Current Genomics - Volume 5, Issue 2, 2004

The complex nature and polygenic determination of most bone disorders require new approaches to search for genes and genetic mechanisms underlying these diseases. The present article overviews powerful and promising methodologies, which have been used to study these disorders. One of the most commonly used approaches is that of candidate gene association study, which seeks to test the association between a particular genetic variant (i.e. allele) and a specific phenotypes. These candidate genes are identified a priori based on known biologic function of gene product. As a complement of the candidate gene approach, the whole-genome scan studies employ polymorphic makers throughout the human genome to search genomic regions responsible for determining a trait of interest by linkage and / or linkage disequilibrium analyses. High-throughput methods for differential gene expression profiling are another powerful approach for searching genes underlying complex traits. Depending on their design, these methods allow researchers to get a “snapshot” of either the whole genome or any part. Importantly, in contrast to the various linkage or linkage disequilibrium tests, gene expression profiling provides information about how genes contribute to a trait. These methods of gene expression analysis are a rapidly developing field of functional genomics. It has a very high potential for applications in bone field, including diagnostics, prevention, and treatment of complex bone disorders. Genes usually function via the protein level. Based on the two-dimensional polyacrylamide gel electrophoresis, mass spectrometry and the yeast two-hybrid techniques, proteomics becomes a potentially useful tool for identifying genes and gene functions underlying complex traits. Focused on protein expression profiles and protein-protein interactions, proteomics turns out to be a complement to functional genomics and one of the best ways to clarify complicated biochemical mechanisms underlying complex bone disorders. Promisingly, proteomics may be eventually applied to seeking and screening genes and drug targets for complex bone disorders.

The growing number of completely deciphered genomic sequences provides an enormous reservoir of data, which can be used for addressing questions related to functional and evolutionary biology. The wealth of this approach is documented by the fast growing numbers of recent publications in the field of evolutionary biology based on comparative genomics. Many proteins of the recombination machinery are conserved between plants, fungi and animals but some of them also show remarkable differences regarding their presence, copy number or molecular structure. For example, the protein responsible for double strand break (DSB) induction during meiosis, SPO11, which is related to the subunit A of the archaebacterial topoisomerase VI, is coded by a single gene in animals and fungi. In contrast, plants harbour three distantly related homologues, which seem to have non-redundant functions either in meiosis or in somatic cells and are indispensable for viability. Moreover, plants possess a homologue of the subunit B of the archaebacterial topoisomerase VI, not present in other eukaryotes. We also summarise the recent progress in the usage of genomic data to analyse the evolution of other DNA recombination factors. Finally, several recent studies report on a strong conservation of a reasonable number of intron positions between plants, animals and fungi. This kind of study provides a basis for comparative genomic analyses across kingdoms and demonstrates the existence of ancient introns, a topic of intensive debate.

Sister chromatids are preferred substrates for the recombinational repair of DNA lesions. Sister chromatid recombination (SCR) results in the exchange of genetic information between newly replicated chromatids and ensures that DNA lesions are either tolerated or repaired. Faulty recombinational repair has been correlated to several genetic diseases, including Bloom's syndrome, inheritable breast cancer (BRCA1 and BRCA2), and Fanconi's Anemia. One approach to understand SCR mechanisms is to clarify the SCR phenotypes in mutants defective in well-conserved radiation repair (RAD) genes. These RAD genes include those that participate directly in the recombinational repair of double-strand breaks (DSBs), known as the RAD51 subgroup, and those that participate in the processing of the DNA break, known as the RAD50 subgroup. A systematic analysis of SCR recombination phenotypes in rad mutants revealed multiple pathways for spontaneous and DNA damage-associated SCR in yeast. Studies focused on vertebrate RAD51 genes have suggested similar pathways. In this review, we shall discuss methods for detecting SCR, recombination mechanisms that generate SCR, mammalian and yeast genes that participate in SCR, and genetic diseases characterized by SCR phenotypes.

The mechanism of NADH oxidation varies between living organisms, and is by far the most complex oxidizing system found in mitochondria. In human mitochondria, a unique, but huge structure, with more than 45 subunits, known as complex I, copes with NADH oxidation. This review compiles our present knowledge on the organization of this complex and the putative role of a small subset of its subunits. This review also describes the major progress that has been made in understanding the molecular bases of respiratory chain complex I deficiency in humans, with mutations identified in both the mitochondrial and the nuclear genes encoding complex I subunits. Finally, the puzzling questions raised by the varying clinical presentations of patients with complex I deficiency are discussed in light of our limited knowledge on complex I function in mammalian cells.

Recent studies on host defense against microbial pathogens have demonstrated that innate immunity predated adaptive immune response. Present in all multicellular organisms, the innate defense uses genome-encoded receptors, to distinguish self from non-self. The invertebrate innate immune system employs several mechanisms to recognize and eliminate pathogens: (i) blood coagulation to immobilize the invading microbes, (ii) lectin-induced complement pathway to lyse and opsonize the pathogen, (iii) melanization to oxidatively kill invading microorganisms and (iv) prompt synthesis of potent effectors, such as antimicrobial peptides. Serine proteases play significant roles in these mechanisms, although studies on their functions remain fragmentary, and only several members have been characterized, for example, the serine protease cascade in Drosophila dorsoventral patterning; the Limulus blood clotting cascade; and the silk worm prophenoloxidase cascade. Additionally, serine proteases are involved in processing Späetzle, the Toll ligand for signaling in antimicrobial peptide synthesis. The recent completion of the Drosophila and Anopheles genomes offers a tantalizing promise for genomic analysis of innate immunity of invertebrates. In this review, we discuss the latest genome-wide studies conducted in invertebrates with emphasis on serine proteases involved in innate immune response. We seek to clarify the analysis by using empirical research data on these proteases via classical approaches in biochemical, molecular and genetic methods. We provide an update on the serine protease cascades in various invertebrates and map a relationship between their involvement in early embryonic development, blood coagulation and innate immune defense.

Analysis of whole genome sequences has revealed that genes constitute only a small fraction of the DNA. The function of the remaining part of the genome is still enigmatic. The role of chromosome structure in gene regulation and maintenance of genome stability is increasingly appreciated, which has led to the hypothesis that large parts of the genome may be dedicated to controlling the formation of specific chromosome conformations. Here we review recent advances in our knowledge of chromosome structure and nuclear organization. We describe possible mechanisms by which genomes encode their spatial conformation, such as the use of specific DNA sequence elements to set up local chromatin structures and the potential exploitation of more global sequence characteristics to influence large-scale chromosome conformation. Complete insight into the processes that govern the spatial conformation of chromosomes will reveal new mechanisms of gene regulation and may also explain the large amount of non-coding DNA in genomes.

Two decades of linkage and association studies with candidate genes have attempted to decipher the genetic contributions of complex behavioral disorders such as Schizophrenia and Substance Use Disorders with only limited success. We suggest refining these efforts by: 1) using association studies on epidemiologically sound, general population samples with large sample sizes (2000 - 5000) to detect small effect loci, 2) developing continuous behavioral measures that “cut at nature's joint”, 3) capitalizing on DNA haplotypes showing evidence of positive selection as “candidate gene” regions and 4) integrating the results with translational biological methodologies. We review our studies of a large, conserved Xq13 haplotype and discuss directions for future studies in genetic dissection integrating across complementary linkage, association, and microarray strategies.