Basics of human genetics

Human genetics is the study of the human genome and how genes are transmitted through generations. The human genome consists of 23 pairs of chromosomes (22 pairs of homologous chromosomes and one pair of sex chromosomes), each containing genes that code for proteins within the cell. On all homologous chromosome pairs, there are two forms of the same gene that are known as alleles, which are passed on from parent to offspring. Heritable diseases are also passed down from parent to offspring via different patterns of inheritance, such as autosomal dominant, autosomal recessive, X-linked, and mitochondrial inheritance. These diseases often result from alterations within an individual's genes called mutations. Although some mutations are benign, many cause cellular dysfunction that manifests as disease. Understanding the mechanisms underlying the transmission of genetic material is vital for understanding and treating diseases that have a genetic component.

Genes

• Gene: basic unit of genetic information; DNA segment containing information that is defined by its nucleotide sequence and encodes a specific protein or an RNA as gene product
• Locus: location of a gene on a chromosome
• Allele: one of at least two possible DNA sequences at a particular locus. An allele is usually normal (or wild-type) and common, whereas other alleles are mutations (e.g., polymorphisms) and are rare.
  • A diploid cell that has the same allele at a specific locus (on each chromosome) is homozygote. Heterozygotes are cells that have two different alleles at a given locus.
  • Polymorphism: a rare allele occurring in a population with a certain frequency
    • Example
      • A population is composed of 500 individuals with a diploid set of chromosomes.
      • All genes occur in pairs, with a total number of 1,000 gene copies in the given population.
      • There are two alleles for a specific gene, with one allele representing a polymorphism.
      • The allele frequency of the mutated gene is 1% = 0.01, i.e., 10 of 1,000 gene copies carry the same mutation.
      • If the individual with the polymorphism is a heterozygote, i.e., carries only one copy of the mutated gene with the other gene copy normal, 10 individuals have the polymorphism.
      • Ten individuals in a population of 500 individuals is 10/500 = 0.02 = 2%.
• Gene frequency: the proportion of a particular allele of a gene within a population

For molecular structure of DNA and chromosomes, see nucleotides, DNA, and RNA.

Chromosomes

The chromosomes are only visible during cell division, especially in metaphase.

• Each human cell contains 23 pairs of homologous (identical) chromosomes.
• A chromosome pair contains one chromosome from each parent, respectively.
• Chromosomes are classified in 2 ways:
  • Classification based upon autosomes and allosomes (sex chromosomes)
    • 22 pairs of autosomes, each comprised of 2 homologous chromosomes
    • 1 allosome pair; , which consists of either 2 X chromosomes (♀) or one X and one Y chromosome (♂)
      • Pseudoautosomal regions (PAR): homologous DNA sequences on X and Y chromosomes, with PAR1 on the short arm and PAR2 on the long arm of the sex chromosomes
  • Classification based upon the number of chromosome sets
    • Diploid (double chromosome set): Human somatic cells contain 46 chromosomes, i.e., 46 DNA molecules, of various lengths, which form 23 pairs of homologous chromosomes.
    • Haploid (single chromosome set): After meiosis, germ cells only contain 23 chromosomes (no chromosome pairs!), i.e., a haploid chromosome set with only one copy of each chromosome.
• Depending on the cell cycle phase, a chromosome consists of one or two identical DNA helices, the chromatids.
  • Chromatid: one of the two identical copies of a chromosome resulting from DNA replication
  • Sister chromatids: two identical copies of a chromosome joined at the centromere (i.e. the duplicated chromosome)
    • The genetic information encoded in the two chromatids of a chromosome is identical (with the exception of individual mutations that may occur during cell division).

Characteristics of chromosomes (chromosome morphology)

• Kinetochore: a protein complex found at the centromore of chromosomes that serve as an attachment point for microtubules during mitosis
• Centromere: a region on chromosomes that joins sister chromatids
  • Divides the chromatids into a short p arm and a long q arm
  • Depending on the position of the centromere, the chromosome is considered:
    • Submetacentric: The centromere is not quite positioned in the middle, i.e., the short p arm and the long q arm are markedly visible.
    • Metacentric: The centromere is positioned approx. in the middle, i.e., the p and q arms are approx. of identical length.
    • Acrocentric The centromere is positioned near one end of the chromosome arm, i.e., one arm is much shorter than the other.
      • The acrocentric autosomes (chromosomes 13, 14, 15, 21, and 22) contain the genes for the large ribosomal subunits on their short arms (within the nucleolar organizer region, NOR).
  • Kinetochores are assembled at the centromere, which is why it is often regarded as the place on the chromosome where the spindle attaches.
• Telomere: repetitive, noncoding DNA sequence at the chromosome ends

Chromosomes are classified based on their length, position of the centromere, and pattern of the bands!

For information on genetic testing, see chromosome testing in laboratory methods.

Traits, their genetic basis, and manifestations

• Genotype: genetic composition of an individual
  • The term genotype is often used when describing a certain set of alleles at one or several specific loci.
  • The phenotype, with environmental interaction, develops from the genotype.
  • Based on the genotype, the following states can be distinguished:
    • Homozygote: Two genetically identical alleles present at the same locus.
    • Heterozygote: Two genetically different alleles present at the same locus.
    • Hemizygote: Only one allele is present at the given locus. This occurs for genes located on the X or Y chromosome of males.
• Phenotype: observable traits of an individual
  • The phenotype is determined by a combination of the genotype and environmental factors. Traits include the physical appearance (e.g., eye or hair color) and characteristics (e.g., behavior, personality) of an individual.
  • An inherited trait may manifest very differently depending on the individual phenotype.
    • Dominant and recessive: If only one of both alleles is phenotypically apparent in heterozygous individuals, the allele is dominant. If the allele does not show its effect in the phenotype, it is recessive.
    • Codominance: both alleles are expressed in the phenotype. The phenotype in heterozygous individuals differs from both possible phenotypes in homozygous individuals.
      • Example: In the ABO blood group system, the blood groups A and B are codominant (however, they are expressed dominantly over blood group O)
      • Incomplete dominance: special case occurring in codominance in which there is no one trait dominates completely, but an intermediate trait is expressed
      • Example: sickle cell anemia

Individuals, family, and population

• Multiple alleles: occurrence of more than two different alleles in a population
  • Occurrence: most likely in all human genes
  • Example: ABO blood group system
• Fundamental terms related to genetic diseases
  • Penetrance: the probability of individuals with a particular genotype manifesting signs of the condition
  • Expressivity: the extent of expression of a given genotype at the phenotypic level, e.g., the extent of disease manifestation
    • Example: Patients with Marfan syndrome present with highly variable manifestations of the disease, ranging from mild (arachnodactyly) to life-threatening (aortic aneurysm).
  • Pleiotropy: one gene contributes to multiple seemingly unrelated phenotypic effects, e.g., affects multiple organ systems
    • Examples: phenylketonuria, Marfan syndrome, sickle cell disease
  • Compound heterozygosity: If the same gene on both chromosomes is altered due to differently mutated alleles, the affected gene may lose its function despite the mutation not being homozygous.
  • Anticipation: occurs when a disease increases in severity over several generations or manifests earlier with each generation
    • Often occurs in trinucleotide repeat disorders
    • Example: Huntington disease
  • Allelic heterogeneity: Different mutations in the same allele result in the same phenotype.
    • Example: most diseases with a Mendelian pattern of inheritance, e.g., G6PD deficiency, familial hypercholesterolemia, β-thalassemia
  • Locus heterogeneity: Mutations in genes at different loci cause the same phenotype.
    • Examples: osteogenesis imperfecta, albinism

References:^[1][2]

Mutations are heritable alterations in the genome of a cell. They can occur as a result of errors during DNA replication or cell division as well as ineffective DNA repair mechanisms. Damage to DNA caused by endogenous and exogenous toxins can also lead to gene and chromosome mutations.

Mutations according to the affected cell population

Based on the affected cells, mutations can be classified as follows:

• Germline mutation: (gametic mutation): mutations that are passed on to offspring via the ova or sperm
• Somatic mutation: acquired mutations that are present only in certain somatic cells
  • Primarily affect only one allele of a gene
  • Do not occur in the germline and therefore cannot be passed on to offspring via the ovum or sperm
  • Almost all malignancies are preceded by a somatic mutation.
• Mosaic (mosaicism): mutation affecting only one population of the body's cells
  • In chromosomal mosaicism, cell populations with different karyotypes are present in one organism.
    • Example: In Turner syndrome; , gonosomal mosaicism is frequently present (several cells with X0; , several cells with XX).
  • Gonadal mosaicism: only some germ cells of an individual carry a mutation
    • Caused by a mutation in the genome of a primordial germ cell during mitosis in germline development
  • Somatic mosaicism: only some of the individual's somatic cells carry a mutation
    • Caused by a mutation during mitosis after fertilization
  • Chimerism: the presence of two genetically distinct cell lines in one individual that originate from two different zygotes; occurs, e.g., when linked oocytes are fertilized.
• Chromosomal instability: caused by mutations in genes that are responsible for DNA repair
  • Results in an increase in chromosomal translations, inversions, and deletions
  • Example: Fanconi anemia, ataxia-telangiectasia
• Two-hit hypothesis: If a mutation is present in one allele of a tumor suppressor gene and the other allele is deactivated by a second mutation, the cell becomes a tumor cell.
  • Example: retinoblastoma
• Loss of heterozygosity (LoH): loss of one allele of a gene after inactivation of the other gene by a mutation

Chromosomal aberrations

Chromosomal aberrations are changes in the genome that are usually visible on karyogram.

• Numerical chromosomal aberrations: altered number of chromosomes
  • Aneuploid: Individual chromosomes are multiplied or absent.
    • Trisomies: trisomy 21, trisomy 13, trisomy 18, trisomy 8, trisomy 9
    • Monosomies: Turner syndrome (45, X0)
    • Polysomies of the gonosomes: Klinefelter syndrome (47, XXY), XYY syndrome, triple X syndrome
  • Polyploid: The entire chromosome set is multiplied, e.g., triploidy (3 x 23 = 69,XXY/XXX).
• Structural chromosomal aberrations: altered chromosome structure with an identical number of chromosomes
  • Detection primarily via FISH
  • Deletion: loss of a chromosome segment, e.g., Cri-du-Chat syndrome (46,XX/XY del(5))
  • Duplication: duplication of a chromosome segment
  • Inversion: inversion of a chromosome segment (e.g., 46,XY,inv(3)(p23q27))
  • Chromosomal translocation: relocation of one chromosome segment onto another (nonhomologous) chromosome
    • Balanced translocation: sum of the genetic material remains identical → no phenotypical alterations
      • Offspring have an increased risk of an unbalanced translocation.
      • E.g., balanced Robertson translocation (45,XY/XX rob(14;21))
    • Unbalanced translocation: The sum of the genetic material is altered.
      • Deletion or duplication of the chromosome segments
      • Normal number of chromosomes, which may have altered phenotype
• Uniparental disomy
  • Offspring has two copies of one chromosome from a single parent and no copies from the other parent.
    • Heterodisomy: two different homologous chromosomes; from one parent passed to offspring → error in meiosis I
    • Isodisomy: identical homologous chromosomes; from one parent passed to offspring → error in meiosis II
  • Should be considered if individual presents with an autosomal recessive condition when only one parent is a carrier
  • Usually results in normal phenotype
  • Uncommon cause of Angelman syndrome and Prader-Willi syndrome (see genomic imprinting)

Uniparental disomy cannot be detected via karyotyping because the number of chromosomes is normal with no loss of genetic material!

Gene mutations

• Various types of gene mutations include:
  • Point mutation: alteration of a single DNA base pair
  • Deletion: loss of one or several base pairs
  • Insertion: addition of one or several base pairs
  • Substitution: One or several base pairs are replaced by one or several different base pairs.
  • Trinucleotide repeat expansion: increased repetition of base triplets that leads to faulty protein synthesis or folding
    • Examples: fragile X syndrome, Huntington's disease, myotonic dystrophy
• Gene mutations can be classified based on the outcome:
  • Frameshift mutation: shift in the reading frame caused by insertion or deletion of a number of nucleotides not divisible by 3, which leads to modified amino acid coding in the gene segments downstream
    • Examples: Tay-Sachs disease, Duchenne muscular dystrophy
  • In-frame deletion or insertion: deletion or insertion of three, six, nine, or more base pairs (always in triplets!), without a shift in the reading frame, but with deletion or insertion of one, two, three, or more amino acids in the protein during translation
  • Silent mutation: altered codon, which codes for the identical amino acids
  • Nonsense mutation: formation of a stop codon, which leads to alterations in the splicing process and early termination of translation
  • Missense mutation: altered codon, which codes for a different amino acid
  • Splice mutation: alterations (especially point mutations) in the nucleotide sequences required for splicing (e.g., on the exon-intron border or at the junction) that lead to defective mRNA and shortened proteins.

Nonsense and frameshift mutations usually change the fundamental structure of the coded protein. For this reason, they typically lead to more severe disease manifestations than missense and silent mutations!

Epigenetics focuses on the heritable chemical modifications of DNA and histone proteins caused by environmental factors. Through these modifications, gene activity can be regulated, i.e., genes can be switched on or off. In contrast to genetic studies, epigenetics is not concerned with the information encoded in the DNA sequence itself.

Main epigenetic mechanisms of the regulation of gene activity

The epigenetic regulation of genes is all about whether or not genes are transcribed. Classical genetics, on the other hand, is concerned with the presence or absence of a gene. Gene expression or repression is determined by chemical modifications of bases (methylation) and histone proteins (various covalent modifications), which are carried out by specialized enzymes.

• DNA methylation: linkage of CH₃ groups with specific cytosine bases of DNA by DNA methyltransferases (DNMTs) → formation of 5-methylcytosine
  • Process: newly synthesized DNA strand is methylated after DNA replication (using the matrix strand as a template)
  • Site: CpG islands
  • Influencing factors
    • Defective DNA repair mechanisms
    • Environmental factors (e.g., arsenic, food substances) that affect DNMT activity
    • Insufficient dietary uptake of methyl donors (e.g., methionine)
    • Cancer cells often contain genes that are activated as a result of defective methylation.
  • Result: Methylation inhibits transcription of the respective gene.
• Histone modification: chemical modification of histone proteins
  • Result: chromatin remodeling → alterations in chromatin structure through histone protein modifications
  • Types of modifications
    • Acetylation
    • Methylation
    • Phosphorylation
    • Ubiquitination
    • Sumoylation
    • ADP ribosylation
  • Examples
    • Acetylation of specific lysine residues in histone proteins catalyzed by histone acetyltransferases → less positively charged histones that do no longer strongly bind to DNA → DNA can be transcribed.
    • Methylation of specific amino acid residues of histone proteins (especially lysines) can lead to increased or decreased transcriptional activity, depending on the amino acid residue on which it occurs.
  • Inheritance: Histone modification pattern and therefore the gene activity state is passed on to daughter cells during cell division!
• Regulatory RNAs: short RNA molecules (e.g., miRNAs) involved in the deactivation of specific target genes at various levels
  • miRNA function: regulation of mRNA degradation and, as a result, inhibition of translation
  • Inactivation of the X chromosome: see below

Epigenetics is concerned with the investigation of external factors that activate or deactivate genes and therefore control transcription!

DNA methylation inhibits transcription; histone methylation can inhibit or enhance transcription!

Typical epigenetic processes

• X inactivation (lyonization)
  • Definition: Inactivation of one of the X chromosomes in individuals with two or more X chromosomes
  • Mechanism: Inactivation occurs on the transcriptional level through regulatory RNAs, Xist RNA (X-inactive specific transcript), and methylation.
    • Which one of the two X chromosomes is inactivated is random.
  • Barr body: the inactivated X chromosome (packaged as heterochromatin); in the cells of a female or individuals with Klinefelter syndrome
  • Inactivation occurs during the 12^th to 16^th day of embryonal development.
• Genomic imprinting
  • Definition: : a mechanism of gene regulation in which one allele of a gene is silenced while the other allele is expressed
  • Mechanism: epigenetic silencing via DNA methylation on one chromosome → depending on the gene, either the maternal or paternal chromosome is inactivated
  • Characteristics
    • In each cell division, information regarding gene activity is passed down to daughter cells. However, this information is deleted during germ cell development and then undergoes sex-specific reprogramming.
    • Genomic imprinting only plays a role in some genes.
  • Examples: Angelman syndrome, Prader-Willi syndrome

References:^[3][4]

Pedigree analysis allows the inheritance patterns of certain traits to be identified. It may, for example, detect evidence of autosomal dominant inherited disorders. In addition to clinical features, an affected gene can be investigated for a mutation or the affected gene's karyotype can be determined.

• Definition: depiction of family relationships with special emphasis on certain phenotypical traits of individual family members
• Aim: to draw conclusions from the phenotype to the genotype of family members and to determine patterns of inheritance
• Form:
  • Symbols indicate individual family members.
  • Lines between the symbols show the degree of the relationship.
  • Generations are indicated with Roman numerals.
  • The children of a generation are designated with Arabic numerals in their order of birth.
  • Affected family members, i.e., carriers, are represented by colored symbols.
• Questions in pedigree analysis:
  1. Dominant or recessive inheritance?
     • Question 1: Does every affected family member have a parent that is also affected?
       • Yes → dominant inheritance of the trait
       • No → recessive inheritance of the trait
  2. Autosomal or gonosomal inheritance?
     • Question 2: Are male family members mainly affected? Yes → (most likely) X-linked recessive inheritance
     • Question 3: If the following criteria are met, the disease (most likely) follows an X-linked dominant pattern of inheritance:
       • Do all affected males also have an affected mother?
       • Do all affected males have healthy sons?
       • Does no affected male have a healthy daughter?
     • If one of these criteria are not met and question 1 has been answered in the affirmative, the disorder is most likely autosomal dominant. If question 1 is answered with a negative response, the disorder is most likely autosomal recessive.

Autosomal dominant inheritance

• Leads to disease even if only one allele is altered
• Usually due to mutations in structural genes
• If one parent is affected, every child has a 50% risk of inheriting the altered allele and therefore the disease.
• Examples: polycystic kidney disease, achondroplasia, Huntington disease, Marfan syndrome, Ehlers-Danlos syndrome, myotonic dystrophy

An autosomal dominant disease with complete penetrance will always manifest in each generation!

Autosomal recessive inheritance

• The disease occurs only if both alleles are altered.
  • Inheritance either via homozygosity for the altered allele or compound heterozygosity
  • Examples: cystic fibrosis, phenylketonuria, autosomal recessive polycystic kidney disease
• Heterozygote (healthy) carriers of a recessive disease are known as carriers and show no phenotypic evidence of the disease.
• If both parents are heterozygote carriers, the offspring have a 25% probability of inheriting the disease, a 50% probability of becoming a disease carrier, and a 25% probability of being unaffected (see table below).

		Heterozygote mother
		v (altered allele)	N (normal allele)
Heterozygote father	v (altered allele)	vv child with disease	Nv carrier
	N (normal allele)	vN carrier	NN healthy child

X-linked recessive inheritance

• The allele responsible for the disease is located on the X chromosome.
• Women are usually carriers and, in rare cases, only affected if both X chromosomes carry the altered allele!
  • Children of a carrier have a 50% probability of inheriting the altered X chromosome.
    • Sons develop the disease, while daughters are carriers
    • The probability that a carrier and healthy father conceive a phenotypically diseased child is 25%.
• Men, whose only X chromosome carries the altered allele, always develop the disease!
  • All daughters inherit the altered X chromosome → carriers
  • All sons inherit the healthy Y chromosome → do not carry any altered allele and are healthy
• Examples: color blindness, hemophilia A/B, G6PD deficiency, Duchenne muscular dystrophy, Becker muscular dystrophy

		Heterozygote mother (carrier)
		x (altered allele)	X (normal allele)
Healthy father	X	Xx carrier	XX healthy daughter
	Y	xY diseased son	XY healthy son

In X-linked recessive inheritance, all male offspring of the affected father (and a healthy mother) are healthy, do not carry the altered allele, and therefore cannot pass on the altered allele!

X-linked dominant inheritance

• The allele responsible for the disease is located on the X chromosome.
• Both men and women are affected.
• Affected mothers have a 50% risk of passing the altered allele on to their offspring (regardless of gender).
• Affected fathers pass the altered allele to all daughters and no sons.
• Examples: Rett syndrome, Alport syndrome, hypophosphatemic rickets

		Heterozygote mother (diseased)
		X (altered allele)	x (normal allele)
Healthy father	xX	Xx diseased daughter	xx healthy daughter
	Y	XY diseased son	xY healthy son

		Heterozygote father (diseased)
		X (altered allele)	Y (normal allele)
Healthy mother	x	Xx diseased daughter	xY healthy son
	x	Xx diseased daughter	xY healthy son

Mitochondrial inheritance

• Mitochondrial DNA is maternally inherited → diseases caused by mutations in mitochondrial DNA are only passed down from mother to offspring
• Any offspring of an affected mother may show signs of the disease.
• Heteroplasmy
  • Each mitochondrion has multiple copies of DNA (mtDNA), and each cell has many mitochondria. Normally, all of the DNA within the mitochondria is the same. If a mutation occurs, it usually is observed only in some of the mtDNA copies; . This heterogeneity among mutated and normal mtDNA is known as heteroplasmy.
  • Disease severity often correlates with the proportion of mutated mtDNA copies.
• Examples: mitochondrial myopathies (e.g., MELAS syndrome), Leber hereditary optic neuropathy

Polygenic inheritance

• A polygenic trait is one that is controlled by the interaction of two or more genes.
• Polygenic traits do not follow Mendelian laws of inheritance.
• Examples
  • Human skin color, eye color, and most anthropometric traits (e.g., height, weight)
  • Many common diseases such as type 1 and type 2 diabetes mellitus, hypertension, androgenic alopecia, atopy, schizophrenia, and Alzheimer disease.

Heterozygote frequency

• Definition: proportion of heterozygote carriers of an altered allele in the population; used to estimate a child's risk of a recessive inherited disorder if the genotype of the parents is unknown
• The heterozygote frequency is calculated in accordance with the Hardy-Weinberg equilibrium: (p+q)² = p² + 2pq + q² = 1 (100%)
  • q = probability of carrying an altered allele
  • q² = probability of carrying two altered alleles (homozygous for the altered allele)
  • p = probability of carrying an unaltered allele
  • p² = probability of carrying two unaltered alleles (homozygous for the unaltered allele)
  • 2pq = heterozygote frequency (probability of carrying both an altered and an unaltered allele)
• Example: cystic fibrosis with an incidence of ∼ 1:2,500
  1. As cystic fibrosis is a recessive disorder, only homozygote carriers develop the disease. The incidence corresponds to the homozygote frequency q².
  2. Calculate q: q² = 1/2500 → q = 1/50 = 2%
  3. Calculate p: (p + q)² = 1 → p + q = 1 → p = 1 – q → p = 1 – 1/50 = 0.98 (98%) ≈ 1
  4. Calculate 2pq: 2pq = 2×1×1/50 = 1/25 heterozygote frequency

References:^[3][5][6]