Developing MYC Degraders Bearing the Von Hippel-Lindau Ligand to Target the "Undruggable" MYC.

Although small-molecule inhibitors with moderate efficacy targeting MYC have been previously described, to this point, research efforts have failed to bring a suitable small-molecule MYC inhibitor to the clinic. Herein, the discovery of a series of novel MYC degraders bearing VHL to target the "undruggable" MYC is presented. The molecules are based on connecting a known MYC binder to a VHL ligand or pomalidomide to induce MYC degradation in various cancer cells known to express MYC.

Distinct roles of haspin in stem cell division and male gametogenesis.

The kinase haspin phosphorylates histone H3 at threonine-3 (H3T3ph) during mitosis. H3T3ph provides a docking site for the Chromosomal Passenger Complex at the centromere, enabling correction of erratic microtubule-chromosome contacts. Although this mechanism is operational in all dividing cells, haspin-null mice do not exhibit developmental anomalies, apart from aberrant testis architecture.

An "Onion-like" Model of Protein Unfolding: Collective versus Site Specific Approaches.

Approximating protein unfolding by an all-or-none cooperative event is a convenient assumption that can provide precious global information on protein stability. It is however quickly emerging that the scenario is far more complex and that global denaturation curves often hide a rich heterogeneity of states that are largely probe dependent. In this review, we revisit the importance of gaining site-specific information on the unfolding process.

Progressive Phosphorylation Modulates the Self-Association of a Variably Modified Histone H3 Peptide.

Protein phosphorylation is a key regulatory mechanism in eukaryotic cells. In the intrinsically disordered histone tails, phosphorylation is often a part of combinatorial post-translational modifications and an integral part of the "histone code" that regulates gene expression. Here, we study the association between two histone H3 tail peptides modified to different degrees, using fully atomistic molecular dynamics simulations.

Haspin-dependent and independent effects of the kinase inhibitor 5-Iodotubercidin on self-renewal and differentiation.

The kinase Haspin phosphorylates histone H3 at threonine-3 (H3T3ph), creating a docking site for the Chromosomal Passenger Complex (CPC). CPC plays a pivotal role in preventing chromosome misalignment. Here, we have examined the effects of 5-Iodotubercidin (5-ITu), a commonly used Haspin inhibitor, on self-renewal and differentiation of mouse embryonic stem cells (ESCs).

Heterochromatin remodeling in embryonic stem cells proceeds through stochastic de-stabilization of regional steady-states.

Cell differentiation is associated with progressive immobilization of chromatin proteins, expansion of heterochromatin, decrease of global transcriptional activity and induction of lineage-specific genes. However, how these processes relate to one another remains unknown. We show here that the heterochromatic domains of mouse embryonic stem cells (ESCs) are dynamically distinct and possesses a mosaic sub-structure.

The Leishmania donovani histidine acid ecto-phosphatase LdMAcP: insight into its structure and function.

Acid ecto-phosphatase activity has been implicated in Leishmania donovani promastigote virulence. In the present study, we report data contributing to the molecular/structural and functional characterization of the L. donovani LdMAcP (L.

Revisiting a dogma: the effect of volume exclusion in molecular crowding.

We critically re-examined the mechanisms that affect protein stability under crowding conditions. Much attention has been paid in recent years to the effects of molecular crowding on the properties of proteins in the cellular milieu. It has been suggested that volume exclusion disfavors the unfolded state since this is more expanded than the folded one, leading to an overall stabilizing effect.

POTAMOS mass spectrometry calculator: computer aided mass spectrometry to the post-translational modifications of proteins. A focus on histones.

Mass spectrometry is a widely used technique for protein identification and it has also become the method of choice in order to detect and characterize the post-translational modifications (PTMs) of proteins. Many software tools have been developed to deal with this complication. In this paper we introduce a new, free and user friendly online software tool, named POTAMOS Mass Spectrometry Calculator, which was developed in the open source application framework Ruby on Rails.

The effect of crowding and confinement: a comparison of Yfh1 stability in different environments.

Crowding and confinement can affect protein stability, favouring the more compact species amongst the folded and unfolded conformations. An unbiased assessment of the relative efficacy of crowded and confined environments has been hampered so far by the paucity of homogeneous comparisons on the same protein. This paper reports spectroscopic studies on yeast frataxin (Yfh1), a protein which provides an excellent model system for stability studies since it undergoes both cold and heat denaturation at measurable temperatures.

Structural role of RKS motifs in chromatin interactions: a molecular dynamics study of HP1 bound to a variably modified histone tail.

The current understanding of epigenetic signaling assigns a central role to post-translational modifications that occur in the histone tails. In this context, it has been proposed that methylation of K9 and phosphorylation of S10 in the tail of histone H3 represent a binary switch that controls its reversible association to heterochromatin protein 1 (HP1). To test this hypothesis, we performed a comprehensive molecular dynamics study in which we analyzed a crystallographically defined complex that involves the HP1 chromodomain and an H3 tail peptide.

Solution structure and molecular interactions of lamin B receptor Tudor domain.

Lamin B receptor (LBR) is a polytopic protein of the nuclear envelope thought to connect the inner nuclear membrane with the underlying nuclear lamina and peripheral heterochromatin. To better understand the function of this protein, we have examined in detail its nucleoplasmic region, which is predicted to harbor a Tudor domain (LBR-TD). Structural analysis by multidimensional NMR spectroscopy establishes that LBR-TD indeed adopts a classical β-barrel Tudor fold in solution, which, however, features an incomplete aromatic cage.

ERBIN is a new SARA-interacting protein: competition between SARA and SMAD2 and SMAD3 for binding to ERBIN.

SARA, an early endosomal protein, plays a key role in TGFβ signalling, as it presents SMAD2 and SMAD3 for phosphorylation by the activated TGFβ receptors. Here, we show that ERBIN is a new SARA-interacting protein that can be recruited by SARA to early endosomes. ERBIN was recently shown to bind and segregate phosphorylated SMAD2 and SMAD3 (SMAD2/3) in the cytoplasm, thereby inhibiting SMAD2/3-dependent transcription.

Casein kinase 1 regulates human hypoxia-inducible factor HIF-1.

Hypoxia-inducible factor 1 (HIF-1), a transcriptional activator that mediates cellular response to hypoxia and a promising target of anticancer therapy, is essential for adaptation to low oxygen conditions, embryogenesis and tumor progression. HIF-1 is a heterodimer of HIF-1alpha, expression of which is controlled by oxygen levels as well as by various oxygen-independent mechanisms, and HIF-1beta (or ARNT), which is constitutively expressed. In this work, we investigate the phosphorylation of the N-terminal heterodimerization (PAS) domain of HIF-1alpha and identify Ser247 as a major site of in vitro modification by casein kinase 1delta (CK1delta).

Cold denaturation and aggregation: a comparative NMR study of titin I28 in bulk and in a confined environment.

An NMR study of the thermal stability of titin I28 in the temperature range from -16 to 65 degrees C showed that this protein can undergo cold denaturation at physiological conditions. This is the second case of a protein undergoing unbiased cold denaturation. Comparison of the stability curves in buffer and in crowded conditions shows that it is possible to measure thermodynamics parameters for unfolding even when proteins aggregate at high temperature.

Phosphorylation of histone H3 at Thr3 is part of a combinatorial pattern that marks and configures mitotic chromatin.

We have previously shown that histone H3 is transiently phosphorylated at Thr3 during mitosis. Extending these studies, we now report that phosphorylated Thr3 is always in cis to trimethylated Lys4 and dimethylated Arg8, forming a new type of combinatorial modification, which we have termed PMM. PMM-marked chromatin emerges at multiple, peripheral sites of the prophase nucleus, then forms distinct clusters at the centric regions of metaphase chromosomes, and finally spreads (as it wanes) to the distal areas of segregating chromatids.

Identification of distinct SET/TAF-Ibeta domains required for core histone binding and quantitative characterisation of the interaction.

Background: The assembly of nucleosomes to higher-order chromatin structures is finely tuned by the relative affinities of histones for chaperones and nucleosomal binding sites. The myeloid leukaemia protein SET/TAF-Ibeta belongs to the NAP1 family of histone chaperones and participates in several chromatin-based mechanisms, such as chromatin assembly, nucleosome reorganisation and transcriptional activation. To better understand the histone chaperone function of SET/TAF-Ibeta, we designed several SET/TAF-Ibeta truncations, examined their structural integrity by circular Dichroism and assessed qualitatively and quantitatively the histone binding properties of wild-type protein and mutant forms using GST-pull down experiments and fluorescence spectroscopy-based binding assays.

Chromatin remodeling during mitosis: a structure-based code?

Histone modifications have been associated with particular states of transcriptional activity and are thought to serve as an "information code". However, this principle does not apply to histone phosphorylation, which can be detected in two, seemingly contrasting situations, i.e.

An automated method for gridding in microarray images.

Microarray technology is a powerful tool for analyzing the expression of a large number of genes in parallel. A typical microarray image consists of a few thousands of spots which determine the level of gene expression in the sample. In this paper we propose a method which automatically addresses each spot area in the image.

Parathymosin affects the binding of linker histone H1 to nucleosomes and remodels chromatin structure.

Linker histone H1 is the major factor that stabilizes higher order chromatin structure and modulates the action of chromatin-remodeling enzymes. We have previously shown that parathymosin, an acidic, nuclear protein binds to histone H1 in vitro and in vivo. Confocal laser scanning microscopy reveals a nuclear punctuate staining of the endogenous protein in interphase cells, which is excluded from dense heterochromatic regions.

The factors governing the thermal stability of frataxin orthologues: how to increase a protein's stability.

Understanding the factors governing the thermal stability of proteins and correlating them to the sequence and structure is a complex and multiple problem that can nevertheless provide important information on the molecular forces involved in protein folding. Here, we have carried out a comparative genomic study to analyze the effects that different intrinsic and environmental factors have on the thermal stability of frataxins, a family of small mitochondrial iron-binding proteins found in organisms ranging from bacteria to humans. Low expression of frataxin in humans causes Friedreich's ataxia, an autosomal recessive neurodegenerative disease.

Human recombinant mutated forms of the mitochondrial COX assembly Sco2 protein differ from wild-type in physical state and copper binding capacity.

The human Sco2 protein is a cytochrome c oxidase assembly protein that participates in mitochondrial copper pathway, acting downstream of Cox17 protein. In a previous work, we detected mutations in the human SCO2 gene in three unrelated infants with fatal cardioencephalomyopathy and COX deficiency. In this study, full-length processed recombinant wild-type and two mutated forms of hSco2p (w/t-rhSco2p, E140K-rhSco2p, and S225F-rhSco2p) were produced in bacteria as soluble recombinant peptides for the first time and evaluated for differences in their physical state and ability to bind copper.

Protein stability in nanocages: a novel approach for influencing protein stability by molecular confinement.

Confinement of a protein in a small inert space and microviscosity are known to increase its thermodynamic stability in a way similar to the mechanisms that stabilize protein fold in the cell. Here, to examine the influence of confinement on protein stability we choose four test cases of single domain proteins characterized by a wide range of melting temperatures, from approximately 73 degrees C of titin I27 to approximately 36 degrees C of yeast frataxin. All proteins are stabilized when confined in the gel, the most dramatic stabilization being that of yeast frataxin, whose melting temperature increased by almost 5 degrees C in the gel.

Type III protein translocase: HrcN is a peripheral ATPase that is activated by oligomerization.

Type III protein secretion (TTS) is catalyzed by translocases that span both membranes of Gram-negative bacteria. A hydrophilic TTS component homologous to F1/V1-ATPases is ubiquitous and essential for secretion. We show that hrcN encodes the putative TTS ATPase of Pseudomonas syringae pathovar phaseolicola and that HrcN is a peripheral protein that assembles in clusters at the membrane.