Molecular and genetic factors in disease

Molecular and genetic factors in disease

D.R. FitzPatrick

J.R. Seckl

Almost all diseases have a genetic component. In children and young adults in particular, many of the disorders causing long-term morbidity and mortality are genetically determined. The molecular basis of most Mendelian (or ‘single-gene’) diseases has now been determined, and our understanding of the abnormalities in cell function responsible for the clinical presentation is improving. It has also become clear that variants in many genes contribute to the pathogenesis of several common diseases such as asthma, rheumatoid arthritis and osteoporosis. In this chapter, we review key principles of cell biology, cellular signalling and molecular genetics, with emphasis on the diagnosis and assessment of patients with genetic diseases.

Functional anatomy and physiology

DNA, chromosomes and chromatin

The nucleus is a membrane-bound compartment found in all cells except erythrocytes and platelets. The human nucleus contains 46 chromosomes, each a single linear molecule of deoxyribonucleic acid (DNA) complexed with proteins to form chromatin. The basic protein unit of chromatin is the nucleosome, comprising 147 base pairs (bp) of DNA wound round a core of four different histone proteins. The vast majority of chromosomal DNA is double-stranded, with the exception of the ends of chromosomes, where ‘knotted’ domains of single-stranded DNA, called telomeres, are found. Telomeres prevent degradation and accidental fusion of chromosomal DNA.

The genome comprises approximately 3.1 billion bp of DNA. Humans are diploid organisms, meaning that each nucleus contains two copies of the genome, visible microscopically as 22 identical chromosomal pairs – the autosomes – named 1 to 22 in descending size order (see Fig. 3.11, p. 57), and two sex chromosomes (XX in females and XY in males). Each DNA strand consists of a linear sequence of four bases – guanine (G), cytosine (C), adenine (A) and thymine (T) – covalently linked by phosphate bonds. The sequence of one strand of double-stranded DNA determines the sequence of the opposite strand because the helix is held together by hydrogen bonds between adenine and thymine or guanine and cytosine nucleotides.

Genes and transcription

Genes are functional elements on the chromosome that are capable of transmitting information from the DNA template via the production of messenger ribonucleic acid (mRNA) to the production of proteins. The human genome contains an estimated 21 500 genes, although many of these are inactive or silenced in different cell types. For example, although the gene for parathyroid hormone (PTH) is present in every cell, activation of gene expression and production of PTH mRNA is virtually restricted to the parathyroid glands. Genes that are active in different cells undergo transcription, which requires binding of an enzyme called RNA polymerase II to a segment of DNA at the start of the gene termed the promoter. Once bound, RNA polymerase II moves along one strand of DNA, producing an RNA molecule that is complementary to the DNA template. A DNA sequence close to the end of the gene, called the polyadenylation signal, acts as a signal for termination of the RNA transcript (Fig. 3.1). The activity of RNA polymerase II is regulated by transcription factors. These proteins bind to specific DNA sequences at the promoter, or to enhancer elements that may be many thousands of base pairs away from the promoter. A loop in the chromosomal DNA brings the enhancer close to the promoter, enabling the bound proteins to interact.

The human genome encodes approximately 1200 different transcription factors, and mutations in many of these can cause genetic diseases (Fig. 3.2). Mutation of the transcription factor binding sites within promoters or enhancers also causes genetic disease. For example, the blood disorder alpha-thalassaemia can result from loss of an enhancer located more than 100 000 bp from the alpha-globin gene promoter, leading to greatly reduced transcription. Similarly, variation in the promoter of the gene encoding intestinal lactase determines whether or not this is ‘shut off’ in adulthood, producing lactose intolerance.

The accessibility of promoters to RNA polymerase II depends on the structural configuration of chromatin. Transcriptionally active regions have decondensed (or ‘open’) chromatin (euchromatin). Conversely, transcriptionally silent regions are associated with densely packed chromatin called heterochromatin. Chemical modification of both the DNA and core histone proteins allows heterochromatic regions to be distinguished from open chromatin. DNA can be modified by addition of a methyl group to cytosine molecules (methylation). In promoter regions, this silences transcription, since methyl cytosines are usually not available for transcription factor binding or RNA transcription. The core histones can also be modified via methylation, phosphorylation, acetylation or sumoylation at specific amino acid residues in a pattern that reflects the functional state of the chromatin; this is called the histone code – reflecting an emerging understanding of the ‘rules’ by which specific modifications mark transcriptionally activating (trimethylation of lysine 4 on histone H3; acetylation of many histone residues) or silencing (methylation of lysine 9 on histone H4; deacetylation of many histone residues) effects. Such DNA and protein modifications are termed epigenetic, as they do not alter the primary sequence of the DNA code but have biological significance in chromosomal function. Abnormal epigenetic changes are increasingly recognised as important events in the progression of cancer, allowing expression of genes which are normally silenced during development to support cancer cell de-differentiation (see Box 3.3, p. 54). They also afford therapeutic targets. For instance, the histone deacetylase inhibitor vorinostat has been successfully used to treat cutaneous T-cell lymphoma, due to the re-expression of genes that had previously been silenced in the tumour. These genes encode transcription factors which promote T-cell cell differentiation as opposed to proliferation, thereby causing tumour regression.

RNA splicing, editing and degradation

Transcription produces an RNA molecule that is a copy of the whole gene, termed the primary or nascent transcript. RNA differs from DNA in three main ways:

The nascent RNA molecule then undergoes splicing, to generate the shorter, ‘mature’ mRNA molecule, which provides the template for protein production. Splicing removes the regions of the nascent RNA molecule that are not required to make protein (intronic regions), and retains and rejoins those segments that are necessary for protein production (exonic regions). Splicing is a highly regulated process that is carried out by a multimeric protein complex called the spliceosome. Following splicing, the mRNA molecule is exported from the nucleus and used as a template for protein synthesis. It should be noted that many genes produce more than one form of mRNA (and thus protein) by a process termed alternative splicing. Different proteins from the same gene can have entirely distinct functions. For example, in thyroid C cells the calcitonin gene produces mRNA encoding the osteoclast inhibitor calcitonin (p. 738), but in neurons the same gene produces an mRNA with a different complement of exons via alternative splicing, which encodes the neurotransmitter calcitonin-gene-related peptide.

The portion of the mRNA molecule that directs synthesis of a protein product is called the open reading frame (ORF). This comprises a contiguous series of three sequential bases (codons), which specify that a particular amino acid should be incorporated into the protein. There are 64 different codons; 61 of these specify incorporation of one of the 20 amino acids, whereas the remaining three codons – UAA, UAG and UGA (stop codons) – cause termination of the growing polypeptide chain. In humans, most ORF start with the amino acid methionine, which is specified by the codon AUG. All mRNA molecules have domains before and after the ORF called the 5′ untranslated region (5′UTR) and 3′UTR, respectively. The start of the 5′UTR contains a cap structure that protects mRNA from enzymatic degradation, and other elements within the 5′UTR are required for efficient translation. The 3′UTR also contains elements that regulate efficiency of translation and mRNA stability, including a stretch of adenine bases known as a polyA tail.

However, there are approximately 4500 genes in humans in which the transcribed RNA molecules do not code for proteins. There are various categories of non-coding RNA (ncRNA), including transfer RNA (tRNA), ribosomal RNA (rRNA), ribozymes and microRNA (miRNA). There are more than 1000 miRNAs that bind to various target mRNAs, typically in the 3′UTR, to affect mRNA stability. This usually results in enhanced degradation of the target mRNA, leading to translational gene silencing. Together, miRNAs affect over half of all human genes and have important roles in normal development, cancer and common degenerative disorders. This is the subject of considerable research interest at present.

Translation and protein production

Following splicing and export from the nucleus, mRNAs associate with ribosomes, which are the sites of protein production (see Fig. 3.1). Each ribosome consists of two subunits (40S and 60S), which comprise non-coding rRNA molecules complexed with proteins. During translation, tRNA binds to the ribosome. The tRNAs deliver amino acids to the ribosome so that the newly synthesised protein can be assembled in a stepwise fashion. Individual tRNA molecules bind a specific amino acid and ‘read’ the mRNA ORF via an ‘anticodon’ of three nucleotides that is complementary to the codon in mRNA. A proportion of ribosomes are bound to the membrane of the endoplasmic reticulum (ER), a complex tubular structure that surrounds the nucleus. Proteins synthesised on these ribosomes are translocated into the lumen of the ER, where they undergo folding and processing. From here the protein may be transferred to the Golgi apparatus, where it undergoes post-translational modifications, such as glycosylation (covalent attachment of sugar moieties), to form the mature protein that can be exported into the cytoplasm or packaged into vesicles for secretion. The clinical importance of post-translational modification of proteins is shown by the severe developmental, neurological, haemostatic and soft-tissue abnormalities that occur in patients with mutations of the enzymes that catalyse the addition of chains of sugar moieties to proteins. An example is phosphomannose isomerase deficiency, in which there is a defect in the conversion of fructose-6-phosphate to mannose-6-phosphate. This results in a defect in supply of D-mannose derivatives for glycosylation of a variety of proteins, resulting in a multi-system disorder characterised by protein-losing enteropathy, hepatic fibrosis, coagulopathy and hypoglycaemia. Post-translational modifications can also be disrupted by the synthesis of proteins with abnormal amino acid sequences. For example, the most common mutation in cystic fibrosis (ΔF508) results in an abnormal protein that cannot be exported from the ER and Golgi.

Mitochondria and energy production

The mitochondrion is the main site of energy production within the cell. Mitochondria arose during evolution via the symbiotic association with an intracellular bacterium. They have a distinctive structure with functionally distinct inner and outer membranes. Mitochondria produce energy in the form of adenosine triphosphate (ATP). ATP is mostly derived from the metabolism of glucose and fat (Fig. 3.3). Glucose cannot enter mitochondria directly but is first metabolised to pyruvate via glycolysis. Pyruvate is then imported into the mitochondrion and metabolised to acetyl-coenzyme A (CoA). Fatty acids are transported into the mitochondria following conjugation with carnitine and are sequentially catabolised by a process called β-oxidation to produce acetyl-CoA. The acetyl-CoA from both pyruvate and fatty acid oxidation is used in the citric acid (Krebs) cycle – a series of enzymatic reactions that produces CO2, NADH and FADH2. Both NADH and FADH2 then donate electrons to the respiratory chain. Here these electrons are transferred via a complex series of reactions resulting in the formation of a proton gradient across the inner mitochondrial membrane. The gradient is used by an inner mitochondrial membrane protein, ATP synthase, to produce ATP, which is then transported to other parts of the cell. Dephosphorylation of ATP is used to produce the energy required for many cellular processes.

Fig. 3.3 Mitochondria.
A Mitochondrial structure. There is a smooth outer membrane surrounding a convoluted inner membrane, which has inward projections called cristae. The membranes create two compartments: the inter-membrane compartment, which plays a crucial role in the electron transport chain, and the inner compartment (or matrix), which contains mitochondrial DNA and the enzymes responsible for the citric acid (Krebs) cycle and the fatty acid β-oxidation cycle. B Mitochondrial DNA. The mitochondrion contains several copies of a circular double-stranded DNA molecule, which has a non-coding region, and a coding region which encodes the genes responsible for energy production, mitochondrial tRNA molecules and mitochondrial rRNA molecules. ATP = adenosine triphosphate; NADH = nicotinamide adenine dinucleotide. C Mitochondrial energy production. Fatty acids enter the mitochondrion conjugated to carnitine by carnitine-palmityl transferase type 1 (CPT I) and, once inside the matrix, are unconjugated by CPT II to release free fatty acids (FFA). These are broken down by the β-oxidation cycle to produce acetyl-CoA. Pyruvate can enter the mitochondrion directly and is metabolised by pyruvate dehydrogenase (PDH) to produce acetyl-CoA. The acetyl-CoA enters the Krebs cycle, leading to the production of NADH and flavine adenine dinucleotide (reduced form) (FADH2), which are used by proteins in the electron transport chain to generate a hydrogen ion gradient across the inter-membrane compartment. Reduction of NADH and FADH2 by proteins I and II respectively releases electrons (e), and the energy released is used to pump protons into the inter-membrane compartment. As these electrons are exchanged between proteins in the chain, more protons are pumped across the membrane, until the electrons reach complex IV (cytochrome oxidase), which uses the energy to reduce oxygen to water. The hydrogen ion gradient is used to produce ATP by the enzyme ATP synthase, which consists of a proton channel and catalytic sites for the synthesis of ATP from ADP. When the channel opens, hydrogen ions enter the matrix down the concentration gradient, and energy is released that is used to make ATP.

Each mitochondrion contains 2–10 copies of a 16 kilobase (kB) double-stranded circular DNA molecule (mtRNA). mtDNA contains 13 protein-coding genes, all involved in the respiratory chain, and the ncRNA genes required for protein synthesis within the mitochondria (see Fig. 3.3). The mutational rate of mtDNA is relatively high due to the lack of protection by chromatin. Several mtDNA diseases characterised by defects in ATP production have been described. mtDNA diseases are inherited exclusively via the maternal line (see Fig. 3.7, p. 51). This unusual inheritance pattern exists because all mtDNA in an individual is derived from that person’s mother via the egg cell, as sperm contribute no mitochondria to the zygote. Mitochondria are most numerous in cells with high metabolic demands, such as muscle, retina and the basal ganglia, and these tissues tend to be the ones most severely affected in mitochondrial diseases (Box 3.1). There are many other mitochondrial diseases that are caused by mutations in nuclear genes, which encode proteins that are then imported into the mitochondrion and are critical for energy production: for example, Leigh’s syndrome and complex I deficiency.

Protein degradation

The cell uses several different systems to degrade proteins and other molecules that are damaged, are potentially toxic or have simply served their purpose. The proteasome is the main site of protein degradation within the cell. The first step in proteasomal degradation is ubiquitination – the covalent attachment of a protein called ubiquitin as a side chain to the target protein. Ubiquitination is carried out by a large group of enzymes called E3 ligases, whose function is to recognise specific proteins that should be targeted for degradation by the proteasome. The E3 ligases ubiquitinate their target protein, which is then transported to a large multiprotein complex called the 26S proteasome, where it is degraded. There is mounting evidence that defects in the proteasome contribute to the pathogenesis of many diseases, particularly degenerative diseases of the nervous system like Parkinson’s disease and some types of dementia that are characterised by formation of abnormal protein aggregates (inclusion bodies) within neurons. At least one inherited disease, termed Angelman’s syndrome, is due to a mutation affecting the UBE3 E3 ligase.

Proteins with complex post-translational modifications are degraded in membrane-bound structures called lysosomes, which have an acidic pH and contain proteolytic enzymes that degrade proteins. There are many inherited defects in lysosomal enzymes that result in failure to degrade intracellular toxic substances. For instance, in Gaucher’s disease, mutations of the gene encoding lysosomal (acid) β-glucosidase lead to undigested lipid accumulating in macrophages, producing hepatosplenomegaly and, if severe, deposition in the brain and mental retardation.

Lysosomes are also crucial for the process of autophagy, a process of self-cannibalisation that allows the cell to adapt to periods of starvation by recycling cellular components. Autophagy is triggered by metabolic stress and begins with the formation of a membrane-bound vesicle called the autophagosome, which contains targeted cellular components such as long-lived proteins and organelles. The autophagosome then fuses with the lysosome to start the degradation and recycling process. Mutations in proteins that are crucial for formation of the autophagosome lead to neurodegenerative diseases in humans, such as juvenile neuronal ceroid lipofuscinosis (Batten’s disease), caused by autosomal recessive mutations in CLN3.

Peroxisomes are small, single membrane-bound cytoplasmic organelles containing many different oxidative enzymes such as catalase. Peroxisomes degrade hydrogen peroxide, bile acids and amino acids. However, the β-oxidation of very long-chain fatty acids appears to be their most important function, since mutations in the peroxisomal β-oxidation enzymes (or the proteins that import these enzymes into the peroxisome) result in the same severe congenital disorder as mutations that cause complete failure of peroxisomal biogenesis. This group of disorders is called Zellweger’s syndrome (cerebrohepatorenal syndrome) and is characterised by severe developmental delay, seizures, hepatomegaly and renal cysts; the biochemical diagnosis is made on the basis of elevated plasma levels of very long-chain fatty acids.

The cell membrane and cytoskeleton

The cell membrane is a phospholipid bilayer, with hydrophilic surfaces and a hydrophobic core (Fig. 3.4). The cell membrane is, however, much more than a simple wall. Cholesterol-rich ‘rafts’ float within the membrane, and proteins are anchored to them via the post-translational addition of complex lipid moieties. The membrane also hosts a series of transmembrane proteins that function as receptors, pores, ion channels, pumps and associated energy suppliers. These proteins allow the cell to monitor the extracellular milieu, import crucial molecules for function, and exclude or exchange unwanted substances. Many protein–protein interactions within the cell membrane are highly dynamic, and individual peptides will associate and disassociate to effect specific roles.

The cell membrane is permeable to hydrophobic substances, such as anaesthetic gases. Water is able to pass through the membrane via a pore formed by aquaporin proteins; mutations of an aquaporin gene cause congenital nephrogenic diabetes insipidus (p. 794). Most other molecules must be actively transported using either channels or pumps. Channels are responsible for the transport of ions and other small molecules across the cell membrane. They open and close in a highly regulated manner. The cystic fibrosis transmembrane conductance regulator (CFTR) is an example of an ion channel that is responsible for transport of chloride ions across epithelial cell membranes. Mutation of the CFTR chloride channel, highly expressed in the lung and gut, leads to defective chloride transport, producing cystic fibrosis. Pumps are highly specific for their substrate and often use energy (ATP) to drive transport against a concentration gradient.

Endocytosis is a cellular process that allows internalisation of larger complexes and molecules by invagination of plasma membrane to create intracellular vesicles. This process is typically mediated by specific binding of the particle to surface receptors. An important example is the binding of low-density lipoprotein (LDL) cholesterol-rich particles to the LDL receptor (LDLR) in a specialised region of the membrane called a clathrin pit. In some cases of familial hypercholesterolaemia (p. 453), LDLR mutations cause failure of this binding and thus reduce cellular uptake of LDL. Other LDLR mutations change a specific tyrosine in the intracellular tail of the receptor, preventing LDLR from concentrating in clathrin-coated pits and hence impairing uptake of LDL, even though LDLR bound to LDL is present elsewhere in the cell membrane.

The shape and structure of the cell are maintained by the cytoskeleton, which consists of a series of proteins which form microfilaments (actin), microtubules (tubulins) and intermediate filaments (keratins, desmin, vimentin, laminins) that facilitate cellular movement and provide pathways for intracellular transport. Dysfunction of the cytoskeleton may result in a variety of human disorders. For instance, some keratin genes encode intermediate filaments in epithelia. In epidermolysis bullosa simplex (p. 1292), mutations in keratin genes (KRT5, KRT14) lead to cell fragility, producing the characteristic blistering on mild trauma.

Receptors, cellular communication and intracellular signalling

Several mechanisms exist that allow cells to communicate with one another. Direct communication between adjacent cells occurs through gap junctions. These are pores formed by the interaction of ‘hemichannels’ in the membrane of adjacent cells. Many diseases are due to mutations in gap junction proteins, including the most common form of autosomal recessive hearing loss (GJB2) and the X-linked form of Charcot–Marie–Tooth disease (GJB1).

Communication between cells that are not directly in contact with each other occurs through hormones, cytokines and growth factors, which bind to and activate receptors on the target cell. Receptors then bind to various other proteins within the cell termed signalling molecules, which directly or indirectly activate gene expression to produce a cellular response.

There are many different signalling pathways; for example, in nuclear steroid hormone signalling, the ligands (steroid hormones or thyroid hormone) bind to their cognate receptor in the cytoplasm of target cells and the receptor/ligand complex then enters the nucleus, where it acts as a transcription factor to regulate the expression of target genes (Box 3.2). However, the most diverse and abundant types of receptor are located at the cell surface, and these activate gene expression and cellular responses indirectly. Activation of a cell surface receptor by its ligand results in a series of intracellular events, involving a cascade of phosphorylation of specific residues in target proteins by an important group of enzymes called kinases. This cascade typically culminates in phosphorylation and activation of transcription factors, which bind DNA and modulate gene expression.

image 3.2   Examples of molecules involved in specific signalling cascades

Receptor Receptor type Ligands Signal transduction Clinical significance
TNFR1 TNF receptor superfamily TNF TRAF2/5, TRADD, IKK, IκB, NFκB, CYLD, RANK Mediator of inflammatory diseases and immune responses
RANK TNF receptor superfamily RANKL TRAF6, IKK, IκB, NFκB Regulates bone resorption
Insulin receptor Receptor tyrosine kinase Insulin IRS1, PI3K, PIP3, PKB, PDK1, mTORC2, GSK3 Regulation of energy homeostasis and glucose metabolism
Erythropoietin receptor Receptor tyrosine kinase Erythropoietin JAK2, STAT5, c-Jun, c-Fos, Src PI3K, PIP3, PDK1, PKB Regulates erythropoiesis
THRα and THRβ Nuclear receptor superfamily T3 Ligand/receptor complex Regulates differentiation and function of many cells and tissues
ERα and ERβ Nuclear receptor superfamily Oestrogen Ligand/receptor complex Important for fertility, reproduction and bone health
GnRHR GPCR GnRH Gq/G11, PLCbetal, PLA(2), PLD, PKC, MAPK Regulates fertility
PTHR1 GPCR PTH, PTHLP Gs, adenyl cyclase, cAMP, PKA, CREB, Gq/G11, PLC, DAG, IP3, PKC, Ca++ Regulates calcium homeostasis and bone turnover


(cAMP = cyclic adenosine monophosphate; CREB = Ca++ intracellular calcium; CYLD = cylindromatosis; DAG = diacylglycerol; ER = (o)estrogen receptor; GnRHR = gonadotrophin releasing hormone receptor; GPCR = G protein-coupled receptor; Gq/G11/Gs = guanine nucleotide binding proteins; GSK3 = glycogen synthetase kinase 3; IκB = inhibitor of kappa B; IKK = I kappa B kinase; IP3 = D-myo-inositol-1,4,5-trisphosphate; IRS1 = insulin receptor substrate 1; JAK2 = Janus activated kinase 2; MAPK = mitogen-activated kinase; mTOR = mammalian target of rapamycin; NFκB = nuclear factor kappa B; PDK1 = phosphoinosotide-dependent kinase 1; PIP3 = phosphoinosotol triphosphate; PI3K = phosphoinosotol 3 kinase; PKA/PKB/PKC = protein kinase A/B/C; PLA/PLC/PLD = phospholipase A/C/D; PTHR1 = parathyroid hormone receptor 1; PTHLP = parathyroid hormone-like protein; RANK = receptor activator of nuclear factor kappa B; STAT5 = signal transducer and activator of transcription; TNF = tumour necrosis factor; TNFR1 = TNF receptor 1; TRAF = TNF receptor-associated factors; TRADD = tumour necrosis factor receptor type 1-associated death domain protein; TRH = thyrotrophin releasing hormone)

Figure 3.5 depicts some of the signalling molecules downstream of the tumour necrosis factor (TNF) receptor. On activation of the receptor by the ligand (in this case, TNF), other molecules, including TNF-receptor-associated proteins (TRAFs), are recruited to the intracellular domain of the receptor. These regulate the activity of a kinase termed IKKγ, which in turn regulates activity of two further kinases termed IKKα and IKKβ. These regulate degradation of an inhibitory protein called IκB, which normally binds to the effector protein NFκB, holding it in the cytoplasm. On receptor activation, a signal is transmitted through TRAFs and the IKK proteins to cause phosphorylation and degradation of IκB, allowing NFκB to translocate to the nucleus and activate gene expression. The system also has negative regulators, including the cylindromatosis (CYLD) enzyme, which regulates the activity of TRAFs by de-ubiquitination. Other transmembrane receptors can be grouped into:

Many receptors can signal only when they assemble as a multimeric complex. Mutations which interfere with assembly of the functional receptor multimer can result in disease. For example, mutations of the insulin receptor that inhibit dimerisation lead to childhood insulin resistance and growth failure. Conversely, some fibroblast growth factor receptor 2 (FGFR2) gene mutations cause dimerisation in the absence of ligand binding, leading to bone overgrowth and an autosomal dominant form of craniosynostosis called Crouzon’s syndrome.

It is becoming clear that specialised projections on the cell surface known as cilia are essential for normal signalling in many tissues. Cilia can be motile or non-motile. Motile cilia are crucial for normal respiratory tract function, with primary ciliary dyskinesia (PCD) resulting in early-onset bronchiectasis due to failure to clear lung secretions. PCD is commonly associated with situs inversus (left–right laterality reversal) as a result of failure of a specific signalling process in very early embryogenesis. Mutations in proteins that are essential for non-motile cilia formation or function are responsible for a large number of autosomal recessive disorders known collectively as ciliopathies, which are commonly associated with intellectual disability, renal cystic dysplasia and retinal degeneration. For example, in the Bardet–Biedl syndrome, mutations in a series of genes encoding ciliary structure cause polydactyly, obesity, hypogonadism, retinitis pigmentosa and renal failure.

Cell division, differentiation and migration

In normal tissues, molecules such as hormones, growth factors and cytokines provide the signal to activate the cell cycle, a controlled programme of biochemical events that culminates in cell division. During the first phase, G1, synthesis of the cellular components necessary to complete cell division occurs. In S phase, the cell produces an identical copy of each chromosome – which carries the cell’s genetic information – via a process called DNA replication. The cell then enters G2, when any errors in the replicated DNA are repaired before proceeding to mitosis, in which identical copies of all chromosomes are segregated to the daughter cells. The progression from one phase to the next is tightly controlled by cell cycle checkpoints. For example, the checkpoint between G2 and mitosis ensures that all damaged DNA is repaired prior to segregation of the chromosomes. Failure of these control processes is a crucial driver in the pathogenesis of cancer, as discussed in Chapter 11 (p. 262).

During development, cells must become progressively less like a stem cell and acquire the morphological and biochemical configuration of the tissue to which they will contribute. This process is called differentiation and it is achieved by activation of tissue-specific genes and inactivation or silencing of genes that maintain the cell in a progenitor state. This epigenetic process enables cells containing the same genetic material to have very different structures and functions. The programme of differentiation is often deranged or partially reversed in cancer cells. A similar mechanism allows adult stem cells to maintain and repair tissues. Cell migration is a process that is also necessary for development and wound healing. Migration also requires the activation of a specific set of genes, such as the transcription factor TWIST, that give the cell polarity and enable the leading edge of the cell to interact with the extracellular environment to control the speed and direction of travel. Again, this process can be reactivated in cancer cells and is thought to facilitate tumour metastasis.

Cell death, apoptosis and senescence

With the exception of stem cells, human cells have only a limited capacity for cell division. The Hayflick limit is the number of divisions a cell population can go through in culture before division stops and the cell enters a state known as senescence. This ‘biological clock’ is of great interest in the study of the normal ageing process. Rare human diseases associated with premature ageing, called progeric syndromes, have been very helpful in identifying the importance of DNA repair mechanisms in senescence (p. 168). For example, in Werner syndrome, a DNA helicase (an enzyme that separates the two DNA strands) is mutated, leading to failure of DNA repair and premature ageing. A distinct mechanism of cell death is seen in apoptosis, or programmed cell death.

Apoptosis is an active process that occurs in normal tissues and plays an important role in development, tissue remodelling and the immune response. The signal that triggers apoptosis is specific to each tissue or cell type. This signal activates enzymes, called caspases, which actively destroy cellular components, including chromosomal DNA. This degradation results in cell death, but the cellular corpse contains characteristic vesicles called apoptotic bodies. The corpse is then recognised and removed by phagocytic cells of the immune system, such as macrophages, in a manner that does not provoke an inflammatory response.

A third mechanism of cell death is necrosis. This is a pathological process in which the cellular environment loses one or more of the components necessary for cell viability. Hypoxia is probably the most common cause of necrosis.

Genetic disease and inheritance


Meiosis is a special form of cell division that only occurs in the post-pubertal testis and the fetal and adult ovary (Fig. 3.6). Meiosis differs from mitosis in two main ways; there are two separate cell divisions and before the first of these there is extensive swapping of genetic material between homologous chromosomes, a process known as recombination. The result of recombination is that each chromosome that a parent passes to his or her offspring is a mix of the chromosomes that the parent inherited from his or her own mother and father. The end products of meiosis are sperm and egg cells (gametes), which contain only 23 chromosomes: one of each homologous pair of autosomes and a sex chromosome. When a sperm cell fertilises the egg, the resulting zygote will thus return to a diploid chromosome complement of 46 chromosomes. The sperm determines the sex of the offspring, since 50% of sperm will carry an X chromosome and 50% a Y chromosome, while each egg cell carries an X chromosome.

The individual steps in meiotic cell division are similar in males and females. However, the timing of the cell divisions is very different (see Fig. 3.6). In females, meiosis begins in fetal life but does not complete until after ovulation. A single meiotic cell division can thus take more than 40 years to complete. In males, meiotic division does not begin until puberty and continues throughout life. In the testes, both meiotic divisions are completed in a matter of days.

Patterns of disease inheritance

Five modes of genetic disease inheritance are discussed below and illustrated in Figures 3.7 and 3.8.

Fig. 3.8 Genomic imprinting and associated diseases.
Several regions of the genome exhibit the phenomenon of imprinting, whereby expression of one or a group of genes is influenced by whether the chromosome is derived from the mother or the father; one such region lies on chromosome 15. A Normal imprinting. Under normal circumstances, expression of several genes is suppressed (silenced) on the maternal chromosome (red), whereas these genes are expressed normally by the paternal chromosome (blue). However, two genes in the paternal chromosome (UBE3 and ATP10A) are silenced. B In sporadic Prader–Willi syndrome, there is a non-disjunction defect on chromosome 15, and both copies of the chromosomal region are derived from the mother (maternal uniparental disomy). In this case, Prader–Willi syndrome occurs because there is loss of function of several paternally expressed genes, including MKRN3, MAGEK2, NDN, PWRN2, C15ORF2 and SNURF-SNRNP. C In sporadic Angelman’s syndrome, both chromosomal regions are derived from the father (paternal uniparental disomy) due to non-disjunction during paternal meiosis. As a result, both copies of the UBE3 gene are silenced and this causes Angelman’s syndrome. Note that the syndrome can also be caused by deletion of this region on the maternal chromosome or a loss-of-function mutation on the maternal copy of UBE3, causing an inherited form of Angelman’s, as illustrated in panel D. D Pedigree of a family with inherited Angelman’s syndrome due to a loss-of-function mutation in UBE3. Inheriting this mutation from a father causes no disease (because the gene is normally silenced in the paternal chromosome) (see individuals I-1, II-1, II-3, III-6), but the same mutation inherited from the mother causes the syndrome (individuals III-3, III-4, IV-4), as this is the only copy expressed and the UBE3 gene is mutated.

Autosomal dominant inheritance

Autosomal dominant disorders result from a genetic abnormality in one of the two copies (alleles) of a single gene. The risk of an affected individual transmitting an autosomal disease to his or her offspring is 50% for each pregnancy, since half the affected individual gametes (sperm or egg cells) will contain the affected chromosome and half will contain the normal chromosome. However, even within a family, individuals with the same mutation rarely have identical patterns of disease due to variable penetrance and/or expressivity. Penetrance is defined as the proportion of individuals bearing a mutated allele who develop the disease phenotype. The mutation is said to be fully penetrant if all individuals who inherit a mutation develop the disease. Expressivity describes the level of severity of each aspect of the disease phenotype. Neurofibromatosis type 1 (NF1, neurofibromin, 17q11.2) is an example of a disease that is fully (100%) penetrant but which shows extremely variable expressivity. The environmental factors and/or variation in other genes that act as modifiers of the mutated gene’s function are mostly unknown. A good example of an environmental influence that can profoundly influence expression of autosomal dominant disease is seen in the triggering of malignant hyperpyrexia by anaesthetic agents in the presence of RYR1 mutations. Autosomal dominant disorders may be the result of either loss or gain of function of the affected gene. For example, adult polycystic kidney disease type 1 is caused by loss-of-function mutations in PKD1, which encodes polycystin I on 16p13.1. Hereditary motor and sensory neuropathy type 1 is caused by increased number of copies (resulting in increased gene dosage) of PMP22, encoding peripheral myelin protein 22 on 17p11.2.

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Apr 9, 2017 | Posted by in GENERAL SURGERY | Comments Off on Molecular and genetic factors in disease

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