Experimental characterization of the human non-sequence-specific nucleic acid interactome.

Background: The interactions between proteins and nucleic acids have a fundamental function in many biological processes, including gene transcription, RNA homeostasis, protein translation and pathogen sensing for innate immunity. While our knowledge of the ensemble of proteins that bind individual mRNAs in mammalian cells has been greatly augmented by recent surveys, no systematic study on the non-sequence-specific engagement of native human proteins with various types of nucleic acids has been reported.

Results: We designed an experimental approach to achieve broad coverage of the non-sequence-specific RNA and DNA binding space, including methylated cytosine, and tested for interaction potential with the human proteome.

BCR-ABL uncouples canonical JAK2-STAT5 signaling in chronic myeloid leukemia.

Constitutive activation of STAT5 is critical for the maintenance of chronic myeloid leukemia (CML) characterized by the BCR-ABL oncoprotein. Tyrosine kinase inhibitors (TKIs) for the STAT5-activating kinase JAK2 have been discussed as a treatment option for CML patients. Using murine leukemia models combined with inducible ablation of JAK2, we show JAK2 dependence for initial lymphoid transformation, which is lost once leukemia is established.

Targeting the SH2-kinase interface in Bcr-Abl inhibits leukemogenesis.

Chronic myelogenous leukemia (CML) is caused by the constitutively active tyrosine kinase Bcr-Abl and treated with the tyrosine kinase inhibitor (TKI) imatinib. However, emerging TKI resistance prevents complete cure. Therefore, alternative strategies targeting regulatory modules of Bcr-Abl in addition to the kinase active site are strongly desirable.

Initial characterization of the human central proteome.

Background: On the basis of large proteomics datasets measured from seven human cell lines we consider their intersection as an approximation of the human central proteome, which is the set of proteins ubiquitously expressed in all human cells. Composition and properties of the central proteome are investigated through bioinformatics analyses.

Results: We experimentally identify a central proteome comprising 1,124 proteins that are ubiquitously and abundantly expressed in human cells using state of the art mass spectrometry and protein identification bioinformatics.

BCR-ABL SH3-SH2 domain mutations in chronic myeloid leukemia patients on imatinib.

Point mutations in the kinase domain of BCR-ABL are the most common mechanism of drug resistance in chronic myeloid leukemia (CML) patients treated with ABL kinase inhibitors, including imatinib. It has also been shown in vitro that mutations outside the kinase domain in the neighboring linker, SH2, SH3, and Cap domains can confer imatinib resistance. In the context of ABL, these domains have an autoinhibitory effect on kinase activity, and mutations in this region can activate the enzyme.

A potent and highly specific FN3 monobody inhibitor of the Abl SH2 domain.

Interactions between Src homology 2 (SH2) domains and phosphotyrosine sites regulate tyrosine kinase signaling networks. Selective perturbation of these interactions is challenging due to the high homology among the 120 human SH2 domains. Using an improved phage-display selection system, we generated a small antibody mimic (or 'monobody'), termed HA4, that bound to the Abelson (Abl) kinase SH2 domain with low nanomolar affinity.

The chemokine interleukin-8 and the surface activation protein CD69 are markers for Bcr-Abl activity in chronic myeloid leukemia.

We have identified differentially regulated genes in chronic myeloid leukemia (CML) cells upon short treatment with the broad-spectrum Bcr-Abl inhibitor dasatinib. The highly specific Bcr-Abl inhibitor nilotinib caused a very similar gene expression signature, validating the identified differentially regulated genes as a read-out of Bcr-Abl activity and implying that Bcr-Abl is the functionally central target of dasatinib in CML cells. Among the strongest downregulated genes, we have further validated the activation marker CD69 and the chemokine interleukin (IL)-8.

Charting the molecular network of the drug target Bcr-Abl.

The tyrosine kinase Bcr-Abl causes chronic myeloid leukemia and is the cognate target of tyrosine kinase inhibitors like imatinib. We have charted the protein-protein interaction network of Bcr-Abl by a 2-pronged approach. Using a monoclonal antibody we have first purified endogenous Bcr-Abl protein complexes from the CML K562 cell line and characterized the set of most tightly-associated interactors by MS.

Chemical proteomic profiles of the BCR-ABL inhibitors imatinib, nilotinib, and dasatinib reveal novel kinase and nonkinase targets.

The BCR-ABL tyrosine kinase inhibitor imatinib represents the current frontline therapy in chronic myeloid leukemia. Because many patients develop imatinib resistance, 2 second-generation drugs, nilotinib and dasatinib, displaying increased potency against BCR-ABL were developed. To predict potential side effects and novel medical uses, we generated comprehensive drug-protein interaction profiles by chemical proteomics for all 3 drugs.