover, AaBGL1 and AaBGL2 had high tolerances for glucose. After adding these β-glucosidases to commercial cellulases, the degradation rates of CMC, Avicel, birch sawdust, and corncob powder increased by 37, 42, 33, and 9%, respectively. Overall, this work identifies an alternative potential source of β-glucosidases with potential applications in commercial cellulose utilization and the bioenergy industry.

The aim of this work was to study the accessibility of histone H1(0) and its structural domains to antibody binding in high molecular mass chromatin fragments of different conformations. Three types of specific antibody populations were used: (1) anti-H1(0) which reacted with antigenic determinants situated along the whole polypeptide chain, (2) anti-GH5 or anti-GH1(0) which recognized epitopes located in the globular region of H1(0) and (3) anti-C-tail antibodies reacting specifically with fragment 99-193 of the protein molecule. The immunoreactivity of the chromatin-bound antigen was investigated by solid-phase ELISA performed on glutaraldehyde-cross-linked chromatin and by an inhibition assay carried out with native chromatin in solution. The results of both methods were unidirectional and showed that: (1) the accessibility of H1(0) did not change with the compaction of the fiber; (2) the G-domain was not accessible to antibodies either in the relaxed or in the condensed state of the fragments, (3) the binding of the C-terminus-specific antibodies was different for isolated monosomes and for the chromatin fiber and (4) the degree of exposure of the epitopes of H1(0) in chromatin was much less than that of histone H1.

BACKGROUND

With the paucity of available literature correlating genetic mutation and response to treatment, we aimed to study the genetic makeup of children with growth hormone (GH) deficiency in Western India and correlate the mutation with auxology and response to GH treatment at end of 1 year.

METHODS

Fifty-three (31 boys and 22 girls) children with severe short stature (height for age z-score <-3) and failed GH stimulation test were studied. Those having concomitant thyroid hormone or cortisol deficiencies were appropriately replaced prior to starting GH treatment. A magnetic resonance imaging (MRI) brain scan was done in all. Genetic mutations were tested for in GH1, GHRH, LHX3, LHX4 and PROP1, POU1F1 and HESX1 genes.

RESULTS

Mean age at presentation was 9.7±5.1 years. Thirty-seven children (Group A) had no genetic mutation detected. Six children (Group B) had mutations in the GH releasing hormone receptor (GHRHR) gene, while eight children (Group C) had mutation in the GH1 gene. In two children, one each had a mutation in PROP1 and LHX3. There was no statistically significant difference in baseline height, weight and BMI for age z-score and height velocity for age z-score (HVZ). HVZ was significantly lower, post 1 year GH treatment in the group with homozygous GH1 deletion than in children with no genetic defect.

CONCLUSIONS

Response to GH at the end of 1 year was poor in children with the homozygous GH1 deletion as compared to those with GHRHR mutation or without a known mutation.

Isolated Growth Hormone Deficiency (IGHD) due to GH1 gene defects has a variable inheritance pattern: autosomal recessive, autosomal dominant, and X-linked. the autosomal dominantly inherited form, IGHD II, is mainly caused by heterozygous mutations of splicing around the exon 3/IVs3 boundary region of the GH1 gene resulting in exon 3 skipping of transcripts. We have previously reported findings on GH1 gene mutations in 28 russian patients with severe congenital IGHD (-3.22+/-1.2 height sDs at the age of 1yr); five heterozygous dominant negative splice site mutations in intron 2, intron 3, and exon 4 of the GH1 gene were identified in 32.1% of the cohort. In the present report we describe a novel 456G>A heterozygous mutation of splicing of the last base of the 3'-acceptor splice site of exon 4 within the GH1 in a 4.2-year old, extremely short (-5.32 height sDs) girl with congenital IGHD. the mutation involves a highly conserved GGGgtg sequence of the exon 4/IVs4 boundary region of the GH1 gene. the predicted effect of the 456 G>A mutation is perturbed splicing with possible skipping of exon 4 of the GH1 gene. the novel heterozygous 456 G>A mutation in exon 4 expands the spectrum of dominant negative splicing defects within the GH1 gene, responsible for congenital IGHD.

β -Glucosidases are used in the food industry to hydrolyse glycosidic bonds in complex sugars, with enzymes sourced from extremophiles better able to tolerate the process conditions. In this work, a novel β-glycosidase from the acidophilic organism Alicyclobacillus herbarius was cloned and heterologously expressed in E. coli BL21 (DE3). Ahe GH1 was stable over a broad range of pH values (5-11) and temperatures (4 ºC-55 ºC). The enzyme exhibited excellent tolerance to fructose and good tolerance to glucose, retaining 65% activity in the presence of 10% ( w/v ) glucose. It also tolerated organic solvents, some of which appeared to have a stimulating effect, in particular ethanol with a 1.7-fold increase in activity at 10% ( v/v ). The enzyme was then applied for the cleavage of isoflavone from isoflavone glucosides in an ethanolic extract of soy flour, to produce soy isoflavones which constitute a valuable food supplement, achieving full conversion within 15 min at 30 °C.

Keywords: Extremophile; HPLC; Hydrolases; Isoflavones; biocatalysis.

Many studies have demonstrated that the most common polymorphism (T1663A, rs2665802) in the promoter region of growth hormone 1 (GH1) gene might play an important role in cancer development and progression. This meta-analysis aims to investigate a more precise estimation of the relationship between GH1 T1663A polymorphism and cancer risk. We searched CISCOM, CINAHL, Web of Science, PubMed, Google Scholar, EBSCO, Cochrane Library, and CBM databases from inception through October 1st, 2013. Meta-analysis was performed using the STATA 12.0 software. Seven studies were included with a total of 4,018 cancer patients and 5,308 healthy controls. Our meta-analysis results revealed that GH1 T1663A polymorphism was associated with increased cancer risks. Subgroup analysis by cancer type showed significant associations between GH1 T1663A polymorphism and increased colorectal cancer risk, but there was no evidence of any association with breast cancer. Further subgroup analysis based on ethnicity indicated that GH1 T1663A polymorphism might increase cancer risks among Asian populations. However, no statistically significant association was found among Caucasian populations. Meta-regression analyses also suggested that cancer type and ethnicity may be the main sources of heterogeneity. No publication bias was detected in this meta-analysis. The present meta-analysis indicates that GH1 T1663A polymorphism may contribute to the risk of colorectal cancer, especially among Asian populations.

Certain peptide hormones and/or their cognate receptors influencing normal cellular pathways also have been detected in breast cancers. The hypothesis is that gene subsets of these regulatory molecules predict risk of breast carcinoma recurrence in patients with primary disease.

Gene expression levels of 61 hormones and 81 receptors were determined by microarray with LCM-procured carcinoma cells of 247 de-identified biopsies. Univariable and multivariable Cox regressions were determined using expression levels of each hormone/receptor gene, individually or as a pair.

Molecular signatures for ER+/PR+, ER-/PR-, and ER- carcinoma cells deciphered by LASSO were externally validated at HRs (CI) of 2.8 (1.84-4.4), 1.53 (1.01-2.3), and 1.72 (1.15-2.56), respectively. Using LCM-procured breast carcinoma cells, a 16-gene molecular signature was derived for ER+/PR+ biopsies, whereas a 10-gene signature was deciphered for ER-/PR- cancers. Four genes, POMC, CALCR, AVPR1A, and GH1, of this 10-gene signature were identified in a 6-gene molecular signature for ER- specimens.

Applying these signatures, Kaplan-Meier plots definitively identified a cohort of patients with either ER-/PR- or ER- carcinomas that exhibited low risk of recurrence. In contrast, the ER+/PR+ signature identified a cohort of patients with high risk of breast cancer recurrence. Each of the three molecular signatures predicted clinical outcomes of breast cancer patients with greater accuracy than observed with either single-gene analysis or by ER/PR protein content alone. Collectively, our results suggest that gene expression profiles of breast carcinomas with suspected poor prognosis (ER-/PR-) have identified a subset of patients with decreased risk of recurrence.

The thyroid hormone nuclear receptor is a chromatin-associated protein regulating expression of specific genes. Acetylation of nucleosomal core histones is thought to be one of the factors regulating transcriptional activity of chromatin, and it is suggested that this reaction negatively regulates thyroid hormone receptor levels in GH1 cells (Samuels, H.H., Stanley, F., Casanova, J., and Shao, T. C. J. Biol. Chem. 255, 2499-2508). In the present study, we found that n-butyrate, a potent inhibitor of histone deacetylase, increases thyroid hormone receptor levels in three distinct non-pituitary cells without changing binding affinity. This effect appeared within 30 min and reached a plateau (240% of control) after a 6-h treatment, before important cellular functions were affected. This effect was time-dependent, dose-dependent, reversible, and paralleled the changes in the electrophoretic mobilities of histones H3 and H4. n-Butyrate prolonged the receptor half-life, and this prolongation corresponded to the increase of receptor levels. Thyroid hormone did not reduce its own receptor levels or influence the effect of n-butyrate. Considering the difference between GH1 cells and non-pituitary cells in the regulation of thyroid hormone receptor levels, our observations, together with those of Samuels et al., suggest the possibility that the acetylation of chromatin-associated proteins has a physiological significance in the regulation of thyroid hormone nuclear receptor levels.

We present evidence that T3 can alter the ADP-ribosylation of chromatin associated proteins. Nuclei from GH1 cells were incubated with [adenylate-32P]NAD and the radioactivity incorporated into histone and non-histone proteins was quantitated and analyzed by gel electrophoresis and autoradiography. Incubation of GH1 cells for 24 h with T3 lowered by 40-70% the [32P]ADP-ribose incorporated into nuclear proteins. However, incubation for 3 h with T3 resulted in a stimulation instead of a decrease of in vitro [32P]ADP-ribose incorporation. The major ADP-ribosylated component electrophoresed as a 120,000 molecular mass non-histone protein, and radiolabeled histones were also observed. The same protein species were observed for all the experimental groups and T3 affected the extent of ADP-ribosylation but did not alter the sedimentation of the [32P]ADP-ribosylated components excised from chromatin after micrococcal nuclease digestion.

A novel bacterial strain, designated as CF21(T), was isolated from the air of Ailuropoda melanoleuca enclosures in China. Cells were gram-negative, aerobic, non-motile, and rod shaped. Strain CF21(T) grew at 10-40 °C (optimum 28-30 °C) and pH 6.0-9.0 (optimum pH 7.0-8.0) and in the presence of NaCl concentrations ranging from 0.0% (w/v) to 2.0 % (optimum 0.0-1.0%). 16SrRNA gene sequence analysis indicated that strain CF21(T) belonged to genus Lysobacter within class Gammaproteobacteria and was most closely related to Luteimonas dalianensi OB44-3(T) (95.8% similarity), Lysobacter ruishenii CTN-1(T) (95.1%), Lysobacter spongiicola KMM329(T) (94.8 %), and Lysobacter daejeonensis GH1-9T (94.6%). The genomic G+C DNA content was 68.72 mol%. Major cellular fatty acids of CF21(T) were iso-C16:0 (30.22%), iso-C15:0 (25.70%), and the sum of 10-methyl C16 : 0 and/or iso-C17 : 1ω9c (21.94%). The prominent isoprenoid quinone was ubiquinone 8 (Q-8). Primary polar lipids included diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, and an unknown phospholipid. DNA sequence relatedness between strain CF21(T) and L. ruishenii CTN-1(T) was 56%, which was clearly below the 70% threshold for prokaryotic species delineation. These analyses indicated that CF21(T) is a novel member of genus Lysobacter, for which the name Lysobacter chengduensis sp. nov. is proposed. The type strain is CF21(T) (=CGMCC1.15145(T) = DSM 100306(T)).

We have previously shown that 3,5,3'-triiodo-L-thyronine (L-T3) stimulates cell growth and a 4- to 8-fold increase in growth hormone mRNA in GH1 cells. These effects appear to be mediated by a thyroid hormone nuclear receptor with an equilibrium dissociation constant for L-T3 of 0.2 nM and an abundance of about 10,000 receptors per cell nucleus. In this report, we show that L-T3 exerts a pleiotypic effect on GH1 cells to rapidly (within 2 h) stimulate [3H]uridine uptake to a maximal value of 2.5- to 3-fold after 24 h. This results from an increase in the number of functional uridine "transport sites" as shown by studies documenting an increase in the apparent Vmax with no change in the Km, 17 microM. Although the labeling of the cellular uridine pool and pools of all phosphorylated uridine derivatives was increased by L-T3, there was no change in the relative amounts of the individual pools in cells incubated with or without hormone. The intracellular concentration of [3H]uridine was estimated to be similar to that of the medium, suggesting that facilitated transport mediates [3H]uridine uptake. That this increase in [3H]uridine transport was nuclear receptor-mediated is supported by the excellent correspondence of the L-T3 dose-response curve for [3H]uridine uptake and that for L-T3 binding to receptor. Finally, inhibition of protein synthesis by cycloheximide and RNA synthesis by actinomycin D demonstrated that the L-T3 effect required continuing protein and RNA synthesis. These results are consistent with an effect of the L-T3-nuclear receptor complex to increase uridine uptake in GH1 cells by altering the expression of gene(s) essential for the transport process.