Chapter 1
Introduction to Molecular Pathology

A large component of human disease has a clear genetic component and can be described in a number of ways. Some of us (some say most of us) harbour abnormal genes within us that we have inherited from our parents, which may cause disease, perhaps evident at birth, perhaps developing years in the future. These are described as 'germline' or perhaps 'constitutive'. Alternatively, disease may arise from normal genes that have become abnormal due to the action of an external factor, such as ionising radiation. This may occur only within a particular organ (such as the breast or prostate), in which case the abnormalities should not be present elsewhere in the body or, indeed, in phenotypically normal tissues within the same organ. Accordingly, in certain cases, both normal and abnormal tissues may be sent for analysis and so compared. Molecular pathology is crucial in determining the cellular basis of all of these abnormalities, and in many cases, suggestions as to treatment. A third aspect of genetic analysis is the precise recognition of an infectious agent such as a bacterium or a virus.

This chapter will provide a sound and basic introduction for the chapters to follow that will address the following three factors. Section 1.1 will present a brief historical perspective of key developments in biomedical science that led to the development of molecular genetics, and then the application of this knowledge to the study of disease, i.e. molecular pathology. We then move to Section 1.2, an examination of the different types of human disease and how they come about (i.e. their aetiology). This is followed, in Section 1.3, by a view of the broad nature of the practice of molecular pathology. The chapter concludes, in Section 1.4, with an introduction to the modern practice of molecular pathology.

Learning Outcomes

After studying this chapter, you should confidently be able to.

• Describe major landmarks in the development of molecular pathology
• Recall the various forms of human disease and the leading causes of death
• Comment upon major features of chromosomal structure
• List common pathological alterations in sections of chromosomes
• Explain the importance of single nucleotide polymorphisms
• Describe the development of the practice of molecular pathology

1.1 A Historical Perspective

The modern practice of molecular pathology has only become possible through the slow and steady increase in our knowledge of the many facets of cell biology and deoxyribonucleic acid (DNA). We can view this as part of the development of our scientific understanding of the composition and working of the cell, the nucleus, heredity, chromosomes, genes and DNA (i.e. physiology). Only then can these be used as tools to address disease (i.e. pathology).

1.1.1 The Development of Molecular Genetics

The Nineteenth Century

Although it was well known from the Middle Ages (and probably earlier) that certain physiological features and diseases were present in families, and that these moved down the generations, perhaps one place to start is in the Victorian era with Darwin and Mendel. The former observed amendable hereditary features and postulated links with survival, publishing his seminal work 'The Origin of Species' in 1859. Mendel provided the mechanisms for the transfer of these features, which we now know as genes, and published his findings in 1865 and 1866, although it was not widely disseminated until the early 1900s. Neither was aware of each other's work.

Miescher is credited with the first description, in 1871, of a substance he named nuclein, obtained from the nuclei of white blood cells, whilst in 1882, Flemming further analysed this substance, describing it as chromatin, a material we now know to be the basis of chromosomes (Greek: coloured body). He subsequently observed the replication of chromosomes in dividing cells and concluded that these molecules were the source of information passed from generation to generation. By the end of the century, Kossel had isolated adenine, cytosine, guanine, thymine and uracil from nuclein, for which he was awarded a Nobel Prize.

The Twentieth Century - Part 1

The last century opened with Bateson's coining of the word genetics in 1906, followed by Johannsen's introduction of the term gene in 1909, emphasising the distinction between phenotypic characteristics of individuals and the material units of heredity that determine them. Levene and others extended the work of Kossel, showing that the nucleic acid component was dominated by only four of the complex molecules described by Kossel and that they were connected to each other by a sugar, the basics of nucleotide chemistry. This was followed by the recognition by Boveri, Sutton and others that chromosomes were responsible for heredity, whilst Griffith, in 1928, showed that physical characteristics of pneumococci can be transferred horizontally (i.e. not merely from parent to offspring) between different strains of the bacterium.

The 1940s were a key decade. McClintock did crucial and pioneering work on cytogenetics, such as the demonstration of transposable elements (which we now call transposons) that move from one chromosomal position to another, which was recognised by the award of a Nobel Prize in 1983. Ris and Mirsky described chromosomes as a complex system of non-histone protein, (DNA) and histone with a definite structure, whilst Avery and colleagues considered DNA a 'transforming factor' in the phenotypes of strains of pneumococcus. Beadle and Tatum published data supporting the hypothesis that genes can exert an effect on development and function through enzymes, whilst the Avery-MacLeod-McCarty experiment showed the hereditable material in chromosomes to be DNA, although some considered that heredity lay in the protein component. Smith coined the term 'genomics' to describe the total chromosomal and genetic component of an individual (the genome) in 1943, whilst decades later, in 1995, Wasinger and colleagues used 'the proteome' as a shorthand to describe the complete protein component of a genome present within a cell, a tissue, an organ, or an organism.

In 1950, Chargaff and colleagues showed that the amount of adenine in a sample of DNA was equal to the amount of thymine, and that the amounts of guanine and cytosine were also equivalent, a key component of the structure of DNA and the genes therein. In 1952, Hershey and Chase reported their now classic and crucial experiment where a virus infecting a bacterium unequivocally showed that the hereditable material was not the protein component of the chromosome but was indeed DNA. The Lederbergs did fundamental work in virology and bacterial genetics, showing the transfer of DNA between bacteria.

Although the term 'molecular biology' was introduced by Weaver in 1938, it was later developed by Astbury in 1950, whilst Butel and Melnick used the term in 1970 to describe the ability of the virus SV40 to induce in cells a transformation to a malignant phenotype. Although at the time the expression had wide usage in the exploration of any molecule, by the 1990s its use was becoming focused specifically on the gene. A key feature of molecular biology is that of the development and refinement of analytical techniques that drove the embryo discipline of biomedical science (as it was developing into) from cell-based studies to the more refined study of individual macro- (as in DNA) and micro-molecules (such as penicillin and insulin). An example of such a technique is that of X-ray crystallography, a necessity in the discovery of the structure of DNA by Watson and Crick in 1953, and of the structure of vitamin B12 by Hodgkin in 1955. The following year saw Tjio and Levan using cytogenetics to report the human diploid chromosome number to be 46.

The Twentieth Century - Part 2

The work of Watson and Crick led directly to knowledge of the transcription of DNA into RNA, and the translation of RNA into protein at the ribosome, with the description of ribosomal RNA in 1955 and transfer RNA in 1957. The central dogma of molecular biology was described in 1958, whilst proof of the long-hypothesised existence of messenger RNA (mRNA) was published in 1961. Other steps demonstrated the molecular basis of DNA in its four nucleotides (adenine [A], cytosine [C], guanine [G] and thymine [T]), and that a triplet sequence of these nucleotides defined an amino acid - the genetic code. For example, the triplet GGC ultimately codes (through RNA, where uracil is substituted for thymine) for glycine. Thus, a sequence of GGC-TTA-GTT in the DNA becomes GGC-UUA-GUU in the mRNA, and thereby codes for the tripeptide glycine-leucine-valine, and, by extrapolation, a protein.

The discovery in the 1950s by Luria, Bertani and colleagues of microbial enzymes able to cut specific sections of viral DNA led directly to their development in the 1960s and 1970s by Smith, Kelly, Berg and others as tools for analysing sections of DNA. These restriction endonucleases, such as EcoRI, cut DNA only at specific sites in the nucleotide sequence and provided the impetus for the major breakthrough of genetic engineering. Advances in RNA included the description of small nuclear RNA in 1968 and circular RNA in 1976 - the precursors of whole families of non-coding RNAs (ncRNAs). The 1970s also saw the discovery of methyl- groups on the DNA, a feature now known to be important...