Seminario 10 PARTE A Introducción a la Genética Medica - María Paz Bidondo
Introduction to Genotype and Phenotype
Overview of the Seminar
- María Paz Bidondo introduces herself as part of the Molecular Biology and Genetics faculty at the University of Buenos Aires, presenting Seminar 10 focused on genotype-phenotype relationships, medical genetics, and standardized nomenclature for genealogical trees.
- The seminar is divided into two parts; this video covers Part A, focusing on genotype and phenotype concepts.
Defining Genotype
- Genotype is defined as the set of genes in an individual, estimated to be around 20,000 genes in humans, including both nuclear and mitochondrial DNA.
- The classical definition emphasizes gene variants; however, a broader interpretation includes extragenic sequences for association studies.
Reference Sequence Considerations
- To determine genotype accurately, a reference sequence for humans is required. Various versions exist (e.g., version 37 or 38), which are updated based on averages from multiple individuals.
- Alleles compared to this reference are termed wild type (or expected alleles), with caution advised against using "wild" due to potential misinterpretation.
Genotypic Variants Explained
Identifying Homozygosity and Heterozygosity
- By comparing an individual's alleles to the reference sequence, characteristics such as homozygosity or heterozygosity can be identified based on allele similarity.
Example Analysis: Hypothetical Gene P
- A hypothetical gene P located on chromosome one illustrates how individuals can have different allele combinations:
- Pedro: Both alleles are wild type (homozygous).
- Martín: Both alleles have a deletion (homozygous but not wild type).
Distinguishing Between Allele Types
- Pablo has one wild type allele and one with a deletion (heterozygous).
- Juan presents a substitution mutation alongside a deletion; neither allele matches the wild type. This scenario is classified as heterozygous compound due to differing mutations.
Conclusion of Key Concepts
Summary of Genotypic Classifications
Understanding Genetic Variants and Chromosomal Structures
Heterozygous and Homozygous Definitions
- The discussion begins with the identification of a heterozygote possessing one wild type variant and another variant associated with a lesion, alongside a compound heterozygote with two distinct variants.
- The term "homozygous" is introduced, particularly in the context of individuals whose sex chromosomes are not homologous (e.g., X and Y chromosomes).
Chromosome Characteristics
- It is emphasized that non-homologous chromosomes will not share corresponding loci; for example, DAX1 is present on chromosome X but absent on chromosome I.
- Most genes on chromosomes X and I are non-homologous, leading to instances where an individual may be homozygous for alleles found only on one chromosome.
Allelic Representation
- For the gene DAX1 located on chromosome X, only one allele is present since chromosome I lacks this gene. Thus, an individual can be considered homozygous for the wild type variant.
- Similarly, for the SRI gene located solely on chromosome I, individuals will also show homozygosity as there are no corresponding alleles from chromosome X.
Pseudoautosomal Regions
- Two regions marked in red represent pseudoautosomal regions (PAR1 and PAR2), which do exhibit homology between sex chromosomes.
- Genes within these pseudoautosomal regions can have alleles from both chromosomes; thus recombination can occur during meiosis.
Genotype vs Phenotype Analysis
- The importance of understanding genotype versus phenotype is highlighted; while sequencing reveals genotypes, it does not directly inform about phenotypic expressions.
- Techniques such as automated Sanger sequencing or next-generation sequencing are mentioned as methods to analyze genetic sequences effectively.
Real Gene Examples: GJB2 and Sequencing Techniques
Gene Location and Functionality
- The GJB2 gene coding for connexin protein is discussed regarding its specific location on chromosome 13 (long arm Q).
Sequencing Results Interpretation
- Electroferograms from automated sequencing illustrate how different nucleotides (A, T, C, G) are represented by distinct colors in results.
Homozygosity Observations
- An individual homozygous for wild type variants shows consistent peaks in electroferogram readings indicating identical alleles across both copies of the gene.
Deletion Analysis Example
Genetic Variants and Their Impact on Phenotypes
Understanding Genetic Variants
- The speaker discusses the observation of multiple colors in genetic data, indicating variations at specific positions due to missing nucleotides in alleles.
- A specific example is given where an allele has a nucleotide shift, causing discrepancies between two alleles, leading to different peak patterns in genetic analysis.
Heterozygosity and Allelic Variation
- The individual is identified as heterozygous with one allele showing a wild type and another exhibiting a deletion, affecting color representation in genetic sequencing.
- In position 497, adenine is substituted by guanine; this change results in distinct peaks for each nucleotide variant observed.
Genotype vs. Phenotype
- The discussion transitions to defining genotypes (homozygous vs. heterozygous) and how these relate to phenotypic expressions based on environmental interactions.
- The phenotype emerges from the genotype's interaction with environmental factors, which can be measured across various biological levels.
Techniques for Analyzing Phenotypes
- To evaluate phenotypes effectively, techniques beyond sequencing are required; examples include immunofluorescence for protein localization studies.
- Observations of connexin localization using fluorescence microscopy illustrate how proteins express themselves within cellular membranes.
Correlations Between Genotype and Phenotype
- Emphasis is placed on understanding correlations between genotypes and phenotypes within clinical contexts, particularly regarding health conditions.
Genotype and Phenotype: Understanding Blood Groups
Genotype vs. Phenotype
- The genotype refers to specific genetic variants, while the phenotype is the observable manifestation that we aim to measure.
- In the context of blood groups, the ABO system illustrates how genotype influences phenotype through molecular mechanisms and interactions during embryonic and postnatal development.
Blood Group Example
- The ABO blood group system involves glycoproteins present on red blood cells (erythrocytes), which determine an individual's blood type.
- Testing for blood types can be done using a hemagglutination test, where a drop of blood reacts with specific constituents to observe agglutination.
Antigens and Antibodies
- Blood group A has antigen A on its erythrocyte membrane, while group B has antigen B; both are derived from a common precursor known as antigen H.
- Individuals produce antibodies against foreign glycoproteins; for example, group A produces antibodies against group B's glycoproteins.
Genetic Basis of Blood Groups
- The ABO blood groups are determined by a gene coding for a glycosyltransferase enzyme that adds sugars to the basic H protein structure.
- This gene is located on chromosome 9 and exhibits three alleles: A, B, and O. Each individual inherits two alleles (one from each parent).
Allelic Variations
- Possible combinations of alleles in individuals include homozygous AA, BB, OO or heterozygous AB; these combinations affect phenotypic expression.
- The gene consists of seven exons with a total nucleotide count of 165 leading to a protein composed of 355 amino acids.
Amino Acid Differences Among Alleles
- Specific nucleotide variations at position 526 lead to different amino acids in proteins produced by allele A (arginine) versus allele B (glycine).
Understanding Blood Group Genetics
The Role of Glycosyltransferase in Blood Groups
- The translation of the glycosyltransferase protein is crucial for determining blood group phenotypes, which are characterized by specific antibodies.
- There are three different alleles that influence the production of blood group phenotypes. For allele A, the enzyme allows for the transfer of N-acetylgalactosamine to antigens.
- In contrast, allele B produces a different enzyme that transfers galactose instead of N-acetylgalactosamine, leading to distinct blood group characteristics.
- Allele O has a premature stop codon resulting in a non-functional protein, explaining why it lacks additional sugar groups and only presents antigen H.
- The spatial arrangement of glycoproteins on erythrocyte membranes determines antibody reactions, linking genotypes to observable phenotypes.
Genotype and Phenotype Correlation
- Blood group classification (phenotype) is based on glycoprotein presence on erythrocytes and corresponding plasma antibodies; this can be tested through hemagglutination assays.
- Group A individuals have either AA or AO genotypes producing functional acetylgalactosaminetransferase; while group B individuals produce galactosyltransferase.
- Both AA and AO can generate antibodies against type B due to their ability to produce specific glycoproteins despite variations in genotype functionality.
Implications for Genetic Research
- Understanding how residual functional products affect phenotype expression is critical when evaluating genetic conditions related to blood types.
- Current advancements in massive sequencing techniques allow researchers to identify gene variants linked to various phenotypic traits or diseases but often lead into complex interpretations regarding genotype relationships.
Navigating Genetic Variants
- When analyzing multiple gene variants from sequencing data, it's essential to correlate these with observed patient phenotypes (e.g., intellectual disabilities).
- Bioinformatics tools help assess which genetic variants may be pathogenic by correlating them with disease mechanisms after establishing genotype–phenotype links.
Evaluating Pathogenicity
- To determine if a variant contributes to disease, one must consider its population frequency; common variants are less likely involved in rare conditions affecting health outcomes.
Heterozygosity and Homozygosity: Understanding Inheritance Patterns
Clinical Correlation of Genetic Variants
- Heterozygosity and homozygosity are discussed in relation to inheritance patterns, specifically focusing on recessive and dominant traits.
- The importance of family history is emphasized; analyzing relatives can provide insights into genetic variants associated with phenotypes.
- Current international consensus allows for scoring variants to determine their pathogenic potential, classifying them as likely pathogenic or benign based on statistical data.
Variant Classification and Uncertainty
- Variants are classified by probability scores: pathogenic (99%), probably pathogenic (95%), benign (less than 5% chance), or uncertain significance, which creates uncertainty for families.
- The concept of genotype-phenotype correlation introduces complexities such as incomplete penetrance, variable expressivity, and pleiotropy.
Penetrance and Expressivity
- Incomplete penetrance is explained through examples where not all individuals with a specific genotype exhibit the expected phenotype.
- Age-related factors may influence penetrance; older patients may show complete penetrance while younger ones might not fully express the trait.
Variable Expressivity
- Expressivity refers to the degree to which a phenotype manifests among individuals who have the same genotype; it can range from mild to severe symptoms.
Pleiotropy in Genetic Disorders
- Pleiotropy describes how a single genotype can lead to multiple phenotypic expressions across different organs that are not anatomically related.
- An example involving Marfan syndrome illustrates how mutations in fibrillin affect various systems like ocular, cardiovascular, and skeletal structures.
Genotype vs. Phenotype
- The relationship between genotype (specific DNA sequences at loci) and phenotype (observable characteristics influenced by both genetics and environment).
Understanding Phenotypes and Mechanisms
The Concept of Black Box in Genetics
- The discussion revolves around the understanding of phenotypes, where the speaker mentions that one can know or not know the mechanism leading to a specific phenotype.
- When mechanisms are understood, they can be explained clearly, as demonstrated with blood group examples.
- In cases where mechanisms are unknown, this situation is referred to as a "black box," indicating a lack of clarity on how molecular cascades result in certain phenotypes.
- The speaker emphasizes the importance of exploring these black boxes to gain insights into genetic expressions and their outcomes.