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Abstract Background Mammalian olfactory receptors (ORs) are subject to a remarkable but poorly understood regime of transcriptional regulation, whereby individual olfactory neurons each express only one allele of a single member of the large OR gene family. Results We performed a rigorous search for enriched sequence motifs in the largest dataset of OR promoter regions analyzed to date. We combined measures of cross-species conservation with databases of known transcription factor binding sites and ab initio motif-finding algorithms. We found strong enrichment of binding sites for the O/E family of transcription factors and for homeodomain factors, both already known to be involved in the transcriptional control of ORs, but did not identify any novel enriched sequences. We also found that TATA-boxes are present in at least a subset of OR promoters. Conclusions Our rigorous approach provides a template for the analysis of the regulation of large gene families and demonstrates some of the difficulties and pitfalls of such analyses. Although currently available bioinformatics methods cannot detect all transcriptional regulatory elements, our thorough analysis of OR promoters shows that in the case of this gene family, experimental approaches have probably already identified all the binding factors common to large fractions of OR promoters.

Abstract Background DNA methylation is a major epigenetic modification important for regulating gene expression and suppressing spurious transcription. Most methods to scan the genome in different tissues for differentially methylated sites have focused on the methylation of CpGs in CpG islands, which are concentrations of CpGs often associated with gene promoters. Results Here, we use a methylation profiling strategy that is predominantly responsive to methylation differences outside of CpG islands. The method compares the yield from two samples of size-selected fragments generated by a methylation-sensitive restriction enzyme. We then profile nine different normal tissues from two human donors relative to spleen using a custom array of genomic clones covering the euchromatic portion of human chromosome 1 and representing 8% of the human genome. We observe gross regional differences in methylation states across chromosome 1 between tissues from the same individual, with the most striking differences detected in the comparison of cerebellum and spleen. Profiles of the same tissue from different donors are strikingly similar, as are the profiles of different lobes of the brain. Comparing our results with published gene expression levels, we find that clones exhibiting extreme ratios reflecting low relative methylation are statistically enriched for genes with high expression ratios, and vice versa, in most pairs of tissues examined. Conclusion The varied patterns of methylation differences detected between tissues by our methylation profiling method reinforce the potential functional significance of regional differences in methylation levels outside of CpG islands.

The human Y chromosome began to evolve from an autosome hundreds of millions of years ago, acquiring a sex-determining function and undergoing a series of inversions that suppressed crossing over with the X chromosome (1,2). Little is known about the recent evolution of the Y chromosome because only the human Y chromosome has been fully sequenced. Prevailing theories hold that Y chromosomes evolve by gene loss, the pace of which slows over time, eventually leading to a paucity of genes, and stasis (3,4). These theories have been buttressed by partial sequence data from newly emergent plant and animal Y chromosomes (5-8), but they have not been tested in older, highly evolved Y chromosomes such as that of humans. Here we finished sequencing of the male-specific region of the Y chromosome (MSY) in our closest living relative, the chimpanzee, achieving levels of accuracy and completion previously reached for the human MSY. By comparing the MSYs of the two species we show that they differ radically in sequence structure and gene content, indicating rapid evolution during the past 6 million years. The chimpanzee MSY contains twice as many massive palindromes as the human MSY, yet it has lost large fractions of the MSY protein-coding genes and gene families present in the last common ancestor. We suggest that the extraordinary divergence of the chimpanzee and human MSYs was driven by four synergistic factors: the prominent role of the MSY in sperm production, 'genetic hitchhiking' effects in the absence of meiotic crossing over, frequent ectopic recombination within the MSY, and species differences in mating behaviour. Although genetic decay may be the principal dynamic in the evolution of newly emergent Y chromosomes, wholesale renovation is the paramount theme in the continuing evolution of chimpanzee, human and perhaps other older MSYs., This study required that we first complete the sequencing of the chimpanzee MSY (9-11) using large-insert bacterial artificial chromosome (BAC) clones and the iterative mapping and sequencing strategy used to [...]

Several studies of the extensive copy-number variation observed in the human olfactory receptor (OR) gene families are conducted to explain the different selective and mechanistic forces that act on such families. The OR family is shown to be highly affected by the homology-based, as well as homology-independent processes.

Author(s): Elena V. Linardopoulou [1, 2]; Eleanor M. Williams [1]; Yuxin Fan [1, 4]; Cynthia Friedman [1]; Janet M. Young [1]; Barbara J. Trask (corresponding author) [1, 2, 3] The [...]

We have developed a fluorescence-based method for mapping single or multiple protein-binding sites on straightened, large-size DNA molecules (> 5 kbp). In the described method, protein-DNA complexes were straightened and immobilized on a fiat surface using surface tension. A fraction of the immobilized complexes displayed a sharp DNA bend with two DNA segments extending from the apex. The presence of DNA-binding proteins at the apex was verified by atomic force microscopy. The position of protein binding relative to the ends of the DNA molecule was determined by measuring the length of two DNA segments using fluorescence microscopy. We demonstrate the potential of the fluorescence-based method to localize protein-binding sites on the DNA template and to evaluate relative binding affinity. The proposed protein-binding-site mapping technique is simple and easy to perform. Practical applications include screening for DNA-binding proteins and the localization of protein-binding sites on large segments of DNA.