Eukaryote DNA is wrapped around nucleosomes that are comprised of octamers of core histones (two heterodimers of histones H2A and H2B and a heterotetramer of histones H3 and H4). Nucleosomes are joined by linker DNA and histone H1 to form chromatin. A central mechanism for regulating chromatin activity is the (reversible) covalent modification of histones by enzymes. A complex interplay between posttranslational modifications (PTMs) 1 either represses or activates transcription in a site-specific and cooperative manner (1). The NH 2 -and COOH-terminal tails of the core histones are subjected to multisite modifications by methylation, acetylation, phosphorylation, ADP-ribosylation, ubiquitination (2, 3), and sumoylation (4). A "histone code" model has been hypothesized where the cell uses a combinatorial system of (clustered) covalent modifications to regulate specific genomic functions (5-7). The current hypothesis is that the pattern and types of modifications modulate protein-protein and protein-DNA interactions, e.g. for recruitment of transcription factors, as a function of the cellular state and environment (8).Acetylation of lysine residues is a major mediator of the histone code. Hyperacetylation of histone tails is usually a sign of high transcriptional activity, whereas hypoacetylation indicates transcriptional silencing (9). Reversible acetylation/ deacetylation of lysine residues is a dynamic balance between the activities of histone acyltransferases and histone deacetylases (HDACs). Inhibition of HDAC activity results in hyperacetylation of lysine residues in the NH 2 tails of histones H3 and H4 (10), and it leads to the expression of genes that induce growth arrest, cell differentiation, and apoptotic cell death in cultured tumor cells (11,12). Non-toxic levels of several HDACis have shown promising pharmacological properties by inhibiting tumor growth in animal models (13)(14)(15). Consequently there is a great interest in developing HDACis as anticancer drugs, and some of these compounds are currently being evaluated in clinical trails (16,17).Mass spectrometry is rapidly becoming a key analytical technology in molecular cell biology and proteomics. Mass spectrometry is particularly suited for the examination of protein primary structure and for determining post-translational modifications of proteins as it reveals covalent modifications From the ‡Danish Technological Institute,

Activating mutations in the epidermal growth factor receptor gene occur as early cancer-driving clonal events in a subset of patients with non-small cell lung cancer (NSCLC) and result in increased sensitivity to EGFR-tyrosine-kinase-inhibitors (EGFR-TKIs). Despite very frequent and often prolonged clinical response to EGFR-TKIs, virtually all advanced EGFR-mutated (EGFRM+) NSCLCs inevitably acquire resistance mechanisms and progress at some point during treatment. Additionally, 20–30% of patients do not respond or respond for a very short time (<3 months) because of intrinsic resistance. While several mechanisms of acquired EGFR-TKI-resistance have been determined by analyzing tumor specimens obtained at disease progression, the factors causing intrinsic TKI-resistance are less understood. However, recent comprehensive molecular-pathological profiling of advanced EGFRM+ NSCLC at baseline has illustrated the co-existence of multiple genetic, phenotypic, and functional mechanisms that may contribute to tumor progression and cause intrinsic TKI-resistance. Several of these mechanisms have been further corroborated by preclinical experiments. Intrinsic resistance can be caused by mechanisms inherent in EGFR or by EGFR-independent processes, including genetic, phenotypic or functional tumor changes. This comprehensive review describes the identified mechanisms connected with intrinsic EGFR-TKI-resistance and differences and similarities with acquired resistance and among clinically implemented EGFR-TKIs of different generations. Additionally, the review highlights the need for extensive pre-treatment molecular profiling of advanced NSCLC for identifying inherently TKI-resistant cases and designing potential combinatorial targeted strategies to treat them.

To select the gene coding for an isoleucine permease, an isoleucine dependent strain (ilv1 cha1) was transformed with a yeast genomic multicopy library, and colonies growing at a low isoleucine concentration were selected. Partial sequencing of the responsible plasmid insert revealed the presence of a previously sequenced 609 codon open reading frame of chromosome II with homology to known permeases. Deletion, extra dosage and C-terminal truncation of this gene were constructed in a strain lacking the general amino acid permease, and amino acid uptake was measured during growth in synthetic complete medium. The following observations prompted us to name the gene BAP2 (branched-chain amino acid permease). Deletion of BAP2 reduced uptake of leucine, isoleucine and valine by 25-50%, while the uptake of 8 other L-alpha-amino acids was unaltered or slightly increased. Introduction of BAP2 on a centromere-based vector, leading to a gene dosage of two or slightly more, caused a 50% increase in leucine uptake and a smaller increase for isoleucine and valine. However, when the 29 C-terminal codons of the plasmid-borne copy of BAP2 were substituted, the cells more than doubled the uptake of leucine, isoleucine and valine, while no or little increase in uptake was observed for the other 8 amino acids.

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