Altered Rates Of Transcription Research Articles

Maturation of UTR-Derived sRNAs Is Modulated during Adaptation to Different Growth Conditions.

Small regulatory RNAs play a major role in bacterial gene regulation by binding their target mRNAs, which mostly influences the stability or translation of the target. Expression levels of sRNAs are often regulated by their own promoters, but recent reports have highlighted the presence and importance of sRNAs that are derived from mRNA 3′ untranslated regions (UTRs). In this study, we investigated the maturation of 5′ and 3′ UTR-derived sRNAs on a global scale in the facultative phototrophic alphaproteobacterium Rhodobacter sphaeroides. Including some already known UTR-derived sRNAs like UpsM or CcsR1-4, 14 sRNAs are predicted to be located in 5 UTRs and 16 in 3′ UTRs. The involvement of different ribonucleases during maturation was predicted by a differential RNA 5′/3′ end analysis based on RNA next generation sequencing (NGS) data from the respective deletion strains. The results were validated in vivo and underline the importance of polynucleotide phosphorylase (PNPase) and ribonuclease E (RNase E) during processing and maturation. The abundances of some UTR-derived sRNAs changed when cultures were exposed to external stress conditions, such as oxidative stress and also during different growth phases. Promoter fusions revealed that this effect cannot be solely attributed to an altered transcription rate. Moreover, the RNase E dependent cleavage of several UTR-derived sRNAs varied significantly during the early stationary phase and under iron depletion conditions. We conclude that an alteration of ribonucleolytic processing influences the levels of UTR-derived sRNAs, and may thus indirectly affect their mRNA targets.

Unfolded Protein Response Differentially Regulates TLR4-Induced Cytokine Expression in Distinct Macrophage Populations.

Cellular stress responses are often engaged at sites of inflammation and can alter macrophage cytokine production. We now report that macrophages in distinct states of differentiation or in different temporal stages of inflammatory response exhibit differential sensitivity to cell stress mediated alterations in M1-like polarized inflammatory cytokine production. Tunicamycin (Tm) treatment of bone marrow derived macrophages (BMDM) cultured with M-CSF cultured bone marrow derived macrophages (M-BMDM) had markedly amplified M1-like responses to LPS, exhibiting higher levels of IL12p40 and IL12p35 mRNAs while BMDM cultured with GM-CSF, which normally express high IL12 subunit production in response to LPS, were relatively unaltered. Anti-inflammatory IL10 mRNA production in LPS-stimulated M-BMDM was greatly reduced by cell stress. These changes in cytokine mRNA levels resulted from altered rates of transcription and mRNA decay. Stress also altered cytokine protein production. Resident liver macrophages isolated from mice treated with Tm showed elevated levels of IL12 subunit mRNA production following LPS stimulation. Furthermore, macrophages infiltrating the liver during the early phase of acetaminophen injury (24 h) had little stress-mediated change in cytokine mRNA production while cells isolated in the later phase (48–72 h) exhibited higher sensitivity for stress elevated cytokine production. Hence cultured macrophages developed using different growth/differentiation factors and macrophages from different temporal stages of injury in vivo show markedly different sensitivity to cell stress for altered inflammatory cytokine production. These findings suggest that cellular stress can be an important modulator of the magnitude and character of myeloid inflammatory activity.

MRNA levels are buffered upon knockdown of RNA decay and translation factors via adjustment of transcription rates in human HepG2 cells

ABSTRACTEvidence from yeast and mammals argues the existence of cross-talk between transcription and mRNA decay. Stabilization of transcripts upon depletion of mRNA decay factors generally leads to no changes in mRNA abundance, attributing this to decreased transcription rates. We show that knockdown of human XRN1, CNOT6 and ETF1 genes in HepG2 cells led to significant alteration in stability of specific mRNAs, alterations in half-life were inversely associated with transcription rates, mostly not resulting in changes in abundance. We demonstrate the existence of the gene expression buffering mechanism in human cells that responds to both transcript stabilization and destabilization to maintain mRNA abundance via altered transcription rates and may involve translation. We propose that this buffering may hold novel cancer therapeutic targets.

Leaf senescence and gene expression.

Abstract The long-standing view of leaf senescence as an organized genetically controlled process has received convincing support from recent genome-wide studies of differential gene expression. Many hundreds of senescence-associated genes (SAGs) appear to be up- or down-regulated for the sake of nutrient salvage, including a wide array of SAGs that encode proteases, nucleases, lipid-, carbohydrate- and amino acid-degrading enzymes. Mobilization and export of nutrients from senescing leaves is accompanied by complex expression of a special set of genes for transporters. Another category of genes with altered transcription rates was shown to control protective- or stress-response functions necessary to keep the cells viable till the task of salvage is completed. Coordinated activation or inactivation of various subsets of SAGs is governed by a comprehensive network of regulatory genes, such as genes for transcription factors and hormone pathways. Transcriptome data and mutant screens provide a foundation for further analysis of physiological functions of senescence-expressed genes and subsequent manipulation for the benefits of plant breeding.

Cancer treatment of the future: Inhibitors of histone methyltransferases

Cancer in humans is the result of a multi-step process. This process often involves the activation of oncogenes and/or the inactivation of tumor suppressor genes. These two steps arise not only due to mutations, but can also be the result of a translocation or an altered transcription rate. One important mechanism is the occurrence of epigenetic alterations like promotor methylation (which may lead to tumor suppressor silencing) or decreased histone acetylation (which can result in the downregulation of proteins involved in apoptosis). Today, histone acetylation and DNA methylation are epigenetic modifications which have been linked closely to the pathology of human cancers and inhibitors of both enzyme classes for clinical use are at hand. In contrast, other fields of epigenetics still lack of similarly thorough knowledge. This is especially true for the group of histone methyltransferases and their inhibitors. Since connections between histone methylation patterns and cancer progression have been recognized, histone methyltransferases represent promising targets for future cancer treatment.

Master Energy-Regulator Kinases in Plants

Plants undergo dramatic changes in metabolism as environmental conditions change and stressful situations are encountered. Most obviously, anabolic processes that operate during the day, when photosynthesis is active, switch to catabolic ones at night, or when sugars are unavailable, or under stresses like exposure to herbicides or to excessive amounts of water. Baena-González et al . identify a pair of protein kinases that have a central role in orchestrating common cellular transcriptional responses to such diverse stimuli. Through analysis of public databases, the authors identified dark-induced ( DIN ) genes whose expression increased in plant cells exposed to dark but decreased after exposure to light or sugars. Experiments monitoring expression of a reporter gene designed from one of the DIN genes revealed sensitivity of the response to a protein kinase inhibitor. Taking cues from work on yeast and mammalian kinases that control metabolism, the authors focused on members of the Snf1-related kinase gene families, which are related to the yeast protein Snf1. Two such proteins, KIN10 and KIN11, activated expression of the DIN reporter when expressed in Arabidopsis leaves. Systematic mutagenesis of regulatory elements implicated a role for G-box binding factors in controlling the response of DIN genes to stress or to KIN10 or KIN11. The authors explored the global transcriptional response to expression of KIN10 with whole-genome GeneChips and identified more than 1000 genes with altered rates of transcription. Comparison to published gene expression patterns showed strong correlation of a subset of these genes with those regulated in plants deprived of sugar or energy. There were negative correlations with expression profiles from plants treated with sugars. A gene expression response highly correlated to that induced by KIN10 was also detected in protoplasts stressed by hypoxia. KIN10 activated (and sugars repressed) genes functioning in major catabolic pathways, whereas genes that promote energy consumption and anabolic processes were repressed in response to KIN10. The authors propose that Kin10 and Kin11 can thus be considered global regulators of primary and secondary metabolism and protein synthesis. The authors went on to generate whole Arabadopsis plants overexpressing KIN10 or depleted of the KIN10 and KIN11 proteins by RNAi. Expression of KIN10 enhanced survival of plants kept in sugar-free medium in the presence of low amounts of light. Overexpression of KIN10 slowed development and--reminiscent of caloric restriction in animals--delayed the onset of senescence. Depletion of KIN10 and KIN11 blocked the transcriptional changes normally associated with transition to dark or stressful conditions and suggested an additional role of the kinases in control of development and flowering. The authors propose that KIN10 and KIN11 may be, like AMPK (adenosine monophosphate-activated protein kinase) of mammals, central regulators of signaling that control energy homeostasis. E. Baena-González, F. Rolland, J. M. Thevelein, J. Sheen, A central integrator of transcription networks in plant stress and energy signaling. Nature 448 , 938-942 (2007). [PubMed]

Insulin and growth hormone stimulate somatostatin receptor (SSTR) expression by inducing transcription of SSTR mRNAs and by upregulating cell surface SSTRs

This study examined the effects of insulin (INS) and growth hormone (GH) on mRNA and functional expression of somatostatin receptors (SSTRs). Rainbow trout liver was used as a model system to evaluate the direct effects of INS and GH on mRNA expression of three SSTR subtypes characterized previously from this species: SSTR1A, SSTR1B, and SSTR2. INS and GH directly stimulated steady-state levels of all SSTR mRNAs in a concentration- and time-dependent manner; however, the pattern of expression was hormone and SSTR subtype specific. INS stimulated SSTR2 expression to a greater extent than SSTR1A or SSTR1B expression, whereas GH stimulated SSTR2 and SSTR1B expression to a similar extent, with SSTR2 and SSTR1B expression being more responsive to GH than SSTR1A. Whether INS- or GH-stimulated SSTR expression resulted from altered rates of transcription and/or changes in mRNA stability also was investigated. Formation of nascent SSTR transcripts in nuclei isolated from rainbow trout hepatocytes was significantly stimulated by INS and GH. Neither INS nor GH, however, affected the stability of SSTR mRNAs. Functional expression of SSTRs was studied in Chinese hamster ovary (CHO-K1) cells stably transfected with SSTR1A or SSTR1B. Surface expression of functional SSTRs was stimulated by INS and GH. These findings indicate that INS and GH stimulate SSTR expression by regulating transcription of SSTR mRNAs and by increasing functional SSTRs on the cell surface, and they suggest that regulation of SSTRs may be important for the coordination of growth, development, and metabolism of vertebrates.

Myosin Loss in Denervated Rat Soleus Muscle after Dexamethasone Treatment

Objective: Critical illness myopathy (CIM) is an acute myopathy that appears in the setting of critical illness or during exposure to corticosteroids and neuromuscular blocking agents. Its pathological feature is selective loss of thick myosin filaments. Our aim is to gain further insight into the pathomechanism of myosin loss in this myopathy. Methods: To clarify the expression of myosin heavy chain (MHC) and ubiquitin ligase atrogin-1 in this myopathy, histological, immunohistochemical, SDS-PAGE, and semiquantitative reverse transcriptase-polymerase chain reaction studies were performed on innervated and denervated rat soleus muscles after saline and dexamethasone treatments. Results: Denervated muscles from dexamethasone-treated rats showed marked MHC loss. The mRNA expression of ubiquitin ligase atrogin-1 was significantly increased in denervated dexamethasone-treated muscles, suggesting that the ubiquitin-proteasome pathway plays an important role in muscular wasting in CIM. Furthermore, mRNA levels of MHC I, a myosin isoform, were decreased in the denervated dexamethasone-treated muscles. Conclusion: Our findings suggest that an altered transcription rate of myosin, as well as the upregulation of multiple ubiquitin ligases, may be responsible for selective myosin loss in this myopathy.

Glucose-stimulated Somatostatin Gene Expression in the Brockmann Bodies of Rainbow Trout (Oncorhynchus mykiss) Results from Increased mRNA Transcription and not from Altered mRNA Stability

Previously, we showed that glucose increases the steady-state levels of the mRNAs encoding two distinct preprosomatostatins (each containing [Tyr7, Gly10]-somatostatin-14 at their C-termini; denoted PPSS II’ and PPSS II”) in the endocrine pancreas (Brockmann body) of rainbow trout (Oncorhynchus mykiss). In the present study, isolated islet cells were used to determine whether glucose-stimulated expression resulted from altered rates of transcription and/or from changes in RNA stability. Nuclear runon assays indicated that the number of PPSS II nascent transcripts were significantly higher in nuclei isolated from islet cells cultured in 10 mM glucose compared to those isolated from cells incubated in 4 mM glucose. High glucose (10 mM) did not, however, affect the stability of PPSS II mRNAs. These results indicate that glucose-stimulated somatostatin expression in the Brockmann bodies of rainbow trout results from increased endogenous mRNA transcription and not from altered mRNA stability.

Transcription of androgen receptor and 5α-reductase II in genital fibroblasts from patients with androgen insensitivity syndrome

Impaired virilisation during embryonic development and pubertal arrest in patients with androgen insensitivity syndromes (AIS) is usually caused by mutations in the androgen receptor (AR)- or the 5α-reductase II (5RII) gene. However, identical mutations may lead to strikingly different phenotypes. To investigate whether this may be caused by individually altered transcription rates in fibroblasts from the genital region (GF) from affected patients, we applied competitive reverse transcribed PCRs (competitive RT-PCR) targeting AR- and 5RII-transcripts. We could demonstrate that AR- and 5RII-mRNA concentrations in cells from patients with partial and complete AIS and missense mutations in the AR- or 5RII-gene are normal or only moderately lowered compared to equally aged normal controls. However, in a patient bearing a premature stop-codon in the AR-gene a considerably lowered AR-transcript level was detected. We conclude, that in patients with incomplete virilisation disorders due to missense mutations, transcription regulation of AR and 5RII generally follows normal patterns. Accordingly, the premature stop-codon found in one patient's AR-gene may rather cause reduced transcript stability than an impairment of transcription activity. Therefore, altered AR- and 5RII-transcription rates in fibroblasts from the GF do not seem to be the cause for the variable genotype-phenotype correlation in androgen insensitivity syndrome.

Acute quadriplegia and loss of muscle myosin in patients treated with nondepolarizing neuromuscular blocking agents and corticosteroids: mechanisms at the cellular and molecular levels.

Long-term treatment with nondepolarizing neuromuscular blocking agents and corticosteroids in the intensive care unit is not benign, and an increasing number of patients with acute quadriplegic myopathy have been reported with increased use of these drugs. The purpose of this study was to investigate the mechanisms underlying acute quadriplegic myopathy. Percutaneous muscle biopsy samples were obtained, and electrophysiologic examinations were performed during the acute phase and during recovery in patients with acute quadriplegic myopathy. Regulation of muscle contraction and myofibrillar protein synthesis was studied using cell physiologic techniques, ultrasensitive electrophoresis, in situ hybridization, and histopathologic techniques. All patients were seen in the intensive care unit of different university hospitals. All patients were critically ill with sepsis. They had been given massive doses of corticosteroids in combination with variable doses of neuromuscular blocking agents. All patients developed paralysis of spinal nerve-innervated muscles. On the other hand, cranial nerve-innervated muscle and sensory and cognitive functions were well maintained after discontinuation of treatment with neuromuscular blocking agents. Muscle biopsy samples were obtained and electrophysiologic examinations were performed in all patients. The major observations in patients with acute quadriplegic myopathy were, as follows: a) a general decrease in myofibrillar protein content; b) specific but highly variable partial or complete loss of myosin and myosin-associated proteins; c) very low thick-filament/thin-filament protein ratios; d) absence of myosin messenger RNA; and e) a dramatically impaired muscle cell force-generating capacity in the acute phase of acute quadriplegic myopathy. During clinical improvement, normal expression of myosin messenger RNAs, reexpression of thick-filament proteins, and increased specific tension were observed. Acute quadriplegic myopathy is associated with a specific decrease in thick-filament proteins related to an altered transcription rate. Although the decreased content of thick-filament proteins is important for prolonged muscle weakness, it is not the primary cause of muscle paralysis in the acute stage, during which impaired muscle membrane excitability probably plays a more significant role. Several factors contribute to this condition, but the action of corticosteroids seems to be the predominant one, along with potentiation by neuromuscular blocking agents, immobilization, and probably also concurrent sepsis.

Differential display approach to quantitation of environmental stimuli on bacterial gene expression.

The differential display of the mRNA technique for eukaryotes is fruitful in identifying genes with altered transcription rates caused by exogenous or endogenous stimuli. Prokaryotic analogues of the method using arbitrary oligonucleotides may reach a complete statistical genome coverage. Thus a genome-wide mass screening for transcriptionally regulated sequences will be possible. However, the primer sets have to be optimized for a given species to result in maximum band yields. Hence the construction of primers requires the calculation of oligonucleotide frequency distributions from known coding regions to choose sequences with frequent occurrence in the bacterial genome. After completion of many whole genome sequencing projects, differentially regulated cDNA sequences are readily identified by sequence comparisons.

Altered transcription control is responsible for the increased level of proliferation-associated P120 in rapidly growing breast carcinoma.

Transcriptional and post-transcriptional regulation of proliferation-associated nucleolar P120 protein expression was examined in 1 normal and 5 malignant breast cell lines. The 6 breast cell lines could be placed into 3 categories on the basis of in vitro growth rate. BT549 and HBL100 grew rapidly; MCF-7/6, MCF-7/AZ and Hs578T grew at a moderate rate; and Hs578N normal epithelia grew slowly. There was a significant correlation between the growth rate, measured by percentage of S-phase fraction of cells or doubling time of cell numbers and the steady-state levels of either P120 protein or P120 mRNA. Next, the mechanisms responsible for the increased level of P120 expression, which is associated with rapidly growing breast carcinoma, were examined. The stability of P120 mRNA was measured by densitometry of quantitative Northern blots of mRNAs from cells treated with actinomycin D. Before the expected decay, a sharp increase in P120 mRNA level was detected shortly after inhibiting the overall RNA or protein synthesis. The calculated half-life of P120 mRNA was very similar (1.8 +/- 0.2 hr) in all 6 cell lines examined. The transcription rate of the P120 gene was determined by densitometric analysis of quantitative nuclear run-off assays. A significant positive correlation was found between the transcription rate of the P120 gene and the steady-state levels of either P120 protein or P120 mRNA. Our conclusion is that the expression of P120 protein is transcriptionally regulated in these breast cells. Therefore, the characteristically high level of P120 protein and mRNA found in most breast carcinomas is due to the altered transcription rate of the P120 gene in transformed cells.

Inactivation of gene expression in plants as a consequence of specific sequence duplication.

Numerous examples now exist in plants where the insertion of multiple copies of a transgene leads to loss of expression of some or all copies of the transgene. Where the transgene contains sequences homologous to an endogenous gene, expression of both transgene and endogenous gene is sometimes found to be impaired. Several examples of these phenomena displaying different features are reviewed. Possible explanations for the observed phenomena are outlined, drawing on known cellular processes in Drosophila, fungi, and mammals as well as plants. It is hypothesized that duplicated sequences can, under certain circumstances, become involved in cycles of hybrid chromatin formation or other processes that generate the potential for modification of inherited chromatin structure and cytosine methylation patterns. These epigenetic changes could lead to altered transcription rates or altered efficiencies of mRNA maturation and export from the nucleus. Where the loss of gene expression is posttranscriptional, antisense RNA could be formed on accumulated, inefficiently processed RNAs by an RNA-dependent RNA polymerase or from a chromosomal promoter and cause the observed loss of homologous mRNAs and possibly the modification of homologous genes. It is suggested that the mechanisms evolved to help silence the many copies of transposable elements in plants. Multicopy genes that are part of the normal gene catalog of a plant species must have evolved to avoid these silencing mechanisms or their consequences.

Altered gene expression in human leukemia K562 cells selected for resistance to etoposide

Sublines of K562 human leukemia cells were selected for resistance (30- to 80-fold) to etoposide by continuous exposure to 0.5 μM VP-16. Two etoposide-resistant cell lines, K/VP.5 and K/VP.5-1, showed a 5-fold reduction in levels of topoisomerase IIα protein compared with K562 cells. Northern analysis indicated a 2.5-fold reduction in topoisomerase IIα mRNA in etoposide-resistant cell lines, due in part to a 1.7-fold decrease in topoisomerase II mRNA stability with no change in transcription rate. Immunoblotting assays of electrophoresed cell lysates from VP-16-treated cells revealed less drug-induced covalent topoisomerase II/DNA adducts in resistant than in sensitive cells, suggesting a functional alteration in resistant cell topoisomerase II. Recent reports of specific topoisomerase II DNA binding sites near the promoter sites of growth response genes and alterations of gene expression in cells treated with topoisomerase II inhibitory drugs led to experiments to determine if the apparent functional alterations of topoisomerase II were accompanied by changes in the regulation of these genes. Therefore, the expression of several growth response genes was compared by northern analysis in parental K562 and both VP-16-resistant cell lines. Basal levels of c- myc were comparable for all three cell lines, but levels of c- jun and c- fos were elevated 2- to 4-fold in VP-16- resistant cell lines. Increased levels of c- fos and c- jun were not a result of altered rates of transcription, as determined by nuclear run-off assays. Exposure of both sensitive and resistant cells to 200 μM VP- 16 for 5 hr resulted in no further changes in topoisomerase II mRNA levels but caused an additional 2- to 3-fold elevation in the level of c- jun mRNA, indicating that altered basal levels of this gene were not due to deregulation of this gene. Acquired VP-16 resistance in K/VP.5 and K/VP.5-1 cells was accompanied by reduced levels and altered activities of DNA topoisomerase II as well as changes affecting the expression of genes important for growth and differentiation.

Anoxia-hyperoxia induces monocyte-derived interleukin-8.

Ischemia-reperfusion and hyperoxia-induced pulmonary injury are associated with the presence of activated neutrophils (PMN) and cellular injury. Although the signals orchestrating the directed migration of these PMN during the pathogenesis of these disease states remain to be fully elucidated, it appears they may be dependent upon the production of certain neutrophil activating/chemotactic factors such as C5a, leukotriene B4, platelet-activating factor, and IL-8. The production of the latter chemotaxin by mononuclear phagocytes is especially intriguing as these cells can mediate inflammatory cell migration by either directly generating IL-8, or by inducing its production from surrounding nonimmune cells. In light of these observations, we propose that ischemia-reperfusion and oxidant stress, in vivo, may be simulated by anoxia-hyperoxia induced stress in vitro, and that this stress may act as a stimulus for the production of IL-8. We now show that isolated human blood monocytes respond to such an oxygen stress with augmented production of IL-8. In initial studies, monocytes demonstrated an increase in the production of IL-8 under anoxic preconditioning. Subsequently, monocytes were cultured under one of the following conditions for 24 h: (a) room air/5% CO2; (b) 95% N2/5% CO2 for 6 h, followed by room air/5% CO2 for 18 h; (c) 95% N2/5% CO2 for 6 h, followed by 95% O2/5% CO2 for 18 h; (d) room air/5% CO2 for 6 h, followed by 95% O2/5% CO2 for 18 h; or (e) 95% O2/5% CO2. Supernatants were isolated and analyzed for IL-8 antigen by specific IL-8 ELISA, demonstrating the production of monocyte-derived IL-8: 5.9 +/- 0.9, 11.4 +/- 1.7, 21.1 +/- 2.3, 14.6 +/- 2.4, and 26.3 +/- 4.7, ng/ml by designated conditions a, b, c, d, and e listed above, respectively. This variance in IL-8 production reflects altered rates of transcription as shown by Northern blot analysis and nuclear run-off assay. Furthermore, when monocytes were concomitantly treated with LPS (100 ng/ml) under in vitro hyperoxic conditions, both IL-8 steady-state mRNA and antigenic activity were two- to threefold greater than under room air conditions. The association of anoxic preconditioning and oxygen stress with augmented production of monocyte-derived IL-8 support the potential role for ischemia-reperfusion and hyperoxia-induced IL-8 production in vivo, providing a possible mechanism for PMN migration/activation in disease states characterized by altered tissue oxygenation.

Rapid transcriptional regulation of the Cab and pEA207 gene families in peas by blue light in the absence of cytoplasmic protein synthesis

We have analyzed the fluence-response and time-course characteristics, and the requirements for protein synthesis, for the blue-light (BL) regulated transcription of the nuclear-coded Cab, pEA207, and pEA25 gene families in pea (Pisum sativum L.). Fluence-response curves indicate two BL responses: a blue low-fluence (BLF) response with a threshold at or below 10(-1) μmol·m(-2) of BL and a blue high-fluence (BHF) response with a threshold between 10(1) and 10(3) μmol·m(-2) of BL. Excitation of the photomorphogenic system responsible for the BLF response results in increased Cab, and decreased pEA25, transcription. Excitation of the photomorphogenic system responsible for the BHF response results in decreased pEA207 transcription and induction of turnover for Cab RNA and pEA207 RNA. Altered rates of transcription for the Cab and pEA207 gene families are apparent within 15 min of BL treatment and remain in effect for at least 24 h. The effect of BL on pEA25 transcription is not apparent until 3-5 h after the BL treatment. Cycloheximide, an inhibitor of cytoplasmic protein synthesis, has no effect on the altered rates of pEA207 and Cab transcription. We conclude from these results that the BLF response for Cab transcription and the BHF response for pEA207 transcription probably occurs without the expression of intervening regulatory genes coded within the nucleus or the translation of pre-existing transcripts derived from nuclear-coded genes.

Insulin-mediated post-transcriptional regulation of hepatic malic enzyme and albumin mRNAs

Livers of insulin-treated diabetic rats accumulate albumin and malic enzyme mRNAs at very different rates. We now report that in normal rats insulin directs a specific increase in malic enzyme mRNA, while albumin mRNA levels remain unaltered. These studies support the contention that insulin regulates the accumulation of hepatic mRNAs in a highly specific manner. To evaluate whether or not albumin and malic enzyme mRNA levels are determined by altered rates of transcription, in vitro transcription assays were performed. The results of these studies demonstrate that increased malic enzyme mRNA levels in insulin-treated normal rats and increased malic enzyme and albumin mRNA levels in insulin-treated diabetic rats do not involve altered rates of transcription of the genetic sequences encoding these proteins. For these two specific proteins, insulin mediates changes in mRNA levels by a post-transcriptional mechanism.

Effects of deletions in the spacer region of the rrnB operon on the transcription of the large ribosomal RNAs from Escherichia coli.

A series of deletions was constructed within the spacer region of the genes for the 16S and 23S RNA on plasmids bearing the rrnB operon. The accumulation and synthesis rates for the 16S and 23S RNAs were determined from normal growing cells and maxicells after transformation with the mutated plasmids. A marked difference in the transcription efficiency of the plasmid-encoded ribosomal 16S and 23S RNAs was observed with cells carrying plasmids, where a sequence motif analogous to the antitermination recognition sequence (Box A) had been deleted. The overall synthesis rate of ribosomal RNAs of such cells was not altered, however, indicating that the difference in transcription rates from the plasmid genes is compensated by altered transcription rates of the corresponding chromosomal genes. In addition, the accumulation of various tRNA species encoded on rRNA operons and non rRNA operons was quantitated and compared. From these results we infer that the regulation of ribosomal RNA transcription does not only occur at the promoter sites but sequence regions possibly involved in antitermination within the operon are crucial for a coordinated synthesis of all ribosomal RNAs.