Genomic sequencing has made it clear that a large fraction of the genes specifying the core biological functions are shared by all eukaryotes. Knowledge of the biological role of such shared proteins in one organism can often be transferred to other organisms. The goal of the Gene Ontology Consortium is to produce a dynamic, controlled vocabulary that can be applied to all eukaryotes even as knowledge of gene and protein roles in cells is accumulating and changing. To this end, three independent ontologies accessible on the World-Wide Web (http://www.geneontology.org) are being constructed: biological process, molecular function and cellular component.

/pdf/gene-ontology-tool-for-the-unification-of-biology-ppsktrcye3.pdf

Gene Ontology: tool for the unification of biology

The human genome holds an extraordinary trove of information about human development, physiology, medicine and evolution. Here we report the results of an international collaboration to produce and make freely available a draft sequence of the human genome. We also present an initial analysis of the data, describing some of the insights that can be gleaned from the sequence.

/pdf/initial-sequencing-and-analysis-of-the-human-genome-5dapk1rsx1.pdf

Initial sequencing and analysis of the human genome.

A 2.91-billion base pair (bp) consensus sequence of the euchromatic portion of the human genome was generated by the whole-genome shotgun sequencing method. The 14.8-billion bp DNA sequence was generated over 9 months from 27,271,853 high-quality sequence reads (5.11-fold coverage of the genome) from both ends of plasmid clones made from the DNA of five individuals. Two assembly strategies-a whole-genome assembly and a regional chromosome assembly-were used, each combining sequence data from Celera and the publicly funded genome effort. The public data were shredded into 550-bp segments to create a 2.9-fold coverage of those genome regions that had been sequenced, without including biases inherent in the cloning and assembly procedure used by the publicly funded group. This brought the effective coverage in the assemblies to eightfold, reducing the number and size of gaps in the final assembly over what would be obtained with 5.11-fold coverage. The two assembly strategies yielded very similar results that largely agree with independent mapping data. The assemblies effectively cover the euchromatic regions of the human chromosomes. More than 90% of the genome is in scaffold assemblies of 100,000 bp or more, and 25% of the genome is in scaffolds of 10 million bp or larger. Analysis of the genome sequence revealed 26,588 protein-encoding transcripts for which there was strong corroborating evidence and an additional approximately 12,000 computationally derived genes with mouse matches or other weak supporting evidence. Although gene-dense clusters are obvious, almost half the genes are dispersed in low G+C sequence separated by large tracts of apparently noncoding sequence. Only 1.1% of the genome is spanned by exons, whereas 24% is in introns, with 75% of the genome being intergenic DNA. Duplications of segmental blocks, ranging in size up to chromosomal lengths, are abundant throughout the genome and reveal a complex evolutionary history. Comparative genomic analysis indicates vertebrate expansions of genes associated with neuronal function, with tissue-specific developmental regulation, and with the hemostasis and immune systems. DNA sequence comparisons between the consensus sequence and publicly funded genome data provided locations of 2.1 million single-nucleotide polymorphisms (SNPs). A random pair of human haploid genomes differed at a rate of 1 bp per 1250 on average, but there was marked heterogeneity in the level of polymorphism across the genome. Less than 1% of all SNPs resulted in variation in proteins, but the task of determining which SNPs have functional consequences remains an open challenge.

http://science.sciencemag.org/content/sci/291/5507/1304.full.pdf

The sequence of the human genome.

The 4,639,221-base pair sequence of Escherichia coli K-12 is presented. Of 4288 protein-coding genes annotated, 38 percent have no attributed function. Comparison with five other sequenced microbes reveals ubiquitous as well as narrowly distributed gene families; many families of similar genes within E. coli are also evident. The largest family of paralogous proteins contains 80 ABC transporters. The genome as a whole is strikingly organized with respect to the local direction of replication; guanines, oligonucleotides possibly related to replication and recombination, and most genes are so oriented. The genome also contains insertion sequence (IS) elements, phage remnants, and many other patches of unusual composition indicating genome plasticity through horizontal transfer.

/pdf/the-complete-genome-sequence-of-escherichia-coli-k-12-2lpi4d3nz4.pdf

The Complete Genome Sequence of Escherichia coli K-12

Two large-scale yeast two-hybrid screens were undertaken to identify protein-protein interactions between full-length open reading frames predicted from the Saccharomyces cerevisiae genome sequence. In one approach, we constructed a protein array of about 6,000 yeast transformants, with each transformant expressing one of the open reading frames as a fusion to an activation domain. This array was screened by a simple and automated procedure for 192 yeast proteins, with positive responses identified by their positions in the array. In a second approach, we pooled cells expressing one of about 6,000 activation domain fusions to generate a library. We used a high-throughput screening procedure to screen nearly all of the 6,000 predicted yeast proteins, expressed as Gal4 DNA-binding domain fusion proteins, against the library, and characterized positives by sequence analysis. These approaches resulted in the detection of 957 putative interactions involving 1,004 S. cerevisiae proteins. These data reveal interactions that place functionally unclassified proteins in a biological context, interactions between proteins involved in the same biological function, and interactions that link biological functions together into larger cellular processes. The results of these screens are shown here.

http://web.cs.mun.ca/~harold/Courses/Old/CS6754.W04/Diary/403623.pdf

A comprehensive analysis of protein–protein interactions in Saccharomyces cerevisiae

Born 1932, I was brought up in Stettin. I was trained as an organic chemist at Cologne University, before I entered the Institute of Genetics there in 1962 to work on tRNA structure and function. From 1967 to my retirement in 1997 I worked at the Adolf-Butenandt-Institute (Munich University) on several aspects of molecular biology of the budding yeast, for example tRNA genes, Ty elements, the yeast genome project, and programmed proteolysis. I include facets of my engagement in national and international organizations.

Chapter 4 A Life with Yeast Molecular Biology

Prostate Cancer - Combination of Hormonal Ablation and Conformal Therapy

Isolation and purification of tDNA from Mycoplasma.

Between August, 1986 and October, 1987, 364 hyperthermia treatments combined with radiotherapy were performed in mostly locally advanced and recurrent tumors. Half of the patients could only be exposed to relatively small radiation doses of 20 to 36 Gy. The therapy results of 24 patients with 26 treated tumor sites showed complete remission in 22% and partial remission in 42% of the sites. The dependence of tumor remission on the dose of radiation and heat and the inhomogeneity of the temperature distribution are discussed.

[Radiation and hyperthermia in the treatment of superficial and semi-deep tumors: heat profile, heat doses, tumor remission]

In the yeast Saccharomyces cerevisiae, the majority of the tRNA genes are found associated with transposable elements one of which is the Ty element. We noticed that the transcriptional activity of several of these genes was rather different depending on the type of the accompanying element(s) [Nelbock et al. (1985) Biol. Chem. Hoppe-Seyler 366, 1041-1051]. In order to study the influence of a Ty element on tRNA gene expression, we used the method of micro-injection into Xenopus oocytes taking advantage of a Ty+/Ty- allelic pair of a tRNALys1 gene recently isolated in our laboratory. A direct comparison showed that the expression of the plus allele was c. sixfold higher than that of the minus allele. In order to determine sequences of the Ty responsible for this effect, we constructed a number of variants in which distinct segments of the Ty were deleted or rearranged. The activating ability was localized to a 590-bp segment of the Ty containing the 'promoter delta', independent of its orientation or location towards the tRNA gene thus resembling the effects described for transcriptional enhancers. This is the first time that a long-range effect has been observed on the expression of a gene transcribed by RNA polymerase III.

Horst Feldmann

Papers

Chapter 4 A Life with Yeast Molecular Biology

Prostate Cancer - Combination of Hormonal Ablation and Conformal Therapy

Isolation and purification of tDNA from Mycoplasma.

[Radiation and hyperthermia in the treatment of superficial and semi-deep tumors: heat profile, heat doses, tumor remission]

Enhancer-like stimulation of yeast tRNA gene expression by a defined region of the Ty element micro-injected into Xenopus oocytes.