scholarly journals Role of conserved hydrophobic amino acids in androgen receptor AF-1 function

2003 ◽  
Vol 31 (3) ◽  
pp. 427-439 ◽  
Author(s):  
R Betney ◽  
IJ McEwan
Keyword(s):  
Amino Acids ◽  
Transcription Factor ◽  
Androgen Receptor ◽  
Gene Transcription ◽  
Hydrophobic Residues ◽  
Steroid Receptor Coactivator ◽  
Binding Interface ◽  
Hydrophobic Amino Acids

The intracellular androgen receptor (AR) is a ligand-activated transcription factor. Upon binding the steroids testosterone or dihydrotestosterone, the activated receptor translocates to the nucleus, binds to specific DNA response elements and interacts with the transcription machinery in order to regulate gene transcription. In the present study, we have described a highly conserved region (amino acids 224-258) within the AR AF-1 domain and have investigated the role of conserved bulky hydrophobic residues in gene regulation. Mutating pairs of residues (I229A/L236A; V240A/V242A; L251A/L254A) reduced transactivation activity by 25-40%. Mutating residues M244, L246 and V248 to alanines had a more dramatic affect on receptor activity, disrupting activity by at least 60%. The latter mutations also disrupted binding to the RNA polymerase-associated protein 74 subunit of the general transcription factor TFIIF. The protein conformation and stability of the mutant polypeptide in vitro was not significantly different from the wild type. None of the mutations tested disrupted binding of the AF-1 domain with the coactivator protein steroid receptor coactivator-1a. Thus we have concluded that conserved hydrophobic residues are important for receptor-dependent gene transcription and that M244, L246 and V248 are part of the binding interface for TFIIF.

scholarly journals Substrate specificities of Escherichia coli ItaT that acetylates aminoacyl-tRNAs

2020 ◽  
Author(s):  
Chuqiao Zhang ◽  
Yuka Yashiro ◽  
Yuriko Sakaguchi ◽  
Tsutomu Suzuki ◽  
Kozo Tomita
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Group ◽  
Hydrophobic Pocket ◽  
Hydrophobic Residues ◽  
Substrate Specificities ◽  
E Coli ◽  
Hydrophobic Amino Acids

Abstract Escherichia coli ItaT toxin reportedly acetylates the α-amino group of the aminoacyl-moiety of Ile-tRNAIle specifically, using acetyl-CoA as an acetyl donor, thereby inhibiting protein synthesis. The mechanism of the substrate specificity of ItaT had remained elusive. Here, we present functional and structural analyses of E. coli ItaT, which revealed the mechanism of ItaT recognition of specific aminoacyl-tRNAs for acetylation. In addition to Ile-tRNAIle, aminoacyl-tRNAs charged with hydrophobic residues, such as Val-tRNAVal and Met-tRNAMet, were acetylated by ItaT in vivo. Ile-tRNAIle, Val-tRNAVal and Met-tRNAMet were acetylated by ItaT in vitro, while aminoacyl-tRNAs charged with other hydrophobic residues, such as Ala-tRNAAla, Leu-tRNALeu and Phe-tRNAPhe, were less efficiently acetylated. A comparison of the structures of E. coli ItaT and the protein N-terminal acetyltransferase identified the hydrophobic residues in ItaT that possibly interact with the aminoacyl moiety of aminoacyl-tRNAs. Mutations of the hydrophobic residues of ItaT reduced the acetylation activity of ItaT toward Ile-tRNAIlein vitro, as well as the ItaT toxicity in vivo. Altogether, the size and shape of the hydrophobic pocket of ItaT are suitable for the accommodation of the specific aminoacyl-moieties of aminoacyl-tRNAs, and ItaT has broader specificity toward aminoacyl-tRNAs charged with certain hydrophobic amino acids.

scholarly journals Cryo-EM structure of amyloid fibrils formed by the entire low complexity domain of TDP-43

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Qiuye Li ◽  
W. Michael Babinchak ◽  
Witold K. Surewicz
Keyword(s):  
Amyloid Fibrils ◽  
Low Complexity ◽  
Structural Features ◽  
Protein Fragments ◽  
Hydrophobic Residues ◽  
Backbone Conformation ◽  
Hydrophobic Amino Acids ◽  
Insight Into ◽  
Lateral Sclerosis

AbstractAmyotrophic lateral sclerosis and several other neurodegenerative diseases are associated with brain deposits of amyloid-like aggregates formed by the C-terminal fragments of TDP-43 that contain the low complexity domain of the protein. Here, we report the cryo-EM structure of amyloid formed from the entire TDP-43 low complexity domain in vitro at pH 4. This structure reveals single protofilament fibrils containing a large (139-residue), tightly packed core. While the C-terminal part of this core region is largely planar and characterized by a small proportion of hydrophobic amino acids, the N-terminal region contains numerous hydrophobic residues and has a non-planar backbone conformation, resulting in rugged surfaces of fibril ends. The structural features found in these fibrils differ from those previously found for fibrils generated from short protein fragments. The present atomic model for TDP-43 LCD fibrils provides insight into potential structural perturbations caused by phosphorylation and disease-related mutations.

scholarly journals Progression and Differentiation of Alveolar Rhabdomyosarcoma Is Regulated by PAX7 Transcription Factor—Significance of Tumor Subclones

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1870
Author(s):  
Klaudia Skrzypek ◽  
Grażyna Adamek ◽  
Marta Kot ◽  
Bogna Badyra ◽  
Marcin Majka
Keyword(s):  
Transcription Factor ◽  
Cell Lines ◽  
Migration Activity ◽  
Id Proteins ◽  
Rh30 Cell ◽  
Str Profile ◽  
And Migration

Rhabdomyosarcoma (RMS), is the most frequent soft tissue tumor in children that originates from disturbances in differentiation process. Mechanisms leading to the development of RMS are still poorly understood. Therefore, by analysis of two RMS RH30 cell line subclones, one subclone PAX7 negative, while the second one PAX7 positive, and comparison with other RMS cell lines we aimed at identifying new mechanisms crucial for RMS progression. RH30 subclones were characterized by the same STR profile, but different morphology, rate of proliferation, migration activity and chemotactic abilities in vitro, as well as differences in tumor morphology and growth in vivo. Our analysis indicated a different level of expression of adhesion molecules (e.g., from VLA and ICAM families), myogenic microRNAs, such as miR-206 and transcription factors, such as MYOD, MYOG, SIX1, and ID. Silencing of PAX7 transcription factor with siRNA confirmed the crucial role of PAX7 transcription factor in proliferation, differentiation and migration of RMS cells. To conclude, our results suggest that tumor cell lines with the same STR profile can produce subclones that differ in many features and indicate crucial roles of PAX7 and ID proteins in the development of RMS.

scholarly journals Antiproliferative Effect of Androgen Receptor Inhibition in Mesenchymal Stem-Like Triple-Negative Breast Cancer

2016 ◽  
Vol 38 (3) ◽  
pp. 1003-1014 ◽  
Author(s):  
Aiyu Zhu ◽  
Yan Li ◽  
Wei Song ◽  
Yumei Xu ◽  
Fang Yang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Androgen Receptor ◽  
Cyclin D1 ◽  
Cancer Cells ◽  
Triple Negative Breast Cancer ◽  
Breast Cancer Cells ◽  
Triple Negative

Background/Aims: Androgen receptor (AR), a steroid hormone receptor, has recently emerged as prognostic and treatment-predictive marker in breast cancer. Previous studies have shown that AR is widely expressed in up to one-third of triple-negative breast cancer (TNBC). However, the role of AR in TNBC is still not fully understood, especially in mesenchymal stem-like (MSL) TNBC cells. Methods: MSL TNBC MDA-MB-231 and Hs578T breast cancer cells were exposed to various concentration of agonist 5-α-dihydrotestosterone (DHT) or nonsteroidal antagonist bicalutamide or untreated. The effects of AR on cell viability and apoptosis were determined by MTT assay, cell counting, flow cytometry analysis and protein expression of p53, p73, p21 and Cyclin D1 were analyzed by western blotting. The bindings of AR to p73 and p21 promoter were detected by ChIP assay. MDA-MB-231 cells were transplanted into nude mice and the tumor growth curves were determined and expression of AR, p73 and p21 were detected by Immunohistochemistry (IHC) staining after treatment of DHT or bicalutamide. Results: We demonstrate that AR agonist DHT induces MSL TNBC breast cancer cells proliferation and inhibits apoptosis in vitro. Similarly, activated AR significantly increases viability of MDA-MB-231 xenografts in vivo. On the contrary, AR antagonist, bicalutamide, causes apoptosis and exerts inhibitory effects on the growth of breast cancer. Moreover, DHT-dependent activation of AR involves regulation in the cell cycle related genes, including p73, p21 and Cyclin D1. Further investigations indicate the modulation of AR on p73 and p21 mediated by direct binding of AR to their promoters, and DHT could make these binding more effectively. Conclusions: Our study demonstrates the tumorigenesis role of AR and the inhibitory effect of bicalutamide in AR-positive MSL TNBC both in vitro and in vivo, suggesting that AR inhibition could be a potential therapeutic approach for AR-positive TNBC patients.

scholarly journals N-terminal Hydrophobic Amino Acids of Activating Transcription Factor 5 (ATF5) Protein Confer Interleukin 1β (IL-1β)-induced Stabilization

2013 ◽  
Vol 289 (7) ◽  
pp. 3888-3900 ◽  
Author(s):  
Takanori Abe ◽  
Masaki Kojima ◽  
Satoshi Akanuma ◽  
Hiromi Iwashita ◽  
Takashi Yamazaki ◽  
...  
Keyword(s):  
Amino Acids ◽  
Transcription Factor ◽  
Interleukin 1Β ◽  
Hydrophobic Amino Acids ◽  
Activating Transcription Factor

The use of siRNA as a pharmacological tool to assess a role for the transcription factor NF-IL6 in the brain under in vitro and in vivo conditions during LPS-induced inflammatory stimulation

2017 ◽  
Vol 28 (6) ◽  
Author(s):  
Jelena Damm ◽  
Joachim Roth ◽  
Rüdiger Gerstberger ◽  
Christoph Rummel
Keyword(s):  
Transcription Factor ◽  
Brain Cells ◽  
Nuclear Factor ◽  
Si Rna ◽  
The Brain ◽  
Deficient Mice ◽  
Transcription Factor Nuclear Factor

AbstractBackground:Studies with NF-IL6-deficient mice indicate that this transcription factor plays a dual role during systemic inflammation with pro- and anti-inflammatory capacities. Here, we aimed to characterize the role of NF-IL6 specifically within the brain.Methods:In this study, we tested the capacity of short interfering (si) RNA to silence the inflammatory transcription factor nuclear factor-interleukin 6 (NF-IL6) in brain cells underResults:In cells of a mixed neuronal and glial primary culture from the ratConclusions:This approach was, thus, not suitable to characterize the role NF-IL6 in the brain

scholarly journals Enhancement of aromatase activity by D-aspartic acid in the ovary of the lizard Podarcis s. sicula

Reproduction ◽  
2001 ◽  
pp. 803-808 ◽  
Author(s):  
L Assisi ◽  
V Botte ◽  
A D'Aniello ◽  
MM Di Fiore
Keyword(s):  
Amino Acids ◽  
Aspartic Acid ◽  
Reproductive Cycle ◽  
Catalytic Properties ◽  
Ovarian Tissue ◽  
Experimental Treatment ◽  
Tissue Homogenate ◽  
Aromatase Activity

The present study investigated the role of D-aspartic acid (D-Asp) in ovarian steroidogenesis and its effect on aromatase activity in the lizard, Podarcis s. sicula. It was determined that D-Asp concentrations vary significantly during phases of the reproductive cycle: they vary inversely with testosterone concentrations and directly with oestradiol concentrations in the ovary and plasma. Experimental treatment showed that administration of D-Asp induces a decrease in testosterone and an increase in oestradiol, and that treatment with other amino acids (L-Asp, D-Glu and D-Ala) instead of D-Asp has no effects. Experiments in vitro confirmed these results. Furthermore, these experiments showed an increase in aromatase activity, as the addition of D-Asp either to fresh ovarian tissue homogenate or to acetonic powder of ovarian follicles induced a significant increase in the conversion of testosterone to oestradiol. Aromatase activity is four times greater in the presence of D-Asp than in its absence. However, almost equivalent values of the two K(m) values (both approximately 25 nmol l(-1)) indicate that aromatase has the same catalytic properties in both cases.

scholarly journals Differential role of cAMP, cGMP and Ca2+ and involvement of kinases and CBP-CREB-CRE pathway in regulation of arylalkylamine N-acetyltransferase 2 gene in the pineal organ of air-breathing catfish, Clarias gariepinus

2021 ◽  
Author(s):  
Hijam Nonibala ◽  
Braj Bansh Prasad Gupta
Keyword(s):  
Gene Expression ◽  
Map Kinase ◽  
Gene Transcription ◽  
Pineal Organ ◽  
Clarias Gariepinus ◽  
Specific Inhibitor ◽  
P38 Map Kinase ◽  
Camp And Cgmp

Abstract Transcription of arylalkylamine N-acetyltransferase 2 (aanat2) gene leads to formation of AANAT2 - the rate-limiting enzyme in melatonin synthesis pathway in photosensitive fish pineal organ. However, unlike in avian and mammalian pineal gland, there is practically no information on signal transduction pathway(s) involved in regulation of aanat2 gene transcription in the fish pineal organ. Therefore, we investigated the role of important molecular components of signalling via cAMP, cGMP, Ca2+ involving PKA, PKG, PKC, MeK and p38 MAP kinase as well as possible role of serine/threonine phosphatases, CREB and CBP using their specific inhibitors and/or activators in aanat2 gene transcription in the fish pineal organ maintained under in vitro culture-conditions. db-cAMP and db-cGMP stimulated the expression of aanat2 gene. db-cAMP- and cGMP-induced aanat2 gene expression was significantly reduced in the presence of H-89 (specific inhibitor of PKA), KT5823 (specific inhibitor of PKG), chelerythrine chloride (specific inhibitor of PKC), U0126 ethanolate (specific inhibitor of MeK) and SB 202190 monohydrochloride hydrate (specific inhibitor of p38 MAP kinase). Inhibitors of PP1 and PP2A significantly increased aanat2 gene expression as well as significantly reduced cAMP- and cGMP-induced gene transcription, while inhibitor of PP2B had no effect on aanat2 gene expression. Inhibitors of both CREB and CBP-CREB interaction completely blocked cAMP-induced aanat2 gene transcription. Based on these findings, we suggest that cAMP, cGMP and Ca2+ stimulate aanat2 gene transcription via PKA, PKG and PKC, respectively. Further, protein phosphatases and CBP-CREB-CRE pathway are actively involved in regulation of on aanat2 gene expression in the fish pineal organ.

scholarly journals Lineage-specific requirements of β-catenin in neural crest development

2002 ◽  
Vol 159 (5) ◽  
pp. 867-880 ◽  
Author(s):  
Lisette Hari ◽  
Véronique Brault ◽  
Maurice Kléber ◽  
Hye-Youn Lee ◽  
Fabian Ille ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Neural Crest ◽  
Neural Crest Cells ◽  
Neural Crest Stem Cells ◽  
Downstream Signaling ◽  
Neural Crest Development ◽  
Sensory Neurogenesis

β-Catenin plays a pivotal role in cadherin-mediated cell adhesion. Moreover, it is a downstream signaling component of Wnt that controls multiple developmental processes such as cell proliferation, apoptosis, and fate decisions. To study the role of β-catenin in neural crest development, we used the Cre/loxP system to ablate β-catenin specifically in neural crest stem cells. Although several neural crest–derived structures develop normally, mutant animals lack melanocytes and dorsal root ganglia (DRG). In vivo and in vitro analyses revealed that mutant neural crest cells emigrate but fail to generate an early wave of sensory neurogenesis that is normally marked by the transcription factor neurogenin (ngn) 2. This indicates a role of β-catenin in premigratory or early migratory neural crest and points to heterogeneity of neural crest cells at the earliest stages of crest development. In addition, migratory neural crest cells lateral to the neural tube do not aggregate to form DRG and are unable to produce a later wave of sensory neurogenesis usually marked by the transcription factor ngn1. We propose that the requirement of β-catenin for the specification of melanocytes and sensory neuronal lineages reflects roles of β-catenin both in Wnt signaling and in mediating cell–cell interactions.

Insights in the molecular mechanisms of an azole stress adapted laboratory-generated Aspergillus fumigatus strain

2021 ◽  
Author(s):  
Marion Aruanno ◽  
Samantha Gozel ◽  
Isabelle Mouyna ◽  
Josie E Parker ◽  
Daniel Bachmann ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Aspergillus Fumigatus ◽  
Molecular Mechanisms ◽  
Drug Transporters ◽  
Major Facilitator Superfamily ◽  
Ergosterol Biosynthesis ◽  
Azole Resistance ◽  
Drug Transporter

Abstract Aspergillus fumigatus is the main cause of invasive aspergillosis, for which azole drugs are the first-line therapy. Emergence of pan-azole resistance among A. fumigatus is concerning and has been mainly attributed to mutations in the target gene (cyp51A). However, azole resistance may also result from other mutations (hmg1, hapE) or other adaptive mechanisms. We performed microevolution experiment exposing an A. fumigatus azole-susceptible strain (Ku80) to sub-minimal inhibitory concentration of voriconazole to analyze emergence of azole resistance. We obtained a strain with pan-azole resistance (Ku80R), which was partially reversible after drug relief, and without mutations in cyp51A, hmg1, and hapE. Transcriptomic analyses revealed overexpression of the transcription factor asg1, several ATP-binding cassette (ABC) and major facilitator superfamily transporters and genes of the ergosterol biosynthesis pathway in Ku80R. Sterol analysis showed a significant decrease of the ergosterol mass under voriconazole exposure in Ku80, but not in Ku80R. However, the proportion of the sterol compounds was similar between both strains. To further assess the role of transporters, we used the ABC transporter inhibitor milbemycine oxime (MLB). MLB inhibited transporter activity in both Ku80 and Ku80R and demonstrated some potentiating effect on azole activity. Criteria for synergism were reached for MLB and posaconazole against Ku80. Finally, deletion of asg1 revealed some role of this transcription factor in controlling drug transporter expression, but had no impact on azole susceptibility. This work provides further insight in mechanisms of azole stress adaptation and suggests that drug transporters inhibition may represent a novel therapeutic target. Lay Summary A pan-azole-resistant strain was generated in vitro, in which drug transporter overexpression was a major trait. Analyses suggested a role of the transporter inhibitor milbemycin oxime in inhibiting drug transporters and potentiating azole activity.

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