This study explores the roles of genome copy number abnormalities (CNAs) in breast cancer pathophysiology by identifying associations between recurrent CNAs, gene expression, and clinical outcome in a set of aggressively treated early-stage breast tumors. It shows that the recurrent CNAs differ between tumor subtypes defined by expression pattern and that stratification of patients according to outcome can be improved by measuring both expression and copy number, especially high-level amplification. Sixty-six genes deregulated by the high-level amplifications are potential therapeutic targets. Nine of these (FGFR1, IKBKB, ERBB2, PROCC, ADAM9, FNTA, ACACA, PNMT, and NR1D1) are considered druggable. Low-level CNAs appear to contribute to cancer progression by altering RNA and cellular metabolism.

In this study we examine the molecular basis for the synergistic regulation of the minimal TCRa enhancer by multiple proteins. We find that reconstitution of TCRc~ enhancer function in nonlymphoid cells requires expression of the lymphoid-specific proteins LEF-1, Ets-1 and PEBP2a (CBFc~I, and a specific arrangement of their binding sites in the enhancer. We show that Ets-1 cooperates with PEBP2a to bind adjacent sites at one end of the enhancer, forming a ternary complex that is unstable by itself. Stable occupancy of the Ets-1-and PEBP2oL-binding sites in a DNase I protection assay was found to depend on both a specific helical phasing relationship with a nonadjacent ATF/CREB-binding site at the other end of the enhancer and on LEF-1. The HMG domain of LEF-1 was found previously to bend the DNA helix in the center of the TCRot enhancer. We now show that the HMG domain of the distantly related SRY protein, which also bends DNA, can partially replace LEF-1 in stimulating enhancer function in transfection assays. Taken together with the observation that Ets-1 and members of the ATF/CREB family have the potential to associate in vitro, these data suggest that LEF-1 can coordinate the assembly of a specific higher-order enhancer complex by facilitating interactions between proteins bound at nonadjacent sites.

Analysis of a T-cell antigen receptor (TCR) a promoter from a variable gene segment (V) revealed a critical GT box element which is also found in upstream regions of several Va genes, TCR enhancer, and regulatory elements of other genes. This element is necessary for TCR gene expression and binds several proteins. These GT box-binding proteins were identified as members of a novel Spl multigene family. Two regions (for reviews, see references 5 and 27). The process of V-D-J gene rearrangements is developmentally and tissue specifically regulated. The TCR a and 8 genes are particularly interesting because the 8 locus is located within the TCR a-gene segments. In T cells committed to the a,3 lineage, the a-specific V-gene segments are rearranged to the a locus, whereas in cells committed to the -yb lineage, the b-specific V-gene segments are rearranged to their respective D/Jb gene segments. Some V gene segments, however, can rearrange to either the a or 8 locus (these gene segments are also called Va).Studies using transgenic mice and cell lines suggest that specific gene segment transcription correlates with differential accessibility of each locus to the recombinase during development (8, 7). Production of sterile transcripts from unrearranged gene segments, in particular, has been found to precede gene rearrangement. It is hypothesized that sterile transcription is either the cause or the consequence of differential opening of chromatin structure and hence accessibility of the gene segment to the recombination enzymes (references 7 and 28 and references therein). Since each of the V-gene segments contains its own promoter, it is possible that the specificity of sterile transcription from these V promoters could contribute to differential accessibility to recombinases during T-cell development, which could result in specific gene segment usage in aot and/or 9y T cells (35). * Corresponding author.Therefore, the study of TCR a promoters can serve the dual purpose of analyzing TCR gene expression as well as regulation of rearrangement.We have focused on the well-characterized Vall.1 gene segment, which is the gene segment predominantly used in T-cell response to pigeon cytochrome c (9, 38). We have analyzed the critical elements for the Vall.1 promoter and found one element which we called the GT box. This GT box is also present in most other available Va upstream sequences (presumably promoters) as well as in the TCR ax enhancer. Homologous regulatory elements (AC boxes) were previously defined in the ,-globin enhancers, promoters, and locus-controlling region (LCR) as well as in the bovine papillomavirus (BPV) promoters and simian virus (SV40) enhancer. This novel element for T-cell gene expression was shown to bind ubiquitously expressed proteins, one of which was identified as the transcription factor Spl (18). Although Spl is one of the earliest-characterized transcription factors which also binds to a consensus GC box sequence, we have made the assumption that additional GT box-binding proteins contain se...

Transmembrane receptors for hormones, neurotransmitters, light, and odorants mediate their cellular effects by activating heterotrimeric guanine nucleotide-binding proteins (G proteins). Crystal structures have revealed contact surfaces between G protein subunits, but not the surfaces or molecular mechanism through which Galphabetagamma responds to activation by transmembrane receptors. Such a surface was identified from the results of testing 100 mutant alpha subunits of the retinal G protein transducin for their ability to interact with rhodopsin. Sites at which alanine substitutions impaired this interaction mapped to two distinct Galpha surfaces: a betagamma-binding surface and a putative receptor-interacting surface. On the basis of these results a mechanism for receptor-catalyzed exchange of guanosine diphosphate for guanosine triphosphate is proposed.