The TP53 Arg72Pro and MDM2 309G>T polymorphisms are not associated with breast cancer risk in BRCA1 and BRCA2 mutation carriers.

BACKGROUND: The TP53 pathway, in which TP53 and its negative regulator MDM2 are the central elements, has an important role in carcinogenesis, particularly in BRCA1- and BRCA2-mediated carcinogenesis. A single nucleotide polymorphism (SNP) in the promoter region of MDM2 (309T>G, rs2279744) and a coding SNP of TP53 (Arg72Pro, rs1042522) have been shown to be of functional significance. METHODS: To investigate whether these SNPs modify breast cancer risk for BRCA1 and BRCA2 mutation carriers, we pooled genotype data on the TP53 Arg72Pro SNP in 7011 mutation carriers and on the MDM2 309T>G SNP in 2222 mutation carriers from the Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA). Data were analysed using a Cox proportional hazards model within a retrospective likelihood framework. RESULTS: No association was found between these SNPs and breast cancer risk for BRCA1 (TP53: per-allele hazard ratio (HR)=1.01, 95% confidence interval (CI): 0.93-1.10, P(trend)=0.77; MDM2: HR=0.96, 95%CI: 0.84-1.09, P(trend)=0.54) or for BRCA2 mutation carriers (TP53: HR=0.99, 95%CI: 0.87-1.12, P(trend)=0.83; MDM2: HR=0.98, 95%CI: 0.80-1.21, P(trend)=0.88). We also evaluated the potential combined effects of both SNPs on breast cancer risk, however, none of their combined genotypes showed any evidence of association. CONCLUSION: There was no evidence that TP53 Arg72Pro or MDM2 309T>G, either singly or in combination, influence breast cancer risk in BRCA1 or BRCA2 mutation carriers.

Effect of the position of TAR on transcriptional activation by HIV-1 Tat in vivo.

Efficient expression of the human immunodeficiency virus (HIV) genome requires the viral-encoded transactivator Tat. Tat interacts with the highly structured trans-activation-response (TAR) RNA that is found at the 5' end of all viral transcripts, and mediates the formation of transcription complexes that are capable of elongation through the entire length of the viral genome. By placing TAR immediately downstream from the P2 promoter of the mouse c-myc gene, we have previously shown that Tat can also direct transcriptional elongation through potential sites of premature termination within c-myc in transfected HeLa cells. We now demonstrate that Tat can activate c-myc transcription when TAR is positioned internally within the c-myc transcript at distances up to 353 nt downstream from the P2 promoter. We show that Tat can also activate transcription from the c-myc P1 promoter, which is located 165 nt upstream from P2 in these hybrid gene constructs. These novel findings show that Tat can activate transcription in vivo when TAR is positioned at distances up to 518 nt downstream from the site of transcriptional initiation. The ability of TAR to mediate Tat-activated transcription over distances greater than previously appreciated has important implications for the mechanism of action of Tat.

Single-nucleotide polymorphisms in the RB1 gene and association with breast cancer in the British population.

A substantial proportion of the familial risk of breast cancer may be attributable to genetic variants each contributing a small effect. pRb controls the cell cycle and polymorphisms within it are candidates for such low penetrance susceptibility alleles, since the gene has been implicated in several human tumours, particularly breast cancer. The purpose of this study was to determine whether common variants in the RB1 gene are associated with breast cancer risk. We assessed 15 tagging single-nucleotide polymorphisms (SNPs) using a case-control study design (n< or = 4474 cases and n < or = 4560 controls). A difference in genotype frequencies was found between cases and controls for rs2854344 in intron 17 (P-trend = 0.007) and rs198580 in intron 19 (P-trend = 0.018). Carrying the minor allele of these SNPs appears to confer a protective effect on breast cancer risk (odd ratio (OR) = 0.86 (0.76-0.96) for rs2854344 and OR = 0.80 (0.66-0.96) for rs198580). However, after adjusting for multiple testing these associations were borderline with an adjusted P-trend = 0.068 for the most significant SNP (rs2854344). The RB1 gene is not known to contain any coding SNPs with allele frequencies > or = 5% but several intronic variants are in perfect linkage disequilibrium with the associated SNPs. Replication studies are needed to confirm the associations with breast cancer.

The structural organisation of LAMA4, the gene encoding laminin alpha4.

We have determined the complete structural arrangement of LAMA4, the gene encoding the laminin alpha4 chain. Using both yeast artificial chromosome clones and total human genomic DNA and primers derived from the cDNA sequence, regions of the gene were amplified and sequenced to determine the splice donor and acceptor sites. The introns were sized by agarose gel electrophoresis of the PCR products. The gene consisted of 39 exons spanning 122 kb. All of the splice sites conformed to the GT/AG rule, except intron 7 which possessed a GC dinucleotide at the donor splice site. The intron/exon ratio was large at 17.8:1, mainly due to large introns at the 5' end of the gene. Regions at both the 5' and 3' end of the gene were subcloned from the yeast artificial chromosomes to enable untranscribed DNA to be sequenced. The gene represents the second of the laminin A gene family to be characterised and its structural organisation is similar to the equivalent regions of the LAMA2 gene.

A point mutation in an intronic branch site results in aberrant splicing of COL5A1 and in Ehlers-Danlos syndrome type II in two British families.

Ehlers-Danlos syndrome (EDS) is a heterogeneous group of connective-tissue disorders characterized by skin fragility, joint laxity, and skeletal deformities. Type V collagen appears to have a causal role in EDS types I and II, which show phenotypic overlap and may sometimes be allelic. Type V collagen can exist as a heterotrimer, [alpha1(V)]2alpha2(V), and it both coassembles with and regulates type I collagen-fibril diameter. Using an intragenic COL5A1 polymorphism, we have demonstrated linkage, at zero recombination, to the same allele in two large British EDS type II families (LOD scores 4.1 and 4.3). Affected members from each family were heterozygous for a point mutation in intron 32 (IVS32:T-25G), causing the 45-bp exon 33 to be lost from the mRNA in approximately 60% of transcripts from the mutant gene. This mutation lies only 2 bp upstream of a highly conserved adenosine in the consensus branch-site sequence, which is required for lariat formation. Although both families shared the same marker allele, we have been unable to identify a common genealogy. This is the first description of a mutation at the lariat branch site, which plays a pivotal role in the splicing mechanism, in a collagen gene. Very probably, the resulting in-frame exon skip has a dominant-negative effect due to incorporation of the mutant proalpha chain into the triple-helical molecule. These findings further confirm the importance of type V collagen in the causation of EDS type II, and the novel collagen mutation indicates the importance of the lariat branch site in splicing.