How to query the Homophila database. The user enters the text query in the form of human disease name, OMIM number, fly gene name, or keyword search through the human disease entry box. A window with information on the disease name, and human and fly genes that match the key word query is opened. The user then can examine the details of an individual human-to-
1.1. Cancer is a genetic disease
Cancer is a major health concern of our time, being responsible for more than 25% of deaths worldwide. The past two decades have produced strong evidence for the genetic basis of cancer. Cancer develops as a clonal disease occuring by the accumulation in multiple steps of genetic (or epigenetic) changes in oncogenes, tumor suppressor genes and “guardian” genes that support expansion of a new clone over the old one. Subsequently, it is the natural selection which helps expansion of a new clone carrying characteristics advantageous for proliferation [1, 2, 3].
Nevertheless, the events contributing to cancer are not restricted to the cancer cells. The most encountered example is the case of
Recently, it has been estimated that up to seven rate-limiting genetic / epigenetic events are needed for the development of a common human cancer . These can appear in multiple different combinations depending on which particular tissue or cell-specific “anticancer” barriers are to be circumvented.
A lot of knowledge about cancer was obtained from studying rare familial “monogenic” cancer syndromes. Although the most of the cancer cases appear to be “sporadic”, when cancer- causing gene mutations occurred only in adult somatic cells, these cases also proved to be important for understanding this intricate disease. Other important aspect refferes to the fact that many key molecular factors to cancer progression may not be deregulated at the gene level. Downstream signaling proteins may become up-regulated by alterations in upstream growth factor signaling, altered catabolism, genes inactivated by epigenetic factors, protein expression altered by enzyme activity, degradation, chaperones, etc .
The idea that genes determine the growth behavior of a cell is now widely accepted, with abnormal growth reflecting the action of abnormal genes. The transformation of a normal cell to a cancer cell starts with changes in growth regulatory genes, and in the course of tumor progression further escape from normal growth control is caused by additional alterations in genes that direct cell multiplication and cell survival. Therefore, either somatic or germline mutations are both considered the root cause of cancer. These heritable changes in cancer cells are subsequently the targets for current attempts to develop effective and specific therapies for this disease . As a general rule, one can say that most things related to cancer are a matter of timing and are determined by many other factors like the cell of origin, the mutations accumulated and the environment, together referred to as the molecular “road map” leading to cancer.
At this time, more than 1% of all human genes are believed to be “cancer genes”. Approximately 90% of them represent somatic mutations in cancer, 20% bear germ-line mutations that predispose to cancer and 10% show both somatic and germ-line mutations . Therefore, it was assumed that there are far fewer “pathways” implicated in cancer than genes. The identification of disease-related genes has led to a number of available genetic tests that detect disease or an individual’s risk of disease. Gene tests are available for different disorders and also in cancer testing, some good examples being for the
1.1.1. The origin of cancer
Cancer had been recognized throughout recorded history and was known to the ancient Egyptians, from around 1600 BC, but it was not studied until the seventeenth century, when the formal study of cancer (oncology) was first documented . Nevertheless, only rather recently were registered spectacular progresses in describing the fundamental molecular basis of cancer, following the entry of molecular biology and especially of genetics.
Usually, cancer is behaving exactly as a clonal disease, beginning with a mutational episode in a single cell and then develops in multiple stages through the acquisition of further mutations which are inherited through division, by the progeny of that cell. As the same outcome can also arise by epigenetic factors that alter chromatin structure, without altering the coding DNA, mutations are not the only way in which a cancer cell acquires inactivation or activation of a key gene/ protein. Therefore, sometimes the term “epimutations” is used to encompass both major routes by which cancer cells acquire aberrant expression/activity of key genes and proteins.
The adult human has been estimated to contain in average as many as 1014 cells, most of which could theoretically become a cancer cell given the right sort of genetic (mutations) and epigenetic changes. Replicating cells may be most vulnerable to cancer-causing mutations. Even if some cell type, of which adult nerve cell are good examples, may avoid becoming cancer cell because they are essentially non-proliferating in the adult, most cells either regularly do or can at a pinch replicate . It seems that most adult cells survive on average for 4-6 weeks and then have to be replaced. Also, over a thousand billion cells may die each day and are renewed either by replication of existing cells or from stem cells precursors. Given that every cell gets a significant amount of daily DNA damage and 1011 or more of them will replicate each day- that is a lot of potential cancer cells. Keeping this in mind a cancer might be expected to be a frequent incidence although so far this only happens in 1 in 3 people and usually even then only after 60 or 70 years of potentially mutation-causing events .
It was well documented that there is a geographical variation in cancer incidence and death, and this likely reflects socioeconomic factors. The different roles of genetic predisposition, gene-environment interaction and infectious agents shared importance in causing cancer. Recent research points to the considerable overlap between the behavior of cancer cells and that of cells during normal physiological wound healing and during embryogenesis. Similarities refers to replication, less differentiated state, invasion/migration, with the major differences reflecting the lack of control and the unscheduled nature of replication which characterizes cancer. One intrigued question addressed later was how the organism is able to distinguish between normal growth and tissue repair (normal cell cycle) on one hand and neoplastic growth (cancer cell cycles) on the other. Several theories [2, 3] sustain that for the initial expansion of a clone of cells more than one mutation is needed. Efforts in several science laboratories sustain that in certain cases the mutational route to cancer may be rather short (in molecular terms) with as few as two interlocking mutations required for initiation or progression of cancer- especially in animal models, unlike in man where at least one of these lesions involves particularly “dangerous” oncogenes such as c-
1.2. The origin of oncogenes
An oncogene is a gene that contributes to converting a normal cell into a cancer cell when mutated or expressed at abnormally-high levels. Although the discovery of the origin of oncogenes came in parallel with the study of retroviruses, known in general as cancer inductors, not all retroviruses are tumor viruses [4, 5]. Even from early 1972 many researchers set out to explore the "oncogene hypothesis" proposed by Robert J. Huebner and George J. Todaro of the National Cancer Institute . Looking for one mechanism to explain the induction of cancer by many different agents, Huebner and Todaro had suggested that there are the retroviral oncogenes as part of the genetic baggage of all cells, perhaps acquired through viral infection early in evolution. They supposed that the oncogenes would be innocuous as long as they remained quiescent. When stimulated into activity by a carcinogenic agent, however, they could convert cells to cancerous growth. It was reasoned that if the hypothesis was correct, the
It was thus supposed that the biologically active cellular oncogenes are mutant forms of normal proto-oncogenes that differ in the regulation of their expression or in the structure and function of their gene products. Nevertheless, biologically active cellular oncogenes were also identified by the ability of tumor DNAs to induce transformation in gene transfer assays. Such experiments have led to the identification of more than a dozen distinct human oncogenes that are activated either by point mutations or by DNA rearrangements in human neoplasm, or by DNA rearrangements that occur in the process of gene transfer. Both somatic mutations and DNA rearrangements of such oncogenes activated in human tumors suggest implication of cellular oncogene activation in the pathogenesis of human cancers. Accordingly, in the last decade, many researchers focused their attention to studies extremely helpful in elucidating these questionable aspects by implying model organisms.
1.2.1. Classification of cancer-genes
The genetic injure present in a parental tumorigenic cell, if not correctable, is maintained such that it is a heritable trait of all subsequent generations cells. Most, if not all cancer cells contain genetic damage that appears to be the responsible event leading to tumorigenesis. Two types of genetic damage are generally found in cancer cells:
A. Dominant genetic damage- and the involved genes have termed proto-oncogenes
A proto-oncogene is a gene whose altered protein product has the capacity to induce cellular transformation. The distinction between proto-oncogene and oncogene relates to the activity of the protein product of the gene. Therefore, an oncogene is believed to be a gene that has sustained some genetic damage and produces a protein capable of cellular transformation. The process of activation of proto-oncogenes to oncogenes can include retroviral transduction or retroviral or transposon integration, point mutations, insertion mutations, gene amplification, chromosomal translocation and/or protein-protein interactions. In general, proto-oncogenes have been classified based upon sequence homology to other known proteins or based on their normal function within cells . As predicted, proto-oncogenes have been identified at all levels of the various signal transduction cascades that control cell growth, proliferation and differentiation. A common rule ascertains that proto-oncogenes which were originally identified as inhabitants in transforming retroviruses are designated as
B. Recessive genetic damage- and the involved genes are variously termed tumor suppressors/ growth suppressors/ recessive oncogenes or anti-oncogenes
There is an equally important category of cancer genes that contribute to tumorigenesis through a loss of function named tumor suppressor genes. In contrast to the growth stimulatory oncogenes, tumor suppressors normally function as attenuators and as inhibitors of growth. This category includes any gene that has the potential of becoming a constitutive growth stimulator and determinant of oncogenic cellular properties. The oncogenicity of these genes is therefore correlated with a gain of function.
The normal versions of both oncogenes and tumor suppressor genes serve in diverse regulatory systems of the cell. Most proto-oncogenes encode for components of signal transduction pathways that convert an extracellular stimulus into a programmed pattern of gene expression. A functional relatedness of tumor suppressors is less apparent; some domains of tumor suppressor gene action include cell surface properties, signal transduction, gene transcription, DNA repair, and checkpoints for cell division .
1.3. Identification of new target cancer-related genes by insertional mutagenesis
Insertional mutagenesis is a good mechanism for identifying new cellular proto-oncogenes, especially when correlating with neoplasms induction by different viruses or transposons. The activation of a candidate oncogene by such an insertion not only provides a means of identifying such potential oncogenes, but also permits their isolation as molecular clones for subsequent investigation. After integrating its viral/ transposon DNA, the virus/ transposon itself represents a marker that could help to isolate surrounding molecular clones. These clones can furthermore provide the flanking genomic sequences, which can reveal the targeted proto-oncogene activated/ targeted by the insertion. A good example for this mechanism is provided by
Most recently studies revealed that one of the best examples of how a transposon insertion could facilitate for oncogene/ candidate-cancer gene identification is given by numerous studies from
1.4. How similar are the fruit flies and humans
The fundamental aspects of the cellular biology, regulation the gene expression, neuronal connectivity, synaptogenesis, cellular signaling and cellular death are commonly accepted as being similar between humans and fruit flies. The structural homology between
A list of disease phenotypes resulting from mutations in genes that are highly related to
2. Why studying cancer in the fruit flies?
The expansion of human cancer is a multistep route, involving the cooperation of mutations in signaling, cell-cycle and cell-death pathways, as well as interactions between the tumor and the microenvironment. In this context, to
In 2009, the second BioMed Conference from Barcelona entitled "Modelling Cancer in
2.1. Testing for preserved function between Drosophila genes and human counterparts
There are a number of ways to test the function of a foreign gene/ protein in transgenic
Herein are presented some results of a study concerned on the clarification of the putative functional conservation between
2.2. Exploration of the DmManf in comparison with Manf putative oncogene from human
Mesencephalic astrocyte-derived neurotrophic factor,
Their finding of an AGG-to-AAG (arg-to-lys) mutation in the adjacent codon 51 in 2 tumors emphasized further the importance of this region. Other evidence  suggested that only a single copy of the
Mesencephalic astrocyte-derived neurotrophic factor sequences are referring to a family of small proteins of approximately 170 residues which contain four di-sulfide bridges that are highly conserved, from nematodes to humans (Table 2).
|1||mesencephalic astrocyte-derived neurotrophic factor||182 aa|
|2||mesencephalic astrocyte-derived neurotrophic factor||246 aa|
|3||mesencephalic astrocyte-derived neurotrophic factor||179 aa|
|4||mesencephalic astrocyte-derived neurotrophic factor||179 aa|
|5||mesencephalic astrocyte-derived neurotrophic factor||179 aa|
|6||mesencephalic astrocyte-derived neurotrophic factor||179 aa|
|7||mesencephalic astrocyte-derived neurotrophic factor||180 aa|
|9||Mesencephalic astrocyte-derived neurotrophic factor||173 aa|
|11||hypothetical protein||168 aa|
The amino acid sequences of this highly conserved protein in evolution shows 51% identity on average between
The higher homology revealed by the comparison of the whole aminoacid sequences of the
To study whether
In this study we took advantage of our first reported loss-of-function mutant
Most of the
A wealth of data support the view that cancer is a multistage disease progressing via the accumulation of multiple genetic changes lesions that compromise the normal control of cell proliferation, survival, differentiation, migration and social interactions with neighboring cells . It is also important to note that apparently phenotypically similar cancers may arise through different combinations of lesions: there are likely many different routes to cancer even in the same cell type. Many key cancer –relevant signaling pathways may be activated or inactivated by mutations at various points that could result in largely identical cell behavior.
From this point of view, interesting cancer-like phenotypes were observed for different mutants previously obtained after
The appearances of the melanotic masses in flies were extensively characterized. Although the
Nucleolus, another important cellular structure displayed an oval shape with irregular contour (Figs. 5a, 5b), without specific membrane, being surrounded by a network of chromatic filaments of nuclear origin which twisted into perinucleolar chromatin. As typical, is prearranged in a fibrillar-granular structure consisting of fibrils and ribo-nucleo-proteinic granules, like ribosomes.
In contrast, in the „old tumor” dark black masses, resembling senescent cells (Fig. 5c) appeared having ceased its function, many structural changes occurring probably due to the intensification of autolitic processes. As a result, the cytoplasmic content decreased significantly, most of the cellular volume being replaced by the vacuolar system. In this cell the nucleus still persists, although the nucleolus have disappeared. Only a few blocks of chromatin and an increased number of autolytic vacuoles can be detected. These changes lead to the idea that such a cell is doomed to apoptosis. Other notable melanotic-like nodule phenotype was observed in case of the
Particular mutants expressing characteristic melanotic phenotype were also obtained in other insertional/ excisional
Half a century ago, melanotic tumors in
Experiments in our laboratory indicated that an artificial transposon,
2.3. Application of the rescue phenotype (gene therapy) technique in flies
Gene therapy consists of the insertion of genes into an individual's cells and tissues to treat a disease, and hereditary diseases, in which a defective mutant allele is replaced by a functional one. Although the technology is still in its infancy, the researchers have already successfully tested it in
2.3.1. DmManf is the true ortholog of the human Manf putative oncogene
To investigate whether human
We proved that the lethality was only caused by deletion of
Herein is described a pilot experiment that supports the evolutionary conservation of the
By the experiments involving
3. Materials and methods
Cancer can be measured as a complex multistep pathology that requires the accumulation of several mutations giving to cells an aberrant proliferative advantage, improved resistance to pro-apoptotic stimuli and loss of differentiation markers. Increasing evidences underline the importance of the tumor microenvironment in the growth of cancer cells. Since it has been shown to exert both pro- and anti-tumoral effects, the role of the immune system in fighting cancer progression has been contradictory. Due to the simplicity of genetic manipulations,
We owe our special thanks to Dr. J. Szidonya for kindly providing EP(3)3171 and w;TM3SerGFP/TM6TbSb Drosophila laboratory strains. We are grateful to Dr. T.I.Heino for providing UAS-DmManf and UAS-HuManf transgenic stocks. We are appreciative to C. Parvu for EM analysis, to A. Burcea for the help with the Figures and really indebted to Dr. I Kiss for kindly review this paper. This research was supported by UEFISCDI Romania, IDEAS grant no.1004/ 2009, ID code 1936/2009.