DNA compaction by the higher-order assembly of PRH/Hex homeodomain protein oligomers

Soufi, Abdenour; Sawasdichai, Anyaporn; Shukla, Alok Kumar; Noy, Peter J.; Dafforn, Timothy R.; Smith, Corinne J.; Jayaraman, Padma-Sheela; Gaston, Kevin

doi:10.1093/nar/gkq659

Cited by 5 publications

(18 citation statements)

References 41 publications

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“…However, rigid body association of two 8.6 nm radius spheres produces an apparent hydrodynamic radius of 17.2 and an apparent molecular weight of 2,618 kDa and the association of three, four or five spheres produces apparent molecular weights of 6,762 kDa, 13,256 kDa and 22,344 kDa, respectively. Species of this nature would be entirely consistent with previous data obtained using AUC and electron microscopy [6]. The largest species (∼500 nm) is not highly populated and it is not removed by centrifugation or filtration; it is therefore not a non-specific aggregate (data not shown).…”

Section: Resultssupporting

confidence: 90%

“…Rather, the resistance to denaturation shown by PRH suggests that this protein forms obligate oligomers in which there are large hydrophobic interacting surfaces, and possibly the exchange of secondary structure units or domain swaps, between monomers. This produces oligomers in which dissociation of the subunits and protein unfolding are coupled and it could explain why we have not observed PRH assemblies smaller than octamers in AUC or gel filtration chromatography [6], [15]. In contrast, in more dynamic assemblies, the subunits usually interact and fold independently with the result that oligomer dissociation does not required unfolding of the monomers and there can be rapid subunit exchange.…”

Section: Discussionmentioning

confidence: 91%

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The Proline Rich Homeodomain Protein PRH/Hhex Forms Stable Oligomers That Are Highly Resistant to Denaturation

et al. 2012

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BackgroundMany transcription factors control gene expression by binding to specific DNA sequences at or near the genes that they regulate. However, some transcription factors play more global roles in the control of gene expression by altering the architecture of sections of chromatin or even the whole genome. The ability to form oligomeric protein assemblies allows many of these proteins to manipulate extensive segments of DNA or chromatin via the formation of structures such as DNA loops or protein-DNA fibres.Principal FindingsHere we show that the proline rich homeodomain protein PRH/Hhex forms predominantly octameric and/or hexadecameric species in solution as well as larger assemblies. We show that these assemblies are highly stable resisting denaturation by temperature and chemical denaturants.ConclusionThese data indicate that PRH is functionally and structurally related to the Lrp/AsnC family of proteins, a group of proteins that are known to act globally to control gene expression in bacteria and archaea.

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Section: Resultssupporting

confidence: 90%

Section: Discussionmentioning

confidence: 91%

The Proline Rich Homeodomain Protein PRH/Hhex Forms Stable Oligomers That Are Highly Resistant to Denaturation

et al. 2012

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“…Moreover, mechanically compacted DNA has recently been used as nanostructure template4 and also applied as protection against chemical, biochemical and mechanical stresses5. Learning from the nature, researchers have started applying the compaction tricks using polyamines67, surfactants89, liposomes10, nanoparticles1112, polymer13, osmoticants like polyethylene glycol (PEG)14, dendrimers1516, multivalent ions17, metal complex18, cyclodextrin19, peptides20 and proteins21 for compaction and storage of DNA for prolonged duration. Among these compacting agents, most important naturally occurring DNA compaction agents are proteins and polyamines like of spermine and spermidine etc2021.…”

mentioning

confidence: 99%

“…Learning from the nature, researchers have started applying the compaction tricks using polyamines67, surfactants89, liposomes10, nanoparticles1112, polymer13, osmoticants like polyethylene glycol (PEG)14, dendrimers1516, multivalent ions17, metal complex18, cyclodextrin19, peptides20 and proteins21 for compaction and storage of DNA for prolonged duration. Among these compacting agents, most important naturally occurring DNA compaction agents are proteins and polyamines like of spermine and spermidine etc2021. Under physiological pH (~7.4), polyamines generally exists in positively charged form, which participates in strong electrostatic interaction with negatively charged phosphate backbone of DNA, and thereby results compaction and enhances the thermal stability of DNA322.…”

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confidence: 99%

A Green Solvent Induced DNA Package

Satpathi

Sengupta

Hridya

et al. 2015

Sci Rep

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Mechanistic details of DNA compaction is essential blue print for gene regulation in living organisms. Many in vitro studies have been implemented using several compaction agents. However, these compacting agents may have some kinds of cytotoxic effects to the cells. To minimize this aspect, several research works had been performed, but people have never focused green solvent, i.e. room temperature ionic liquid as DNA compaction agent. To the best of our knowledge, this is the first ever report where we have shown that guanidinium tris(pentafluoroethyl)trifluorophosphate (Gua-IL) acts as a DNA compacting agent. The compaction ability of Gua-IL has been verified by different spectroscopic techniques, like steady state emission, circular dichroism, dynamic light scattering and UV melting. Notably, we have extensively probed this compaction by Gua-IL through field emission scanning electron microscopy (FE-SEM) and fluorescence microscopy images. We also have discussed the plausible compaction mechanism process of DNA by Gua-IL. Our results suggest that Gua-IL forms a micellar kind of self aggregation above a certain concentration (≥1 mM), which instigates this compaction process. This study divulges the specific details of DNA compaction mechanism by a new class of compaction agent, which is highly biodegradable and eco friendly in nature.

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“…An amount of 20 µg of cell extract was used in western blotting experiments with PRH and phosphorylated PRH (pPRH) antibodies (16). …”

Section: Methodsmentioning

confidence: 99%

Protein kinase CK2 inactivates PRH/Hhex using multiple mechanisms to de-repress VEGF-signalling genes and promote cell survival

Noy

Sawasdichai

Jayaraman

et al. 2012

Nucleic Acids Research

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Protein kinase CK2 promotes cell survival and the activity of this kinase is elevated in several cancers including chronic myeloid leukaemia. We have shown previously that phosphorylation of the Proline-Rich Homeodomain protein (PRH/Hhex) by CK2 inhibits the DNA-binding activity of this transcription factor. Furthermore, PRH represses the transcription of multiple genes encoding components of the VEGF-signalling pathway and thereby influences cell survival. Here we show that the inhibitory effects of PRH on cell proliferation are abrogated by CK2 and that CK2 inhibits the binding of PRH at the Vegfr-1 promoter. Phosphorylation of PRH by CK2 also decreases the nuclear association of PRH and induces its cleavage by the proteasome. Moreover, cleavage of phosphorylated PRH produces a stable truncated cleavage product which we have termed PRHΔC (HhexΔC). PRHΔC acts as a transdominant negative regulator of full-length PRH by sequestering TLE proteins that function as PRH co-repressors. We show that this novel regulatory mechanism results in the alleviation of PRH-mediated repression of Vegfr-1. We suggest that the re-establishment of PRH function through inhibition of CK2 could be of value in treatment of myeloid leukaemias, as well as other tumour types in which PRH is inactivated by phosphorylation.

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DNA compaction by the higher-order assembly of PRH/Hex homeodomain protein oligomers

Cited by 5 publications

References 41 publications

The Proline Rich Homeodomain Protein PRH/Hhex Forms Stable Oligomers That Are Highly Resistant to Denaturation

The Proline Rich Homeodomain Protein PRH/Hhex Forms Stable Oligomers That Are Highly Resistant to Denaturation

A Green Solvent Induced DNA Package

Protein kinase CK2 inactivates PRH/Hhex using multiple mechanisms to de-repress VEGF-signalling genes and promote cell survival

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