《医学遗传学》PPT课件 (2).ppt

Progress of Strategy, Technique and Methodology in Medical Genetics,CONTENTS,Landmarks in genetics and genomics Educating health-care professionals about genetics and genomics Basic knowledge of medical genetics Theoretical、techniques and data base From chromosomes to genes、 genomes、transcriptomes、proteomes、metabonomics Epigenetics/epigenomics Cancer genetics and genomics as an example Personalized medicine,Educating health-care professionals about genetics and genomics,A sampling of resources for genetics education for health-care professionals,BASELINE COMPETENCIES,1 KNOWLEDGE All health professionals should understand:,1 KNOWLEDGE All health professionals should understand:,2 SKILLS All health professionals should be able to:,3 ATTITUDES All health professionals should:,A. Principles related to biological variation,A. Principles related to biological variation,A. Principles related to biological variation,A. Principles related to biological variation,A. Principles related to biological variation,B. Principles related to cell biology,B. Principles related to cell biology,C. Principles related to classical (Mendelian) genetics,C. Principles related to classical (Mendelian) genetics,D. Principles related to molecular genetics,D. Principles related to molecular genetics,D. Principles related to molecular genetics,E. Principles related to development,E. Principles related to development,E. Principles related to development,E. Principles related to development,F. Principles related to new genetic technology,Genetics is the study of single genes and their effects. Genetics in the health-care setting concerns heritable variation that is related to health and disease（Medical Genetics）Genomics, on the other hand, is a relatively new term describing the study of the function and interaction of all the genes in the genome Molecular biology is the study of the structures and functions of macromolecules such as nucleic acids and proteins. Chromosomes: the self-replicating genetic structures of cells containing the cellular DNA that carries in its nucleotide sequence the linear array of genes. Gene: specific segments of DNA composed of distinctive sets of nucleotide pairs in discrete region of a chromosome that encodes a particular protein.,Transcriptomics: the study of transcriptome which is consist of the complete set of mRNA in any given cell. Proteomics：technology by which systematic surveys are made of the expression of large number of distinct species in a biological sample, such as a cell lysate or a biological fluid. Metabonomics encompasses the comprehensive and simultaneous systematic profiling of multiple metabolite levels and their systematic and temporal changes caused by factors such as diet, lifestyle, environment, genetic effects and pharmaceutical effects both beneficial and adverse, in whole organisms.,DNA methylation DNA methylation occurs predominantly in repetitive genomic regions that contain CpG residues. DNA methylation represses transcription directly by inhibiting the binding of specific transcription factors, and indirectly by recruiting methyl-CpG-binding proteins and their associated repressive chromatin remodelling activities. Epigenetic Refers to mitotically or meiotically heritable changes in gene expression that do not involve a change in DNA sequence. Genomic imprinting The epigenetic marking of a locus on the basis of parental origin, which results in monoallelic gene expression. Epigenome The global epigenetic patterns that distinguish or are variable between cell types. These patterns include DNA methylation, histone modifications and chromatin associated proteins.,Genetic testing has been described as “the analysis of human DNA, RNA, chromosomes, proteins and certain metabolites in order to detect heritable disease-related genotypes, mutations, phenotypes or karyotypes for clinical purposes”. Cancer genomics malignant phenotype can rarely, if ever, occur as a result of a single genetic defect. As such we should probably now discard the term cancer genetics and talk of cancer genomics among the repertoire of diseases encompassed by genomic medicine Aspects of genomic medicine relevant to cancer as a multifactorial disorder include inherited mutations that confer an increased predisposition, somatic mutations in the process of carcinogenesis and genetic variations that may be pathological or protective in the context of disease expression or management.,Categories of human genetic disorders,Unifactorial disorders Multifactorial disorders Chromosomal disorders Somatic cell genetic disorders Mitochondrial genetic disorders,Methodologies of Medical Genetics,Population Screening Pedigree Analysis Karyotyping Twin Analysis Human races comparation Constitutive analysis Linkage and Association analysis,Fluorescence in-situ hybridization (FISH) Comparative genomic hybridization (CGH) Multiplex ligation- dependent probe amplification Denaturing high performance liquid chromatography Temperature gradient capillary electrophoresis Single-strand confirmation polymorphisms (SSCP) Denaturing gradient gel electrophoresis (DGGE) Sequencing Haplotype PCR Southern blot Metabolite analysis 2D-gel electrophoresis MS or MS/MS GC-EI/ToF-MS NMR Spectroscopy Techniques,The biogenesis of microRNAs,MicroRNAs can function as tumour suppressors and oncogenes,The different forms of RNA silencing.,RNA-modification strategies for genetic medicine,A Depiction of Chromosome 16 Based on the Determination of Its Actual Sequence by the Human Genome Project.,50,From DNA to protein,Alternative Splicing. A single gene can produce multiple related proteins, or isoforms, by means of alternative splicing.,Examples of Point Mutations.,Reciprocal chromosomal translocations in Burkitt's lymphoma, a solid tumour of B lymphocytes. The genes for making the heavy chains of antibodies (CH) are located on chromosomes 14, whereas those for making the light chains are on chromosomes 2 and 22. These genes are expressed exclusively in B lymphocytes, because only these cells have the necessary transcription factors to switch on their expression. In most (over 90%) of Burkitt's lymphoma cases, a reciprocal translocation moves the proto-oncogene c-myc from its normal position on,A.Painting probes stain entire chromosomes. B. Regional painting probes can be generated by chromosome microdissection. C. Centromeric-repeat probes are available for almost all human chromosomes. D. Large insert clones are available for most genomic regions. e | Special probe sets can be designed to facilitate diagnosis of known structural rearrangements. F.Genomic DNA is used as the probe in comparative genomic hybridization (CGH) to establish copy number. G. For high-resolution analysis, DNA fibres can be used as the target for probe hybridization. H. Microarrays can be used as targets for hybridization to provide resolutions down to the single-nucleotide level. A BAC array is shown, to which test DNA and reference DNA are hybridized,Comparison of cytogenetic techniques for identifying chromosomal abnormalities,Combining cytogenetic approaches to understand a complex chromosomal rearrangement,Studying genome organization using three-dimensional fluorescence in situ hybridization,60,61,What is a DNA Microarray? A DNA micorarray allows scientists to perform an experiment on thousands of genes at the same time. Each spot on a microarray contains multiple identical strands of DNA. The DNA sequence on each spot is unique. Each spot represents one gene.Thousands of spots are arrayed in orderly rows and columns on a solid surface (usually glass).The precise location and sequence of each spot is recorded in a computer database. Microarrays can be the size of a microscope slide, or even smaller.,Study and reference DNA are labeled with a green (a) and red (b) fluorochrome, respectively, and cohybridized to normal metaphase spreads or to microarray. Green and red regions of chromosomes correspond respectively to an overrepresentation (green/red 1.2) and an under representation (green/red 0.8) of the study DNA.,How to sequence DNA,“Sequencing does put microarrays at risk in some areas of the life-sciences research market.”,Individual-cell analysis of signalling. A.Tumour cells that have been isolated from a patient are treated with different environmental cues or therapeutic agents as a way to identify which signalling networks are active. It is possible to study cancer cells from the tumour (Tu), stromal cells (S), cells of the vasculature (V), or immune cells such as T cells (T). B.The same technique can be used to study signalling in subsets ofnormal primary cells, such as T cells (T), B cells (B) or monocytes (M) aftertreatment with various stimuli, such as interleukin (IL)-7 (red circles) or IL-4 (blue circles),Actin fibers (red - stained with rhodamine) and microtubuls (green - stained with fluorescein) in fibroblasts in an in vitro culture. A bottom image - superposition of images of green and red fluorescence,Pedigree analysis,The scope of proteomics,Proteins can be studied in various contexts, including sequence, structure, interactions, expression, localization, and modification. Proteomics is divided into several major but overlapping branches, that embrace these different contexts: (a) sequence & structural proteomics, (b) expression proteomics, (c) interaction proteomics, and (d) functional proteomics.,Sequence and structural proteomics,Three primary nucleic acid sequence databases: Genbank, the EMBL nucleotide sequence database, and the DNA database of Japan (DDBJ). Protein sequence databases: SWISS-PROT, and TrEMBL Protein structure database : The Protein Data Bank (www.rscb.org),Expression proteomics,The analysis of protein abundance and involves the separation of complex protein mixtures, the identification of individual components and their systematic quantitative analysis The key technologies in expression proteomics are 2D-gel electrophoresis and multi-dimensional liquid chromatography (MDLC) for protein separation, mass spectrometry for protein identification, and image analysis or mass spectrometry for protein quantification.,The genome, transcriptome, and proteome,Studying the transcriptome using DNA microarrays,PM: perfectly matched; MM: mismatched (a single base difference in a central position, compared with PM),Interaction proteomics,It studies the genetic and physical interactions among proteins as well as interactions between proteins and nucleic acids or small molecules. One of its ambitious goals is the creation of proteome linkage maps based on binary interactions between individual proteins and higher-order interactions determined by the systematic analysis of protein complexes. Key technologies in this area include (1) the yeast two-hybrid system, and mass spectrometry for the analysis of protein complexes, & (2) biochemical assays and structural analysis methods (e.g., X-ray crystallography, NMR) for protein-ligand interaction study.,Functional proteomics: The study of protein function on a large scale.,The challenges of proteomics,Combination of different protein interaction techniques to increase the level of confidence in the human interactome while minimizing the number of experiments to perform.,Steps involved in protein interaction mapping by affinity purification coupled to mass spectrometry. Possible issues are highlighted.,Cancer is caused by alterations in oncogenes, tumor-suppressor genes, and microRNA genes. These alterations are usually somatic events, although germ-line mutations can predispose a person to heritable or familial cancer. A single genetic change is rarely sufficient for the development of a malignant tumor. Most evidence points to a multistep process of sequential alterations in several, often many, oncogenes, tumor-suppressor genes, or microRNA genes in cancer cells. Tumors often possess cytogenetically different clones that arise from the initialtransformed cell through secondary or tertiary genetic alterations. This heterogeneity contributes to differences in clinical behavior and responses to treatment of tumors of the same diagnostic type.,Functional Properties of Oncogenes,Transcription Factors Chromatin Remodelers Growth Factors Growth Factor Receptors Signal Transducers Apoptosis Regulators,MicroRNA Genes,MicroRNA genes, unlike other genes involved in cancer, do not encode proteins. Instead, the products of these genes consist of a single RNA strand of about 21 to 23 nucleotides; their function is to regulate gene expression. A microRNA molecule can anneal to a messenger RNA (mRNA) containing a nucleotide sequence that complementsthe sequence of the microRNA,Prospects for integrative analysis of comprehensive approaches used to characterize cancer.,Cancer pathways and targeted therapy. a, Multiple signalling pathways upregulated in cancer cells owing to specific alterations in oncogenes or tumour suppressors stimulate tumour-cell proliferation, often by promoting G1S cell-cycle progression b, Classical chemotherapy and radiotherapy eliminates cancer cells by inducing DNA damage and subsequent apoptosis,Complementary approaches to understanding cancer genetics.,Stages of Metastatic Progression,Pressures that Drive Selection for Metastatic Traits,Distinct Fates for Disseminated Cancer Cells,Patterns of Metastatic Colonization,A model of the influence of genetic background on metastatic efficiency,Comparing the different factors that might influence tumour-gene- and metastasisgene-expression patterns,The deepening of our understanding of normal biology has made it clear that stem cells have a critical role not only in the generation of complex multi-cellular organisms, but also in the development of tumors. Recent findings support the concept that cells with the properties of stem cells are integral to the development and perpetuation of several forms of human cancer. Eradication of the stem-cell compartment of a tumor also may be essential to achieve stable, long-lasting remission, and even a cure, of cancer. Advances in our knowledge of the properties of stem cells have made specific targeting and eradication of cancer stem cells a topic of considerable interest.,Embryonic and somatic stem cells as a source of genetic medicines,Summary of LOH studies in ovarian cancer,Summary of molecular cytogenetic studies quantifying genomic imbalance,Gene Silencing in Normal Cells Heritable gene silencing involves, among other processes, the interplay between DNA methylation, histone covalent modifications, and nucleosomal remodeling. Some of the enzymes that contribute to these modifications include DNA methyltransferase (DNMTs), histone deacetylases (HDACs), histone methyltransferases (HMTs), and complex nucleosomal remodeling factors (NURFs). The interplay between these processes establishes a heritable repressive state at the start site of genes resulting in gene silencing. Physiologically, sil