202 The Reduced and Altered Activities of PAX5 are Linked to the Protein-Protein Interaction Motif (Coiled-Coil Domain) of the PAX5-PML Fusion Protein in t(9;15)-Associated Acute Lymphocytic Leukemia

2011 ◽  
Vol 11 ◽  
pp. S124
Author(s):  
Jihui Qiu ◽  
Huimin Chu ◽  
Shuo Dong
Keyword(s):  
Fusion Protein ◽  
Protein Interaction ◽  
Acute Lymphocytic Leukemia ◽  
Coiled Coil ◽  
Lymphocytic Leukemia ◽  
Protein Protein Interaction ◽  
Coiled Coil Domain

The Reduced and Altered Activities of PAX5 Are Linked to the Protein-Protein Interaction Motif (coiled-coil domain) of the PAX5-PML Fusion Protein In t(9;15)-Associated Acute Lymphocytic Leukemia.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1022-1022 ◽  
Author(s):  
Jihui Qiu ◽  
Huimin Chu ◽  
Shuo Dong
Keyword(s):  
Fusion Protein ◽  
Protein Interaction ◽  
Conflicts Of Interest ◽  
Lymphoblastic Leukemia ◽  
Consensus Sequence ◽  
Coiled Coil ◽  
Lymphocytic Leukemia ◽  
Wild Type ◽  
Hematopoietic Stem ◽  
Coiled Coil Domain

Abstract Abstract 1022 The paired box domain of PAX5 was reported to fuse with the sequence of promylocytic leukemia (PML) to produce PAX5-PML chimeric protein in two patients with B-cell acute lymphoblastic leukemia (B-ALL). In the present studies, we found, by gel-shift assays, that PAX5-PML bound to a panel of PAX5-consensus sequence as a homodimer with reduction of its DNA-binding affinities in comparison with wild-type PAX5. In transient transfection assays using 293T and HeLa cells, and retrovirus transduction of murine hematopoietic stem/progenitor cells together with quantitative real-time polymerase chain reaction (RQ-PCR) analysis, PAX5-PML inhibited wild-type PAX5 target gene transcriptional activity. Studies comparing PAX5-PML with PAX5-PML(ΔCC) demonstrated that the coiled-coil protein interaction domain located within PML moiety was required for PAX5-PML homodimer complex formation and partial transcriptional repression of genes controlled by PAX5. Fluorescent microscopic examination of transiently expressed YFP-tagged proteins in HeLa and 293T cells demonstrated that YFP-PAX5-PML and YFP-PAX5-PML(ΔCC) exhibited a diffuse granular pattern within the nucleus, similar to PAX5 but PML. By fluorescent recovery after photobleach (FRAP), we have shown that PAX5-PML fusion protein has reduced intranuclear mobility compared to wild-type PAX5. Furthermore, the dimerization domain (coiled-coil) of PML was responsible for the reduced intranuclear mobility of PAX5-PML. These results indicate that the coiled-coil domain of PAX5-PML is important for each of the known activities of PAX5-PML fusion proteins. Disclosures: No relevant conflicts of interest to declare.

Characterization of Two New Imatinib-Responsive Fusion Genes Generated by Disruption of PDGFRB in Eosinophilia-Associated Chronic Myeloproliferative Disorders.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 667-667
Author(s):  
Christoph Walz ◽  
Georgia Metzgeroth ◽  
Claudia Schoch ◽  
Torsten Haferlach ◽  
Rudiger Hehlmann ◽  
...  
Keyword(s):  
Amino Acid ◽  
Fusion Protein ◽  
Tyrosine Kinase ◽  
Coiled Coil ◽  
Myeloproliferative Disorders ◽  
Fusion Genes ◽  
Protein Protein Interaction ◽  
Coiled Coil Domain ◽  
Exon 11

Abstract Fusion genes involving PDGFRA, PDGFRB, FGFR1 and JAK2 are seen in a substantial number of patients with BCR-ABL negative myeloproliferative disorders (MPD) and result in constitutive activation of the corresponding tyrosine kinase moiety. The vast majority of tyrosine kinase fusion partners contain coiled-coil domains or other dimerization motifs properties that are essential for malignant transformation. We have identified two patients presenting with eosinophilia-associated MPD and a t(5;12)(q31;q24) or a complex translocation t(1;5;11) with involvement of 5q31, respectively, suggesting a possible involvement of the PDGFRB gene which is located at chromosome band 5q31–33. 5′-rapid amplification of cDNA ends (5′-RACE) for the t(5;12) identified an in-frame mRNA fusion between ’G protein-coupled receptor kinase interactor 2′ (GIT2) exon 12 at 12q24 and PDGFRB exon 11. GIT2 is a member of the GIT protein family that is extensively alternative spliced in many distinct forms causing its functional diversity. A reciprocal transcript was amplified by RT-PCR with a fusion between PDGFRB exon 10 and GIT2 exon 13. GIT2-PDGFRB is predicted to be translated into a 742 amino acid fusion protein that retains the GIT2 N-terminal protein-protein interaction motif Ankyrin and an Arf GTPase activating protein (ArfGAP) domain fused to the transmembrane and catalytic domain of PDGFRB. The truncated GIT2 protein lacks coiled-coil domains as they are lost in the fusion protein due to the breakpoint within GIT2 intron 12. We therefore speculate that the Ankyrin repeat, which is one of the most common protein-protein interaction motifs in nature, may have replaced the function of a coiled-coil domain offering dimerization properties to the fusion protein. 5′-RACE for the complex t(1;5;11) identified an in-frame mRNA fusion between ’GPI-anchored membrane protein 1′ (GPIAP1) exon 7 at 11p13 and PDGFRB exon 11. Normal GPIAP1 is a cytoplasmic phosphoprotein which plays a mainly uncharacterized role in cellular activation or proliferation. The chimeric mRNA is predicted to encode an 803 amino acid fusion protein retaining the coiled-coil domain of GPIAP1 fused to the transmembrane and catalytic domains of PDGFRB. Both patients have been treated with 400 mg/day imatinib, which is a selective inhibitor of PDGFRB, and achieved rapid complete clinical and hematological remission. Residual GIT2-PDGFRB transcripts could be detected repeatedly during a 17 months follow up in case 1 whereas no follow-up samples have been available for case 2. These data give further evidence that numerous partner genes fuse to PDGFRB in BCR-ABL negative MPDs. In addition, the data demonstrate that cytogenetic analysis is a mandatory technique for the identification of tyrosine kinase fusion genes. In cases with abnormalities of chromosome 5q, a possible involvement of PDGFRB should be screened by adequate FISH and PCR-based techniques. Although their occurrence is rare in general, the identification of these fusion genes is essential for the successful treatment with tyrosine kinase inhibitors.

scholarly journals Correction: Solid-state nanopore analysis on conformation change of p53TAD–MDM2 fusion protein induced by protein–protein interaction

Nanoscale ◽  
2018 ◽  
Vol 10 (38) ◽  
pp. 18423-18423
Author(s):  
Hongsik Chae ◽  
Dong-Kyu Kwak ◽  
Mi-Kyung Lee ◽  
Seung-Wook Chi ◽  
Ki-Bum Kim
Keyword(s):  
Solid State ◽  
Fusion Protein ◽  
Protein Interaction ◽  
Conformation Change ◽  
Protein Protein Interaction

Correction for ‘Solid-state nanopore analysis on conformation change of p53TAD–MDM2 fusion protein induced by protein–protein interaction’ by Hongsik Chae et al., Nanoscale, 2018, DOI: 10.1039/c8nr06423g.

Development of Small Molecule Inhibitors of the AML1-ETO and CBFβ-SMMHC Oncoproteins.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1591-1591
Author(s):  
Jolanta E. Grembecka ◽  
Kristin Graf ◽  
Yali Kong ◽  
Michael Douvas ◽  
Tomasz Cierpicki ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Cell Line ◽  
Small Molecule ◽  
Protein Interaction ◽  
Small Molecule Inhibitors ◽  
Leukemia Cell Line ◽  
Runt Domain ◽  
Protein Protein Interaction ◽  
Lead Compounds ◽  
Binding Interface

Abstract Core binding factor (CBF) is a heterodimeric transcription factor composed of RUNX1 (CBFα) and CBFβ subunits which are essential for normal blood cell development. CBFβ functions to increase the DNA-binding of the RUNX1 subunit 20–40 fold and to protect the RUNX1 subunit against ubiqitination and proteasome degradation, making this protein-protein interaction critical for CBF function. Two of the most common translocations involving the subunits of CBF are the inv(16) and the t(8;21) which produce the chimeric proteins CBFβ-SMMHC and AML1-ETO, respectively, which are associated with the development of Acute Myeloid Leukemia (AML). The AML1-ETO fusion protein is a dominant inhibitor of wildtype RUNX1-CBFβ activity in vivo and causes a blockage in normal hematopoiesis, predisposing for the development of leukemia. The interaction between CBFβ and AML1-ETO is critical for its function, therefore treatments targeting AML1-ETO and blocking its interaction with CBFβ are highly likely to be therapeutically beneficial. The CBFβ-SMMHC fusion protein causes dysregulation of CBF function by means of anomalously tight binding to RUNX1. Since binding to RUNX1 is required for the dysfunction associated with CBFβ-SMMHC, this interaction represents an excellent target for inhibition as a potential therapeutic strategy. We have initiated efforts to develop small molecule inhibitors of the RUNX1-CBFβ interaction as possible therapeutics for the treatment of the associated leukemias. Both virtual screening searches, focused on the X-ray structures of RUNX1 Runt domain and CBFβ, and high-throughput screening of NCI (National Cancer Institute) and Maybridge fragment libraries were used to identify initial lead compounds interacting with these proteins and blocking heterodimerization of CBF. Compounds were tested experimentally by FRET (Fluorescence Resonance Energy Transfer) and ELISA for their inhibition of RUNX1-CBFβ interaction. This resulted in a number of initial lead compounds targeting either the Runt domain or CBFβ and inhibiting this protein-protein interaction. Based on the docking mode selected lead compounds were further optimized using medicinal chemistry approaches to increase their affinity and determine the structure-activity relationships (SAR). This resulted in several compounds with low micromolar affinity (IC50 < 10 μM) which effectively block the heterodimerization of CBF in vitro and in a cell-based assay. Interestingly, compounds targeting CBFβ bind to a site displaced from the binding interface for RUNX1 as shown by the NMR-based docking, i.e. these compounds function as allosteric inhibitors of this protein-protein interaction. The most potent compounds were tested either in the Kasumi-1 leukemia cell line harboring t(8;21) translocation or in the ME-1 cell line with inv(16), resulting in a blockage of proliferation, induction of apoptosis and differentiation of these cells. These compounds represent the first small molecule inhibitors targeting CBF and inhibiting this interaction. They represent good starting points for the development of therapeutically useful inhibitors. Several approaches are being explored to modify these compounds to achieve selectivity towards AML1-ETO or CBFβ-SMMHC oncoproteins versus wild type proteins.

Solid-state nanopore analysis on conformation change of p53TAD–MDM2 fusion protein induced by protein–protein interaction

Nanoscale ◽  
2018 ◽  
Vol 10 (36) ◽  
pp. 17227-17235 ◽  
Author(s):  
Hongsik Chae ◽  
Dong-Kyu Kwak ◽  
Mi-Kyung Lee ◽  
Seung-Wook Chi ◽  
Ki-Bum Kim
Keyword(s):  
Solid State ◽  
Fusion Protein ◽  
Single Molecule ◽  
Protein Interaction ◽  
Conformation Change ◽  
Protein Protein Interaction ◽  
Translocation Event

Nutlin-3-induced conformation change of p53TAD–MDM2 fusion protein were probed by single molecule translocation event using solid-state nanopores.

scholarly journals The features of polyglutamine regions depend on their evolutionary stability

2020 ◽  
Author(s):  
Pablo Mier ◽  
Miguel A. Andrade-Navarro
Keyword(s):  
Amino Acid ◽  
Codon Usage ◽  
Protein Interaction ◽  
Protein Interactions ◽  
Coiled Coil ◽  
Evolutionary Stability ◽  
Protein Protein Interactions ◽  
Protein Protein Interaction ◽  
Structural Conformation ◽  
Taxonomic Groups

Abstract Background Polyglutamine regions (polyQ) are one of the most studied and prevalent homorepeats in eukaryotes. They have a particular length-dependent codon usage, which relates to a characteristic CAG-slippage mechanism. Pathologically expanded tracts of polyQ are known to form aggregates and are involved in the development of several human neurodegenerative diseases. The non-pathogenic function of polyQ is to mediate protein-protein interactions via a coiled-coil pairing with an interactor. They are usually located in a helical context.Results Here we show how these known features related to polyQ depend on their stability in evolution. We have classified the polyQ regions of 60 proteomes from four distinct taxonomic groups (Insecta, Teleostei, Sauria and Mammalia) in three main categories based on their evolutionary stability. Codon usage, amino acid context, structural conformation and the protein-protein interaction capacity of polyQ from all studied taxa critically depend on the region stability.Conclusions Our results show that apart from the sequence of a polyQ, information about its orthologous sequences is needed to assess its function.

scholarly journals The features of polyglutamine regions depend on their evolutionary stability

2020 ◽  
Author(s):  
Pablo Mier ◽  
Miguel A. Andrade-Navarro
Keyword(s):  
Amino Acid ◽  
Codon Usage ◽  
Protein Interaction ◽  
Protein Interactions ◽  
Coiled Coil ◽  
Evolutionary Stability ◽  
Length Variation ◽  
Evolutionary Analysis ◽  
Protein Protein Interaction ◽  
Structural Conformation

Abstract Background: Polyglutamine regions (polyQ) are one of the most studied and prevalent homorepeats in eukaryotes. They have a particular length-dependent codon usage, which relates to a characteristic CAG-slippage mechanism. Pathologically expanded tracts of polyQ are known to form aggregates and are involved in the development of several human neurodegenerative diseases. The non-pathogenic function of polyQ is to mediate protein-protein interactions via a coiled-coil pairing with an interactor. They are usually located in a helical context. Results: Here we study the stability of polyQ regions in evolution, using a set of 60 proteomes from four distinct taxonomic groups (Insecta, Teleostei, Sauria and Mammalia). The polyQ regions can be distinctly grouped in three categories based on their evolutionary stability: stable, unstable by length variation (inserted), and unstable by mutations (mutated). PolyQ regions in these categories can be significantly distinguished by their glutamine codon usage, and we show that the CAG-slippage mechanism is predominant in inserted polyQ of Sauria and Mammalia. The polyQ amino acid context is also influenced by the polyQ stability, with a higher proportion of proline residues around inserted polyQ. By studying the secondary structure of the sequences surrounding polyQ regions, we found that regarding the structural conformation around a polyQ, its stability category is more relevant than its taxonomic information. The protein-protein interaction capacity of a polyQ is also affected by its stability, as stable polyQ have more interactors than unstable polyQ.Conclusions: Our results show that apart from the sequence of a polyQ, information about its orthologous sequences is needed to assess its function. Codon usage, amino acid context, structural conformation and the protein-protein interaction capacity of polyQ from all studied taxa critically depend on the region stability. There are however some taxa-specific polyQ features that override this importance. We conclude that a taxa-driven evolutionary analysis is of the highest importance for the comprehensive study of any feature of polyglutamine regions.

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