scholarly journals Enhancers regulate polyadenylation site cleavage and control 3’UTR isoform expression

2020 ◽  
Author(s):  
Buki Kwon ◽  
Neil D. Patel ◽  
Shih-Han Lee ◽  
Jihye Lee ◽  
Weirui Ma ◽  
...  
Keyword(s):  
Amino Acid Sequences ◽  
Polyadenylation Site ◽  
Protein Activity ◽  
Cleavage Activity ◽  
Lipid Phosphatase ◽  
Gene Level ◽  
Dna Elements ◽  
Isoform Expression ◽  
And Control ◽  
Increase Gene Expression

SummaryEnhancers are DNA elements that increase gene expression. mRNA production is determined by transcript production and polyadenylation site (PAS) cleavage activity. We established an assay to measure enhancer-dependent PAS cleavage activity in human cells because PAS cleavage may control alternative 3’UTR isoform expression. We found that enhancers are widespread regulators of PAS cleavage and consistently increase cleavage of proximal and weak PAS. Half of tested transcription factors exclusively regulated PAS cleavage without affecting transcript production, whereas co-activators changed both parameters. Deletion of an endogenous enhancer of PTEN did not change gene-level mRNA or protein abundance but affected expression of alternative mRNA transcripts, thus preventing 3’UTR shortening. Our data reveal that in addition to controlling transcript production, enhancers also regulate PAS cleavage, thus changing 3’UTR isoform usage and protein activity, as PTEN proteins translated from the alternative 3’UTR isoforms differ in intrinsic lipid phosphatase activity despite having identical amino acid sequences.

scholarly journals Interaction of human CYP17 (P-45017α, 17α-hydroxylase-17,20-lyase) with cytochrome b5: importance of the orientation of the hydrophobic domain of cytochrome b5

1997 ◽  
Vol 321 (3) ◽  
pp. 857-864 ◽  
Author(s):  
Peter LEE-ROBICHAUD ◽  
Mustak A. KADERBHAI ◽  
Naheed KADERBHAI ◽  
J. Neville WRIGHT ◽  
Muhammad AKHTAR
Keyword(s):  
Signal Sequence ◽  
Cytochrome B5 ◽  
Amino Acid Sequences ◽  
Side Chain ◽  
Hydrophobic Domain ◽  
Side Chain Cleavage ◽  
The Core ◽  
Cleavage Activity ◽  
P450 Reductase ◽  
Chain Cleavage

Human CYP17 (P-45017α, 17α-hydroxylase-17,20-lyase)-catalysed side-chain cleavage of 17α-hydroxyprogestogens into androgens is greatly dependent on the presence of cytochrome b5. The native form of cytochrome b5 is composed of a globular core, residues 1Ő98, followed by a membrane insertable C-terminal tail, residues 99Ő133. In the present study the abilities of five different forms of cytochrome b5 to support the side-chain cleavage activity of CYP17 were compared. The five derivatives were: the native pig cytochrome b5 (native pig), its genetically engineered rat counterpart (coreŐtail), the soluble core form of the latter (core), the core with the secretory signal sequence of alkaline phosphatase appended to its N-terminal (signalŐcore) and the latter containing the C-terminal tail of the native rat protein (signalŐcoreŐtail). When examined by Edman degradation and MS, the engineered proteins were shown to have the expected N-terminal amino acid sequences and molecular masses. The native pig was found to be acetylated at the N-terminal. The native pig and coreŐtail enzymes were equally efficient at enhancing the side-chain cleavage activity of human CYP17 and the signalŐcoreŐtail was 55% as efficient. The core and signalŐcore constructs were completely inactive in the aforementioned reaction. All the five derivatives were reduced to varying degrees by NADPH:cytochrome P-450 (NADPH-P450) reductase and the relative efficiencies of this reduction were reminiscent of the behaviour of these derivatives in supporting the side-chain cleavage reaction. In the side-chain cleavage assay, however, NADPH-P450 reductase was used in large excess so that the reduction of cytochrome b5 derivatives was not rate-limiting. The results highlight that productive interaction between cytochrome b5 and CYP17 is governed not only by the presence of a membrane insertable hydrophobic region on the cytochrome b5 but also by its defined spatial orientation at the C-terminal.

scholarly journals Analysis of CRISPR in Streptococcus mutans suggests frequent occurrence of acquired immunity against infection by M102-like bacteriophages

Microbiology ◽  
2009 ◽  
Vol 155 (6) ◽  
pp. 1966-1976 ◽  
Author(s):  
Jan R. van der Ploeg
Keyword(s):  
Streptococcus Mutans ◽  
Genome Sequence ◽  
Sequence Similarity ◽  
Amino Acid Sequences ◽  
Acquired Immunity ◽  
Environment Analysis ◽  
Loss Of Resistance ◽  
Dna Elements ◽  
Resistance To Infection ◽  
Resistant Mutants

Clustered regularly interspaced short palindromic repeats (CRISPR) consist of highly conserved direct repeats interspersed with variable spacer sequences. They can protect bacteria against invasion by foreign DNA elements. The genome sequence of Streptococcus mutans strain UA159 contains two CRISPR loci, designated CRISPR1 and CRISPR2. The aims of this study were to analyse the organization of CRISPR in further S. mutans strains and to investigate the importance of CRISPR in acquired immunity to M102-like phages. The sequences of CRISPR1 and CRISPR2 arrays were determined for 29 S. mutans strains from different persons. More than half of the CRISPR1 spacers and about 35 % of the CRISPR2 spacers showed sequence similarity with the genome sequence of M102, a virulent siphophage specific for S. mutans. Although only a few spacers matched the phage sequence completely, most of the mismatches had no effect on the amino acid sequences of the phage-encoded proteins. The results suggest that S. mutans is often attacked by M102-like bacteriophages, and that its acquisition of novel phage-derived CRISPR sequences goes along with the presence of S. mutans phages in the environment. Analysis of CRISPR1 of M102-resistant mutants of S. mutans OMZ 381 showed that some of them had acquired novel spacers, and the sequences of all but one of these matched the phage M102 genome sequence. This suggests that the acquisition of the spacers contributed to the resistance against phage infection. However, since not all resistant mutants had new spacers, and since the removal of the CRISPR1 array in one of the mutants and in wild-type strains did not lead to loss of resistance to infection by M102, the acquisition of resistance must be based on further elements as well.

Effects of moderate heart failure and functional overload on rat plantaris muscle

2002 ◽  
Vol 92 (1) ◽  
pp. 18-24 ◽  
Author(s):  
Espen E. Spangenburg ◽  
Simon J. Lees ◽  
Jeff S. Otis ◽  
Timothy I. Musch ◽  
Robert J. Talmadge ◽  
...  
Keyword(s):  
Heart Failure ◽  
Skeletal Muscle ◽  
Exercise Training ◽  
Muscle Function ◽  
Skeletal Muscle Function ◽  
Moderate Heart Failure ◽  
Plasmic Reticulum ◽  
Uptake Rates ◽  
Isoform Expression ◽  
And Control

It is thought that changes in sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) of skeletal muscle contribute to alterations in skeletal muscle function during congestive heart failure (CHF). It is well established that exercise training can improve muscle function. However, it is unclear whether similar adaptations will result from exercise training in a CHF patient. Therefore, the purpose of this study was to determine whether skeletal muscle during moderate CHF adapts to increased activity, utilizing the functional overload (FO) model. Significant increases in plantaris mass of the CHF-FO and sham-FO groups compared with the CHF and control (sham) groups were observed. Ca2+ uptake rates were significantly elevated in the CHF group compared with all other groups. No differences were detected in Ca2+ uptake rates between the CHF-FO, sham, and sham-FO groups. Increases in Ca2+ uptake rates in moderate-CHF rats were not due to changes in SERCA isoform proportions; however, FO may have attenuated the CHF-induced increases through alterations in SERCA isoform expression. Therefore, changes in skeletal muscle Ca2+handling during moderate CHF may be due to alterations in regulatory mechanisms, which exercise may override, by possibly altering SERCA isoform expression.

scholarly journals Comparative analysis of ankyrin (ANK) genes of five capripoxviruses isolate strains from Xinjiang province in China

2020 ◽  
Vol 17 (1) ◽  
Author(s):  
Chuanchuan He ◽  
Jianjun Tong ◽  
Xueping Zhang ◽  
Milikaimu Tuohetiniyazi ◽  
Yu Zhang ◽  
...  
Keyword(s):  
Host Range ◽  
Target Genes ◽  
Domain Architecture ◽  
Amino Acid Sequences ◽  
Effective Prevention ◽  
Variable Regions ◽  
Sheeppox Virus ◽  
Goatpox Virus ◽  
C Terminus ◽  
And Control

Abstract Background Sheeppox and goatpox are both economically important animal diseases in which pathogens are goatpox virus (GTPV) and sheeppox virus (SPPV). They can’t cause cross-species infection between sheep and goats in general. But in recent decades, the infection of sheep by goatpox or goats by sheeppox has been reported. The literature has indicated that the occurrence of these cases has a significant and direct relationship with mutations of ankyrin genes families (ANK genes 010,138,140,141.2,145) located in two-terminal regions of capripoxvirus genomes. So it is very important to decipher these nucleotides and their coding amino acid sequences of the five genes regarded as host range and virulence factors for effective prevention and control of capripoxvirus diseases. Methods In this study, all the ankyrin genes of three goatpox virus, two sheeppox virus, and one GTPV vaccine strains from Nanjiang areas of Xinjiang province of China during 2010–2011 were collected, amplified, cloned and sequenced. The sequence of every ankyrin genes has been compared with not only sequences from six viruses but also all sequences from three species of capripoxvirus genus from Gene bank, and every ANK gene’s mutated nucleotides and amino acids have been screened, and the relationship of genetic evolution among different virus strains has been analyzed, as well as the domain architecture of these genes was forecasted and analyzed. Results The six capripoxvirus strains can be well-distinguished GTPV and SPPV based on five ANK genes’ sequence identicalness except for GTPV-SS strain, which showed higher identicalness with SPPV. The ANK gene sequence of the GTPV-SS strain was 100% identical with SPPV-M1 (ANK138,140,145) and SPPV-M2 (ANK138,145), respectively. Phylogenetically, these six capripoxvirus strains were also grouped into the same cluster of India reference strains in lineages and showed extreme identical conservative or variable regions with India capripoxvirus isolates by sequence alignment. Moreover, for the functional domains, these ANK genes of capripoxvirus except for ANK gene 145, are identical in size, and ANK genes 145 of SPPV are usually 100 bp (approximately 30 aa) longer than those of GTPV and eventually form a PRANC domain at C-terminus. Conclusions The isolated strain of GTPV-SS may be a cross-species infection or the collected material was contaminated, and the inferred Capripox outbreak in Xinjiang in 2010 can be introduced from India. ANK genes 138,140,141.2 and 145 of capripoxvirus can be used as the target genes to identify GTPV and SPPV. Moreover, the four ANK genes determining the host range are more significant than the ANK gene 010. These ANK genes play combining roles for their function.

scholarly journals Formation of Pyrazines in Maillard Model Systems: Effects of Structures of Lysine-Containing Dipeptides/Tripeptides

Foods ◽  
2021 ◽  
Vol 10 (2) ◽  
pp. 273
Author(s):  
Furong Wang ◽  
Hailiang Shen ◽  
Ting Liu ◽  
Xi Yang ◽  
Yali Yang ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Sunflower Seed ◽  
Seed Oil ◽  
Reaction System ◽  
Amino Acid Sequences ◽  
Model Systems ◽  
Sunflower Seed Oil ◽  
C Terminus ◽  
And Control

At present, most investigations involving the Maillard reaction models have focused on free amino acids (FAAs), whereas the effects of peptides on volatile products are poorly understood. In our study, the formation mechanism of pyrazines, which were detected as characteristic volatiles in sunflower seed oil, from the reaction system of glucose and lysine-containing dipeptides and tripeptides was studied. The effect of the amino acid sequences of the dipeptides and tripeptides on pyrazine formation was further highlighted. Four different dipeptides and six tripeptides were selected. The results showed that the production of pyrazines in the lysine-containing dipeptide models was higher than that in the tripeptide and control models. Compounds 2,5(6)-Dimethylpyrazine and 2,3,5-trimethylpyrazine were the main pyrazine compounds in the dipeptide models. Furthermore, the C- or N-terminal amino acids of lysine-containing dipeptides can exert an important effect on the formation of pyrazines. In dipeptide models with lysine at the C-terminus, the content of total pyrazines followed the order of Arg−Lys > His−Lys; the order of the total pyrazine content was Lys−His > Lys−Arg in dipeptide models with N-terminal lysine. Additionally, for the tripeptide models with different amino acid sequences, more pyrazines and a greater variety of pyrazines were detected in the tripeptide models with N-terminal lysine/arginine than in the tripeptide models with N-terminal histidine. However, the total pyrazine content and the percentage of pyrazines in the total volatiles were similar in the tripeptide models with the same amino acids at the N-terminus. This study clearly illustrates the ability of dipeptides and tripeptides containing lysine, arginine and histidine to form pyrazines, improving volatile formation during sunflower seed oil processing.

scholarly journals Major role for ACE-independent intrarenal ANG II formation in type II diabetes

2010 ◽  
Vol 298 (1) ◽  
pp. F37-F48 ◽  
Author(s):  
Sungmi Park ◽  
Benjamin J. Bivona ◽  
Hiroyuki Kobori ◽  
Dale M. Seth ◽  
Mark C. Chappell ◽  
...  
Keyword(s):  
Serine Protease ◽  
Type Ii Diabetes ◽  
Ace Inhibition ◽  
Receptor Blockade ◽  
Type Ii ◽  
Afferent Arteriole ◽  
Ang Ii ◽  
Protease Inhibition ◽  
Protein Activity ◽  
And Control

Combination therapy of angiotensin-converting enzyme (ACE) inhibition and AT1 receptor blockade has been shown to provide greater renoprotection than ACE inhibitor alone in human diabetic nephropathy, suggesting that ACE-independent pathways for ANG II formation are of major significance in disease progression. Studies were performed to determine the magnitude of intrarenal ACE-independent formation of ANG II in type II diabetes. Although renal cortical ACE protein activity [2.1 ± 0.8 vs. 9.2 ± 2.1 arbitrary fluorescence units (AFU)·mg−1·min−1] and intensity of immunohistochemical staining were significantly reduced and ACE2 protein activity (16.7 ± 3.2 vs. 7.2 ± 2.4 AFU·mg−1·min−1) and intensity elevated, kidney ANG I (113 ± 24 vs. 110 ± 45 fmol/g) and ANG II (1,017 ± 165 vs. 788 ± 99 fmol/g) levels were not different between diabetic and control mice. Afferent arteriole vasoconstriction due to conversion of ANG I to ANG II was similar in magnitude in kidneys of diabetic (−28 ± 3% at 1 μM) and control (−23 ± 3% at 1 μM) mice; a response completely inhibited by AT1 receptor blockade. In control kidneys, afferent arteriole vasoconstriction produced by ANG I was significantly attenuated by ACE inhibition, but not by serine protease inhibition. In contrast, afferent arteriole vasoconstriction produced by intrarenal conversion of ANG I to ANG II was significantly attenuated by serine protease inhibition, but not by ACE inhibition in diabetic kidneys. In conclusion, there is a switch from ACE-dependent to serine protease-dependent ANG II formation in the type II diabetic kidney. Pharmacological targeting of these serine protease-dependent pathways may provide further protection from diabetic renal vascular disease.

scholarly journals Peptide Characterization of Mature Fluorotic and Control Human Enamel

2016 ◽  
Vol 27 (1) ◽  
pp. 66-71 ◽  
Author(s):  
Isabel Maria Porto Lelis ◽  
Gabriela F. Molina ◽  
Cláudia Souza ◽  
Walter B. Perez ◽  
Helen J. Laure ◽  
...  
Keyword(s):  
Protein Extraction ◽  
Proteinase Inhibitors ◽  
Time Of Flight ◽  
Amino Acid Sequences ◽  
Ionization Time ◽  
Flight Mass Spectrometry ◽  
Human Enamel ◽  
High Fluoride ◽  
And Control

Abstract Exposure to high fluoride levels during amelogenesis causes enamel fluorosis. This study aimed to determine and compare the amino acid sequences in the enamel of fluorotic and control teeth. This investigation included enamel samples obtained from erupted and non-erupted third molars with either TF grade 4-6 (n=7) fluorosis or no sign of fluorosis (controls, n=7). The samples were kept frozen at -20 °C until protein extraction. Samples were etched and processed with a cocktail of proteinase inhibitors and immediately analyzed. Matrix Assisted Laser Desorption/Ionization-Time-Of-Flight/Time-of-Flight Mass Spectrometry (MALDI-TOF/TOF) followed by MASCOT search aided the peptides analysis. The more abundant peptides bore the N-terminal amelogenin sequences WYQSIRPPYP (which is specific for the X-encoded amelogenin) and MPLPPHPGHPGYINF (which does not show sexual dimorphism) were not different in control or fluorotic enamel. There was no missing proteolytic cleavage in the fluorotic samples, which suggested that the increased amount of protein described in fluorotic enamel did not stem from the decreased ability of proteinases to cleave the proteins in humans. This study showed how to successfully obtain peptide from superficial enamel. A relatively low number of teeth was sufficient to provide good data on the actual peptides found in mature enamel.

Abstract 346: Cardiac Expression of Fetal and Adult SCN5A Isoforms in Sudden Unexplained Infant Death (SUID)

2020 ◽  
Vol 127 (Suppl_1) ◽  
Author(s):  
Alexandra F Williams ◽  
Kelsey Tomasek ◽  
Kyle Gregory ◽  
Carlos Fulmer ◽  
Cole Bozeman ◽  
...  
Keyword(s):  
Sodium Current ◽  
Heart Tissue ◽  
Vital Role ◽  
Developmental Expression ◽  
Rna Integrity ◽  
Significant Difference ◽  
Gated Channel ◽  
Isoform Expression ◽  
And Control ◽  
Potential Substrate

Sodium Voltage-Gated Channel Alpha Subunit 5 (SNC5A) plays a vital role in cardiac depolarization and has been implicated in Long QT Syndrome and sudden cardiac death. SCN5A encodes fetal and adult isoforms, differing by splicing of exon 6A or 6B, respectively. Fetal SCN5A decreases peak sodium current, leading to slower channel activation and inactivation and providing a potential substrate for arrhythmia. Little is known about developmental expression of these isoforms. We investigated developmental expression of SCN5A isoforms in infants, toddlers, and adults, including expression ratios in SUID cases and controls (0-12 months). We extracted RNA from heart tissue collected at autopsy (fetus-24 months) with 80 SUID cases and 27 controls, and 5 adult donor hearts. RNA Integrity Numbers ranged from 4.8-9.5. Relative SCN5A isoform expression was quantified by qRT-PCR. Non-parametric t-test and ANOVA were used; P<0.05 was considered statistically significant. Our results show a significant stepwise increase in adult/fetal isoform expression with age. Relative expression of adult/fetal SCN5A increased significantly from fetal-4 months to 5-24 months (P<0.0001), and again in adulthood (P<0.0001). There was no significant difference between case and control groups at 0-4 or 0-12 months. Expression of adult/fetal SCN5A increases significantly from birth to 24 months of age and into adulthood. Relative adult/fetal SCN5A expression is lowest during the neonatal period through 4 months, corresponding with the peak incidence of SUID. Further investigation of protein expression of SCN5A isoforms may shed light onto the age-dependent incidence of sudden death.

scholarly journals Chromatin network markers of leukemia

2020 ◽  
Vol 36 (Supplement_1) ◽  
pp. i455-i463
Author(s):  
N Malod-Dognin ◽  
V Pancaldi ◽  
A Valencia ◽  
N Pržulj
Keyword(s):  
Lymphocytic Leukemia ◽  
Supplementary Information ◽  
Cell Of Origin ◽  
Complement Protein ◽  
Driver Genes ◽  
Chromosome Conformation ◽  
The Road ◽  
Cellular Processes ◽  
Dna Elements ◽  
And Control

Abstract Motivation The structure of chromatin impacts gene expression. Its alteration has been shown to coincide with the occurrence of cancer. A key challenge is in understanding the role of chromatin structure (CS) in cellular processes and its implications in diseases. Results We propose a comparative pipeline to analyze CSs and apply it to study chronic lymphocytic leukemia (CLL). We model the chromatin of the affected and control cells as networks and analyze the network topology by state-of-the-art methods. Our results show that CSs are a rich source of new biological and functional information about DNA elements and cells that can complement protein–protein and co-expression data. Importantly, we show the existence of structural markers of cancer-related DNA elements in the chromatin. Surprisingly, CLL driver genes are characterized by specific local wiring patterns not only in the CS network of CLL cells, but also of healthy cells. This allows us to successfully predict new CLL-related DNA elements. Importantly, this shows that we can identify cancer-related DNA elements in other cancer types by investigating the CS network of the healthy cell of origin, a key new insight paving the road to new therapeutic strategies. This gives us an opportunity to exploit chromosome conformation data in healthy cells to predict new drivers. Availability and implementation Our predicted CLL genes and RNAs are provided as a free resource to the community at https://life.bsc.es/iconbi/chromatin/index.html. Supplementary information Supplementary data are available at Bioinformatics online.

Evaluation of 2-Year-Old Intrasplenic Fetal Liver Tissue Transplants in Rats

2003 ◽  
Vol 12 (4) ◽  
pp. 423-438 ◽  
Author(s):  
Amelie Lupp ◽  
Manfred Danz ◽  
Dieter Müller
Keyword(s):  
Liver Tissue ◽  
Fetal Liver ◽  
Male Rats ◽  
Surrounding Tissue ◽  
Transplant Recipients ◽  
Glutathione S Transferase ◽  
Tissue Transplants ◽  
Preneoplastic Foci ◽  
Isoform Expression ◽  
And Control

Liver cell transplantation into host organs like the spleen may possibly provide a temporary relief after extensive liver resection or severe liver disease or may enable treatment of an enzyme deficiency. With time, however, dedifferentiation or malignant transformation of the ectopically transplanted cells may be possible. Thus, in the present study syngenic fetal liver tissue suspensions were transplanted into the spleen of adult male rats and evaluated 2 years thereafter in comparison to orthotopic livers for histopathological changes and (as markers for preneoplastic transformation) for cytochrome P450 (P450) and glutathione S-transferase (GST) isoform expression. Because inducibility of P450 and GST isoforms may be changed in preneoplastic foci, prior to sacrifice animals were additionally treated either with β-naphthoflavone, phenobarbital, dexamethasone, or the respective solvent. In the 2-year-old grafts more than 70% of the spleen mass was occupied by the transplant. The transplanted hepatocytes were arranged in cord-like structures. Also few bile ducts were present. Morphologically, no signs of malignancy were visible. With all rats, transplant recipients as well as controls, however, discrete nodular structures were seen in the livers. Due to age, both livers and transplants displayed only a low P450 2B1 and 3A2 and GST class α and μ isoform expression. No immunostaining for P450 1A1 was visible. At both sites, β-naphthoflavone, phenobarbital, or dexamethasone treatment enhanced P450 1A1, P450 2B1 and 3A2, or P450 3A2 expression, respectively. No immunostaining for GST class π isoforms was seen in the transplants. The livers of both transplant recipients and control rats, however, displayed GST π-positive foci, corresponding to the nodular structures seen histomorphologically. Compared to the surrounding tissue, these foci also exhibited a more pronounced staining for GST class α and μ isoforms and a stronger inducibility of the P450 1A1 expression due to β-naphthoflavone. In conclusion, in contrast to the livers, no preneoplastic foci seem to appear in the intrasplenic transplants even 2 years after transplantation. This may be due either to the protection of these transplants by the orthotopic livers or to the different humoral and nerval influences at the ectopic site.

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