scholarly journals Histone H3K4 and H3K36 methylation promotes recruitment, but not activity, of the NuA3 histone acetyltransferase complex in S. cerevisiae

2016 ◽  
Author(s):  
Benjamin J.E. Martin ◽  
Kristina L. McBurney ◽  
Vicki E. Maltby ◽  
Kristoffer N. Jensen ◽  
Julie Brind’Amour ◽  
...  
Keyword(s):  
Histone Acetyltransferase ◽  
Pwwp Domain ◽  
H3k36 Methylation ◽  
Phd Finger ◽  
Binding Domains ◽  
Histone Ptms ◽  
Histone H3k4 ◽  
Multiple Domains

AbstractHistone post-translational modifications (PTMs) alter chromatin structure by promoting the interaction of chromatin-modifying complexes with nucleosomes. The majority of chromatin-modifying complexes contain multiple domains that preferentially interact with modified histones, leading to speculation that these domains function in concert to target nucleosomes with distinct combinations of histone PTMs. In S. cerevisiae, the NuA3 histone acetyltransferase complex contains three domains, the PHD finger in Yng1, the PWWP domain in Pdp3, and the YEATS domain in Taf14, which in vitro bind to H3K4 methylation, H3K36 methylation, and acetylated and crotonylated H3K9 respectively. However the relative in vivo contributions of these histone PTMs in targeting NuA3 is unknown. Here we show that in vivo H4K4 and H3K36 methylation, but not acetylated or crotonylated H3K9, recruit NuA3 to transcribed genes. Through genome-wide colocalization and by mutational interrogation, we demonstrate that the PHD finger of Yng1, and the PWWP domain of Pdp3 independently target NuA3 to H3K4 and H3K36 methylated chromatin respectively. In contrast, we find no evidence to support the YEATS domain of Taf14 functioning in NuA3 recruitment. Collectively our results suggest that the presence of multiple histone-PTM binding domains within NuA3, rather than restricting it to nucleosomes containing distinct combinations of histone PTMs, can serve to increase the range of nucleosomes bound by the complex. Interestingly however, the simple presence of NuA3 is insufficient to ensure acetylation of the associated nucleosomes, suggesting a secondary level of acetylation regulation that does not involve control of HAT-nucleosome interactions

scholarly journals H3K36 methylation and DNA-binding are critical for Ioc4 recruitment and Isw1b remodeller function

2021 ◽  
Author(s):  
Jian Li ◽  
Lena Bergmann ◽  
Kimberly M Webb ◽  
Madelaine M Gogol ◽  
Philipp Voigt ◽  
...  
Keyword(s):  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Functional Characterization ◽  
Full Length ◽  
Pwwp Domain ◽  
H3k36 Methylation ◽  
Chromatin Remodelling Complex

The Isw1b chromatin-remodelling complex is specifically recruited to gene bodies to help retain pre-existing histones during transcription by RNA polymerase II. Recruitment is dependent on H3K36 methylation and the Isw1b subunit Ioc4, which contains an N-terminal PWWP domain. Here, we present the crystal structure of the Ioc4-PWWP domain including a detailed functional characterization of the domain on its own as well as in the context of full-length Ioc4 and the Isw1b remodeller. Ioc4-PWWP preferentially binds H3K36me3-containing nucleosomes. The ability of the PWWP domain to bind DNA is required for this interaction. It is also promoted by the unique insertion motif present in Ioc4-PWWP. The ability to bind H3K36me3 as well as DNA are also critical for full-length Ioc4 binding to nucleosomes in vitro as well as its recruitment to gene bodies in vivo. Furthermore, a fully functional Ioc4-PWWP domain is necessary for efficient remodelling by Isw1b and the maintenance of ordered chromatin in vivo, thereby preventing intragenic transcription initiation and the production of non-coding RNAs.

scholarly journals Role of Papillomavirus E1 Initiator Dimerization in DNA Replication

2005 ◽  
Vol 79 (13) ◽  
pp. 8661-8664 ◽  
Author(s):  
Stephen Schuck ◽  
Arne Stenlund
Keyword(s):  
Dna Replication ◽  
Dna Binding ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Severe Effect ◽  
Origin Of Dna Replication ◽  
Initiation Of Dna Replication

ABSTRACT Viral initiator proteins are polypeptides that form oligomeric complexes on the origin of DNA replication (ori). These complexes carry out a multitude of functions related to initiation of DNA replication, and although many of these functions have been characterized biochemically, little is understood about how the complexes are assembled. Here we demonstrate that loss of one particular interaction, the dimerization between E1 DNA binding domains, has a severe effect on DNA replication in vivo but has surprisingly modest effects on most individual biochemical activities in vitro. We conclude that the dimer interaction is primarily required for initial recognition of ori.

scholarly journals The HOX Homeodomain Proteins Block CBP Histone Acetyltransferase Activity

2001 ◽  
Vol 21 (21) ◽  
pp. 7509-7522 ◽  
Author(s):  
Wei-fang Shen ◽  
Keerthi Krishnan ◽  
H. J. Lawrence ◽  
Corey Largman
Keyword(s):  
Dna Binding ◽  
Protein Function ◽  
Histone Acetyltransferase ◽  
Gene Activation ◽  
Homeodomain Proteins ◽  
Hox Proteins ◽  
Reporter Assays ◽  
Hox Protein

ABSTRACT Despite the identification of PBC proteins as cofactors that provide DNA affinity and binding specificity for the HOX homeodomain proteins, HOX proteins do not demonstrate robust activity in transient-transcription assays and few authentic downstream targets have been identified for these putative transcription factors. During a search for additional cofactors, we established that each of the 14 HOX proteins tested, from 11 separate paralog groups, binds to CBP or p300. All six isolated homeodomain fragments tested bind to CBP, suggesting that the homeodomain is a common site of interaction. Surprisingly, CBP-p300 does not form DNA binding complexes with the HOX proteins but instead prevents their binding to DNA. The HOX proteins are not substrates for CBP histone acetyltransferase (HAT) but instead inhibit the activity of CBP in both in vitro and in vivo systems. These mutually inhibitory interactions are reflected by the inability of CBP to potentiate the low levels of gene activation induced by HOX proteins in a range of reporter assays. We propose two models for HOX protein function: (i) HOX proteins may function without CBP HAT to regulate transcription as cooperative DNA binding molecules with PBX, MEIS, or other cofactors, and (ii) the HOX proteins may inhibit CBP HAT activity and thus function as repressors of gene transcription.

Identification of the functional domain of p21WAF1/CIP1 that protects cells from cisplatin cytotoxicity

2005 ◽  
Vol 289 (3) ◽  
pp. F514-F520 ◽  
Author(s):  
Fang Yu ◽  
Judit Megyesi ◽  
Robert L. Safirstein ◽  
Peter M. Price
Keyword(s):  
Dominant Negative ◽  
Nuclear Antigen ◽  
Functional Domain ◽  
Binding Domains ◽  
Cdk2 Activity ◽  
Localization Signal ◽  
Induced Apoptosis ◽  
Proliferating Cell

The p21 cyclin-dependent kinase (cdk) inhibitor protects cells from cisplatin cytotoxicity in vivo and in vitro. However, the mechanism of protection is not known. Separate p21 domains are known to interact with several different proteins having proapoptotic functions. To investigate the mechanism of protection by p21, we have constructed adenoviruses encoding the different domains of p21. We were able to localize the protective activity to a region of 54 amino acids containing the cyclin-cdk interacting moiety. Other protein binding domains of p21, including the NH2-terminal procaspase-3 interactive region and the COOH-terminal region containing the proliferating cell nuclear antigen binding domain and the nuclear localization signal, had little protective effect on cisplatin cytotoxicity. The dependence of cisplatin cytotoxicity on cdk2 activity was also demonstrated because 1) cisplatin caused a marked increase in cdk2 activity, which was prevented by the p21 expression adenovirus, and 2) a cdk2 dominant-negative adenovirus also protected cells from cisplatin-induced apoptosis. Thus the data suggest that the mechanism of p21 protection is by direct inhibition of cdk2 activity and that cisplatin-induced apoptosis is caused by a cdk2-dependent pathway.

scholarly journals Fused protein domains inhibit DNA binding by LexA.

1992 ◽  
Vol 12 (7) ◽  
pp. 3006-3014 ◽  
Author(s):  
E A Golemis ◽  
R Brent
Keyword(s):  
Dna Binding ◽  
Protein Interactions ◽  
In Vitro Assays ◽  
Dimerization Domain ◽  
Activation Domains ◽  
Dna Binding Domains ◽  
Binding Domains ◽  
Operator Binding

Many studies of transcription activation employ fusions of activation domains to DNA binding domains derived from the bacterial repressor LexA and the yeast activator GAL4. Such studies often implicitly assume that DNA binding by the chimeric proteins is equivalent to that of the protein donating the DNA binding moiety. To directly investigate this issue, we compared operator binding by a series of LexA-derivative proteins to operator binding by native LexA, by using both in vivo and in vitro assays. We show that operator binding by many proteins such as LexA-Myc, LexA-Fos, and LexA-Bicoid is severely impaired, while binding of other LexA-derivative proteins, such as those that carry bacterially encoded acidic sequences ("acid blobs"), is not. Our results also show that DNA binding by LexA derivatives that contain the LexA carboxy-terminal dimerization domain (amino acids 88 to 202) is considerably stronger than binding by fusions that lack it and that heterologous dimerization motifs cannot substitute for the LexA88-202 function. These results suggest the need to reevaluate some previous studies of activation that employed LexA derivatives and modifications to recent experimental approaches that use LexA and GAL4 derivatives to detect and study protein-protein interactions.

scholarly journals Yin Yang 1 is required for PHD finger protein 20-mediated myogenic differentiation in vitro and in vivo

2020 ◽  
Vol 27 (12) ◽  
pp. 3321-3336
Author(s):  
Hyunji Lee ◽  
Youngeun Hong ◽  
Gyeyeong Kong ◽  
Dong Hoon Lee ◽  
Minhee Kim ◽  
...  
Keyword(s):  
Myogenic Differentiation ◽  
Phd Finger ◽  
Yin Yang 1 ◽  
Finger Protein ◽  
Yin Yang

scholarly journals The Co-Chaperone HspBP1 Is a Novel Component of Stress Granules that Regulates Their Formation

Cells ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 825
Author(s):  
Hicham Mahboubi ◽  
Ossama Moujaber ◽  
Mohamed Kodiha ◽  
Ursula Stochaj
Keyword(s):  
Stress Response ◽  
Cellular Stress ◽  
Stress Granules ◽  
Biological Properties ◽  
Cellular Stress Response ◽  
Binding Domains ◽  
Hsp70 Family ◽  
Independent Functions

The co-chaperone HspBP1 interacts with members of the hsp70 family, but also provides chaperone-independent functions. We report here novel biological properties of HspBP1 that are relevant to the formation of cytoplasmic stress granules (SGs). SG assembly is a conserved reaction to environmental or pathological insults and part of the cellular stress response. Our study reveals that HspBP1 (1) is an integral SG constituent, and (2) a regulator of SG assembly. Oxidative stress relocates HspBP1 to SGs, where it co-localizes with granule marker proteins and polyA-RNA. Mass spectrometry and co-immunoprecipitation identified novel HspBP1-binding partners that are critical for SG biology. Specifically, HspBP1 associates with the SG proteins G3BP1, HuR and TIA-1/TIAR. HspBP1 also interacts with polyA-RNA in vivo and binds directly RNA homopolymers in vitro. Multiple lines of evidence and single-granule analyses demonstrate that HspBP1 is crucial for SG biogenesis. Thus, HspBP1 knockdown interferes with stress-induced SG assembly. By contrast, HspBP1 overexpression promotes SG formation in the absence of stress. Notably, the hsp70-binding domains of HspBP1 regulate SG production in unstressed cells. Taken together, we identified novel HspBP1 activities that control SG formation. These features expand HspBP1’s role in the cellular stress response and provide new mechanistic insights into SG biogenesis.

Multispecialty Microsurgical Course Utilizing the Blue-Blood Chicken Thigh Model Significantly Improves Resident Comfort, Confidence, and Attitudes in Multiple Domains

2019 ◽  
Vol 36 (02) ◽  
pp. 142-150 ◽  
Author(s):  
Nikita O. Shulzhenko ◽  
Weifeng Zeng ◽  
Nicholas J. Albano ◽  
Sarah M. Lyon ◽  
Aaron M. Wieland ◽  
...  
Keyword(s):  
Technical Skill ◽  
Educational Models ◽  
Training Course ◽  
Ethical Implications ◽  
End To End ◽  
Case Presentations ◽  
High Level ◽  
Multiple Domains

Abstract Background The high level of technical skill required by microsurgical procedures has prompted the development of in vitro educational models. Current models are cost-ineffective, unrealistic, or carry ethical implications and are utilized as isolated experiences within single surgical specialties. The purpose of this study was to assess the educational and interprofessional effect of a microsurgical training course utilizing the nonliving “Blue-Blood” chicken thigh model (BBCTM) in a multidisciplinary environment. Methods A 10-hour course was developed integrating didactic lectures, case presentations, and one-on-one practical sessions utilizing hydrogel microvessels and the BBCTM. Pre- and postcourse surveys were administered assessing participants' self-reported comfort and confidence within fundamental microsurgical domains, assessments of the models utilized, and the effects of a multidisciplinary environment on the experience. Results A total of 19 residents attended the course on two separate occasions (n = 10 and n = 9, respectively). Respondents varied from postgraduate year-2 (PGY-2) to PGY-6+ and represented plastic and reconstructive surgery (n = 10), urology (n = 6), and otolaryngology (n = 3). On average, each participant performed 4.3 end-to-end, 1.3 end-to-side, and 0.4 coupler-assisted anastomoses. Following the course, participants felt significantly more comfortable operating a microscope and handling microsurgical instruments. They felt significantly more confident handling tissues, manipulating needles, microdissecting, performing end-to-end anastomoses, performing end-to-side anastomoses, using an anastomotic coupler, and declaring anastomoses suitable (all p < 0.05). The majority of participants believed that the use of live animals in the course would have minimally improved their learning. All but two respondents believed the course improved their awareness of the value of microsurgery in other specialties “very much” or “incredibly.” Conclusion A microsurgical training course utilizing nonliving models such as the “BBCTM significantly improves resident comfort and confidence in core operative domains and offers an in vivo experience without the use of live animals. Multispecialty training experiences in microsurgery are beneficial, desired, and likely underutilized.

scholarly journals Regulation of the DNA-binding and transcriptional activities of Xenopus laevis NFI-X by a novel C-terminal domain.

1995 ◽  
Vol 15 (10) ◽  
pp. 5552-5562 ◽  
Author(s):  
E Roulet ◽  
M T Armentero ◽  
G Krey ◽  
B Corthésy ◽  
C Dreyer ◽  
...  
Keyword(s):  
Dna Replication ◽  
Dna Binding ◽  
Xenopus Laevis ◽  
Transcriptional Activation ◽  
Binding Activity ◽  
New Members ◽  
Binding Domains ◽  
Dna Binding Activity

The nuclear factor I (NFI) family consists of sequence-specific DNA-binding proteins that activate both transcription and adenovirus DNA replication. We have characterized three new members of the NFI family that belong to the Xenopus laevis NFI-X subtype and differ in their C-termini. We show that these polypeptides can activate transcription in HeLa and Drosophila Schneider line 2 cells, using an activation domain that is subdivided into adjacent variable and subtype-specific domains each having independent activation properties in chimeric proteins. Together, these two domains constitute the full NFI-X transactivation potential. In addition, we find that the X. laevis NFI-X proteins are capable of activating adenovirus DNA replication through their conserved N-terminal DNA-binding domains. Surprisingly, their in vitro DNA-binding activities are specifically inhibited by a novel repressor domain contained within the C-terminal part, while the dimerization and replication functions per se are not affected. However, inhibition of DNA-binding activity in vitro is relieved within the cell, as transcriptional activation occurs irrespective of the presence of the repressor domain. Moreover, the region comprising the repressor domain participates in transactivation. Mechanisms that may allow the relief of DNA-binding inhibition in vivo and trigger transcriptional activation are discussed.

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