Molecular Phenotyping of Mouse Mutant Resources by RNA Expression Profiling Pp. 121-129
J. Beckers, J. Hoheisel, W.
Mewes, M. Vingron, and M. Hrabé de Angelis
The Use of Phylogenetic Profiles for Gene Predictions Pp. 131-137
David A. Liberles, Anna
Thorén, Gunnar von Heijne and Arne Elofsson
A Dilemma of Functional Genomics: Count the Chickens or Study their Eggs? Pp. 139-148
S.K. Dower and E. Kiss-Toth
The Role of local Insulin-like Growth Factor-1 Isoforms in the Pathophysiology of Skeletal Muscle Pp. 149-162
A. Musarò and N. Rosenthal
Adenoviral Vectors for Cancer Gene Therapy Pp. 163-180
Wenli Zhao, Masanobu
Kobayashi, Masuo Hosokawa and Prem Seth
Genetic Location of Heritable Traits Through Association Studies: A Review Pp. 181-200
D. Garcia, J. Cañon and S.
Dunner
Evolutionary Perspective for Functional Divergence of Gene Family and Applications in Functional Genomics Pp. 201-211
Xun Gu, Yufeng Wang, Jianying
Gu and Kent Vander Velden
Multicolor-FISH Approaches for the Characterization of Human Chromosomes in Clinical Genetics and Tumor Cytogenetics Pp. 213-235
Thomas Liehr and Uwe Claussen
[Back to top] Molecular Phenotyping of Mouse Mutant Resources by RNA Expression Profiling
J. Beckers, J. Hoheisel, W.
Mewes, M. Vingron, and M. Hrabé de Angelis
Microarray-based techniques allow us to visualize and
quantify the expression of every single gene in any population of cells. In
yeast the true potential of large-scale transcriptome analysis in identifying
regulatory units and understanding gene function has already been demonstrated
by evaluating expression profiles of a comprehensive group of mutants. We
discuss the potential of DNA-chip technologies for the analysis of gene
expression in complex organisms. The usefulness of transcriptome analysis for
clinical purposes und diagnosis of cancers is already well established. We
argue that microarray-based expression profiling will also be a useful tool for
the analysis of gene function and approaches complementary to classical
phenotypic description in mammals, particularly in regard of the large
resources of mutant models that are currently being generated by gene-targeting
and mutagenesis of the mouse genome. Experimental requirements and potential
future directions are discussed.
[Back to top] The Use of Phylogenetic Profiles for Gene Predictions
David A. Liberles, Anna Thorén, Gunnar von Heijne and Arne Elofsson
Determining gene functions from genomic sequences is a central goal of bioinformatics. Most purely computational approaches to this problem are based on the detection of genes with similar sequences. With the completion of fully sequenced genomes alternative approaches have become feasible. One such method is that of phylogenetic profiles. In this method a gene is described by its phylogenetic profile, i.e. a string that encodes the presence or absence of a homologous gene in other genomes. This string is then used to search for other genes with similar profiles. In this paper we briefly review the field as well as present an analysis on the performance of the method. We also discuss variations on this theme including inverse phylogenetic profiles and non-exact profiles using phylogenetic trees. In conclusion this indicates that phylogenetic profiles might be useful for some, but not all functional annotations. Functional annotation of genomes remains an important problem in genomics when no close homologs exist.
[Back to top] A Dilemma of Functional Genomics: Count the Chickens or
Study their Eggs?
S.K. Dower and E. Kiss-Toth
A challenge modern molecular biology is faced with, is
defining the organisation of intracellular signalling networks. The latest
advances in microarrays and high throughput proteomics allow us to study
changes in transcription, translation and post-translational modification at
the level of the whole genome, potentially leading to the identification of all
genes affected by a given stimulus. These approaches are very robust, however,
no direct information can be obtained about the signalling events leading to
those observed changes. We liken this situation to finding and studying eggs
laid by a group of hens, but having no idea which chicken laid the eggs of
interest. A complementary strategy is to screen for signalling components
(“chickens”), whose activation leads to changes in gene expression ("eggs").
The most common example of this approach is a genetic screen, where mutations
causing particular phenotypes are identified. These screens are mainly applied
in organisms with a relatively short generation time such as bacteria, yeast,
the worm and the fly. In these systems however, detectable phenotypes only
occur for non-redundant signalling molecules. Thus even using all existing
screening techniques including knockout, genome-sequencing projects reveal a
large number of novel genes with no previously identified function, even for
the best-studied organisms. In addition, the possible involvement of the
majority of those gene products with assigned function in complex processes,
such as embryogenesis or inflammation is unknown. It is therefore desirable to
design a high throughput experimental system, which detects gene products based
on their function. One version of such a system has been developed recently and
is being successfully used for identifying components of the IL-1/TNF
signalling network, utilizing a transcription reporter sensitive to these
cytokines. Upon overexpression, most of the known components of this signalling
network mimic the effect of the cytokine. Based on this observation, bioactive
protein-encoding cDNA clones can be identified in complex pools by the
sensitive reporter. This system provides a proof of principle for a general
strategy, whereby gene products are being detected based on their function.
[Back to top] The Role of local Insulin-like Growth Factor-1 Isoforms in the Pathophysiology of Skeletal Muscle
A. Musarò and N. Rosenthal
The
prolongation of skeletal muscle strength in aging and neuromuscular disease has
been the objective of numerous studies employing a variety of approaches. To
date however, efforts to prevent or attenuate age- or disease-related muscle
degeneration have been largely unsuccessful. Cell-based therapies have been
stalled by the difficulty in obtaining sufficient numbers of autologous
myoblasts and by inefficient incorporation into host muscle. Administration of
growth hormone prevents age-related loss of muscle mass, but has failed to
increase muscle strength. In this context, where direct therapeutic approaches
to redress the primary disease are still suboptimal, it may be more effective
to focus on strategies for improving skeletal muscle function. Experimental
models of muscle growth and regeneration have implicated Insulin-like Growth Factor-1
(IGF-1) as an important mediator of anabolic pathways in skeletal muscle cells.
Two major IGF-1 transcripts are characterized: the locally acting isoform with
an autocrine/paracrine action and the circulating isoform with endocrine
effects. The physiological differences between the function of local and
circulating isoform of IGF-1 are not completely established. However the
selective expression of the muscle-specific IGF-1 isoform avoids hypertrophic
effects on distal organs such as the heart, and eliminates risk of possible
neoplasms induced by inappropriate high expression levels of circulating IGF-1.
In this review we discuss the roles of IGF-1 isoforms in myogenesis and the
potential therapeutic role of local IGF-1 isoform on muscle aging and diseases.
[Back to top] Adenoviral Vectors for Cancer Gene Therapy
Wenli Zhao, Masanobu
Kobayashi, Masuo Hosokawa and Prem Seth
Cancer gene therapy is considered a very viable approach
for the treatment of cancer. The basic idea is to introduce the therapeutic
gene, often in the form of a foreign cDNA, into the cancer or other appropriate
target cells. For gene delivery purposes, several viral and non-viral vectors
have been described. One vector that has become very popular for the purpose of
cancer gene therapy is adenovirus. Main reasons are that the recombinant
adenoviruses expressing the foreign therapeutic genes can be easily generated,
and that the infection of the target cells with the recombinant adenoviruses
produces large quantities of the therapeutic proteins. The review describes the
basic characteristics and the molecular biology of adenoviruses. This review
further describes various methods to generate the recombinant adenoviruses.
These include the methods to generate the first generation adenoviruses (E1
deleted), up to the most recently used gutless viruses (with complete viral
deletions). A critical comparison of the advantages and disadvantages of
various methods is also described. The review also describes the use of
adenoviruses which selectively replicate in cancer cells. The review further
describes the various approaches for cancer gene therapy currently being
exploited. These include the gene replacement strategies using tumor suppressor
genes, anti-oncogenes, use of suicide genes, immunomodulation and
anti-angiogenesis approaches. An in depth analysis of the pre-clinical data
obtained using adenoviral vectors for these cancer gene therapy approaches is
also described. The review concludes with a critical discussion of the
adenoviruses for cancer gene therapy.
[Back to top] Genetic Location of Heritable Traits Through Association Studies: A Review
D. Garcia, J. Cañon and S.
Dunner
In the last few years association and linkage
disequilibrium studies have come to play an important role in the search for
the location of genes underlying certain traits, since linkage analyses provide
less accurate estimations of the positions of the genes as the complexity and
rareness of the traits increase, partly due to the difficulty of getting large
and informative enough samples. These approaches have been proven to be able to
narrow the distance between the expected site of the locus and the nearest
marker, and to reduce sample requirements in terms of size and structure when
compared to those needed for linkage studies to obtain evidence for a gene’s
involvement. On the other hand, the lack of robustness with respect to
population history and structure makes them still a subject of constant
research. The kind of sample, the analysis to perform, the approach of seeking
association with a particular marker versus conducting a complete scan of a
wide part of the genome, the type and number of markers used, the nature of the
trait (discrete or continuous), and the underlying model of the disequilibrium
are some of the different factors needed to be taken into account when
considering association studies. Any of them would by itself justify an
individual review, but it is our intention to provide an overall perspective of
the different approaches available at the current time.
[Back to top] Evolutionary Perspective for Functional Divergence of Gene Family and Applications in Functional Genomics
Xun Gu, Yufeng Wang, Jianying
Gu and Kent Vander Velden
Gene family proliferation has provided the raw materials for functional innovations in many organisms. Although many models have been proposed on how duplicate genes have undergone functional divergence and survived in the long-term evolution, the details remain largely unknown. In this review paper, recent developments about this important issue have been discussed with special references to the implication for functional genomics. With a combination of large-scale genome sequencing and powerful computational analysis, we show that a great deal of functional information can be obtained from the evolutionary perspective, which can in turn be used to facilitate high throughput functional assays.
[Back to top] Multicolor-FISH Approaches for the Characterization of Human Chromosomes in Clinical Genetics and Tumor Cytogenetics
Thomas Liehr and Uwe Claussen
A variety of multicolor fluorescence in situ hybridization (FISH) assays have been developed in the last decade. Routine application of such techniques started in 1996 with the simultaneous use of all the 24 human whole chromosome painting probes (i.e. multiplex-FISH = M-FISH and spectral karyotyping = SKY). Since that time different approaches for chromosomal differentiation based on multicolor-FISH (mFISH) assays have been published with the purpose to characterize structurally abnormal chromosomes and supernumerary marker chromosomes of unknown origin after conventional karyotypic analysis. Their characterization is of high clinical impact and is the requisite condition for further molecular investigations aimed at identification of disease related genes. We present an overview of the available different mFISH methods, highlighting their advantages and limitations, as well as their applications in clinical and tumor cytogenetics. Finally, an outlook is given on further possible developments of this special field of molecular cytogenetics.