Genetics and Evolution: Molecular Basis of Inheritance - Eukaryotic Ribosomes

Introduction to Eukaryotic Ribosomes

  • Ribosomes are cellular structures responsible for protein synthesis.
  • Eukaryotic ribosomes are larger and more complex compared to prokaryotic ribosomes.
  • Composed of two subunits: small (40S) and large (60S) subunits.
  • Found both in the cytoplasm and attached to the endoplasmic reticulum (ER).

Structure of Eukaryotic Ribosomes

  • Small Ribosomal Subunit (40S):
    • Composed of one RNA molecule and proteins.
    • Contains binding sites for mRNA and tRNAs.
    • Stabilizes the complex during translation initiation.
  • Large Ribosomal Subunit (60S):
    • Composed of three RNA molecules and proteins.
    • Forms the peptidyl transferase center for peptide bond formation.
    • Acts as a catalyst in protein synthesis.

Role of Eukaryotic Ribosomes in Protein Synthesis

  • Protein synthesis occurs in two main steps: transcription and translation.
  • Ribosomes are involved in translation, which is the process of synthesizing proteins from mRNA.
  • Translation Steps:
    1. Initiation: Ribosomes bind to mRNA and identify the start codon.
    2. Elongation: Amino acids are added to the growing polypeptide chain.
    3. Termination: Ribosome reaches a stop codon, and protein synthesis is terminated.

Ribosomal RNA (rRNA)

  • RNA molecules that are part of the ribosome structure.
  • Three main types of rRNA in eukaryotes:
    1. 18S rRNA: Found in the small 40S subunit.
    2. 5.8S rRNA: Found in the large 60S subunit.
    3. 28S rRNA: Also found in the large 60S subunit.
  • rRNA molecules provide a scaffold for ribosomal proteins and contribute to the catalytic activity of the ribosome.

Ribosomal Proteins

  • Numerous proteins are associated with ribosomal RNA.
  • Proteins help stabilize the overall structure of ribosomes.
  • Different ribosomal proteins have distinct functions, including:
    • Mediating interactions with tRNA and mRNA.
    • Facilitating the formation of the peptide bond during protein synthesis.
    • Assisting in the proper folding of nascent polypeptides.

Ribosome Biogenesis

  • Ribosomes are synthesized through a complex process known as ribosome biogenesis.
  • Ribosome biogenesis involves the production and assembly of ribosomal RNA and ribosomal proteins.
  • Steps in Ribosome Biogenesis:
    1. Transcription of rRNA genes.
    2. Processing and modification of rRNA.
    3. Assembly of ribosomal subunits.
    4. Transport of ribosomal subunits to the cytoplasm.
  • Ribosome dysfunction can lead to various human disorders, including:
    1. Diamond-Blackfan anemia: Reduced production of red blood cells due to ribosome abnormalities.
    2. Shwachman-Diamond syndrome: Affects the bone marrow, pancreas, and skeletal system.
    3. Cartilage Hair Hypoplasia: Characterized by skeletal abnormalities and short stature.

Antibiotics Targeting Ribosomes

  • Some antibiotics target the ribosomes of bacteria, leading to interference with protein synthesis.
  • Examples of antibiotics that target ribosomes include:
    • Tetracycline: Inhibits aminoacyl-tRNA binding.
    • Chloramphenicol: Inhibits peptidyl transferase activity.
    • Erythromycin: Interferes with the movement of tRNA during protein synthesis.
    • Streptomycin: Inhibits the initiation of protein synthesis.

Summary

  • Eukaryotic ribosomes play a crucial role in protein synthesis.
  • Composed of small and large subunits, consisting of rRNA and ribosomal proteins.
  • Ribosome biogenesis involves the production and assembly of ribosomal components.
  • Ribosome dysfunction can lead to various human disorders.
  • Antibiotics can target ribosomes and inhibit bacterial protein synthesis.

Role of Eukaryotic Ribosomes in Cellular Functions

  • Apart from protein synthesis, ribosomes have additional functions:
    • Regulatory roles in gene expression.
    • mRNA quality control, ensuring proper removal of faulty transcripts.
    • Ribosome-associated molecular machinery regulates translation speed and protein folding.
    • Signal recognition particle (SRP) helps guide proteins to the endoplasmic reticulum for membrane insertion.
  • Ribosomes have a dynamic nature, allowing for diverse cellular functions beyond protein synthesis.

Ribosome Structure and Antibiotics

  • Antibiotics target bacterial ribosomes, exploiting differences from eukaryotic ribosomes.
  • Different binding sites and protein structures between bacterial and eukaryotic ribosomes make antibiotics selective.
  • Some examples of antibiotics and their mode of action:
    • Aminoglycosides (e.g., streptomycin): Bind to bacterial ribosomes, causing misreading of the genetic code.
    • Macrolides (e.g., erythromycin): Prevent elongation of the polypeptide chain during translation.
    • Chloramphenicol: Prevents peptide bond formation.

Riboswitches: RNA-Based Gene Regulation

  • Riboswitches are regulatory elements found in bacterial mRNA.
  • They are made up of a folded RNA sequence that can change its structure upon binding to specific molecules.
  • Types of riboswitches and their functions:
    • Metabolite-binding riboswitches: Bind to small molecule ligands, regulating gene expression.
    • Amino acid-binding riboswitches: Control production of enzymes involved in amino acid biosynthesis.
  • Riboswitches offer a unique mechanism for gene regulation through direct binding of ligands to mRNA.

Translation Initiation in Eukaryotes

  • In eukaryotes, translation initiation is a complex process involving multiple factors:
    • Small ribosomal subunit binds to the 5’ cap of mRNA.
    • Scanning mechanism identifies the start codon (usually AUG).
    • Large ribosomal subunit joins and translation begins.
  • Translation initiation ensures accurate and efficient protein synthesis.
  • Example: Upstream Open Reading Frames (uORFs) regulate the translation of main open reading frames (ORFs) by affecting ribosome scanning efficiency.

Ribosome Slippage and Frameshifting

  • Ribosome slippage and frameshifting are rare events, but they play important roles in gene expression regulation.
  • Ribosome slippage:
    • Can occur during translation of specific mRNA sequences.
    • Leads to a shift in the reading frame, resulting in the synthesis of a different protein.
  • Frameshifting:
    • Shifts the reading frame of the ribosome by one or two nucleotides.
    • Often triggered by specific RNA sequences or secondary structures.
  • These mechanisms allow for the production of multiple proteins from a single mRNA.

Ribosome Profiling: A Tool for Studying Translation

  • Ribosome profiling (or Ribo-seq) is a technique used to study translation at a genome-wide scale.
  • It involves mapping the position of ribosomes along mRNA molecules.
  • Steps in ribosome profiling:
    1. Cells are treated with a ribosome stalling agent, which halts ribosomes at their current positions.
    2. Ribosomes are digested away, leaving behind protected segments of mRNA.
    3. Sequencing and analysis of these segments provide information about translation dynamics.
  • Ribosome profiling has revolutionized our understanding of translation and gene expression.

Ribosome Heterogeneity

  • Ribosomes are not uniform in structure and composition; they exist in multiple forms.
  • Different ribosome populations have unique functions in specific cellular contexts.
  • Examples of ribosome heterogeneity:
    • Free ribosomes: Involved in cytosolic protein synthesis.
    • Membrane-bound ribosomes: Synthesize proteins destined for secretion or membrane insertion.
    • Stress granule-associated ribosomes: Play a role in cellular stress response.
  • Ribosome heterogeneity allows for specialization and fine-tuning of protein synthesis.

Ribosome Recycling and Quality Control

  • After completing translation, ribosomes need to be recycled for future rounds of protein synthesis.
  • Steps in ribosome recycling:
    1. Release of the completed polypeptide chain.
    2. Dissociation of the small and large ribosomal subunits.
    3. Recycling factors assist in disassembling the ribosomal complex.
    4. Individual ribosomal subunits are ready to initiate translation again.
  • Quality control mechanisms monitor ribosome fidelity, eliminating any error-prone or aberrant complexes.

Ribosome-targeting Diseases and Therapeutic Approaches

  • Ribosome-targeting diseases result from dysregulation or malfunction of ribosomes.
  • Therapeutic approaches for ribosomal disorders are being explored.
  • Examples of ribosome-targeting diseases:
    • Diamond-Blackfan anemia: Caused by mutations affecting ribosomal proteins.
    • Shwachman-Diamond syndrome: Mutations in a gene involved in ribosome biogenesis.
  • Potential therapeutic approaches include restoring ribosome function or compensating for ribosome dysfunction using targeted therapies.

Conclusion

  • Eukaryotic ribosomes are essential cellular components responsible for protein synthesis.
  • Ribosomes play diverse roles beyond translation, including gene regulation and cellular processes.
  • Antibiotics target bacterial ribosomes, exploiting differences from eukaryotic ribosomes.
  • Ribosome profiling and studies on ribosome heterogeneity have uncovered new insights into translation dynamics.
  • Ribosome-related disorders are being investigated for their mechanisms and potential therapeutic interventions.

Translation and tRNA

  • Transfer RNA (tRNA) functions as an adapter molecule during translation.
  • tRNA carries amino acids to the ribosome and matches them with their corresponding codons on mRNA.
  • Key features of tRNA:
    • Unique three-dimensional structure with an amino acid attachment site (3’ end) and an anticodon loop.
    • Anticodon loop base pairs with the complementary codon on mRNA.
    • Each tRNA is specific for a particular amino acid.
  • tRNA plays a crucial role in accurate and efficient protein synthesis.

Initiation Factors in Translation

  • Initiation factors are proteins that assist in the initiation phase of translation.
  • They help assemble ribosomes at the start codon of mRNA.
  • Examples of initiation factors:
    • eIF1: Enhances the accuracy of initiation and prevents premature ribosome binding.
    • eIF1A: Promotes the binding of initiator tRNA to ribosomes.
    • eIF2: Provides the GTPase activity needed for translation initiation.
  • Initiation factors ensure proper start codon recognition and initiation of protein synthesis.

Elongation Factors in Translation

  • Elongation factors are proteins that facilitate the elongation phase of translation.
  • They help coordinate the addition of amino acids to the growing polypeptide chain.
  • Examples of elongation factors:
    • EF-Tu: Delivers aminoacyl-tRNA to the ribosome.
    • EF-Ts: Catalyzes the exchange of GDP for GTP on EF-Tu.
    • EF-G: Translocates the ribosome along the mRNA during elongation.
  • Elongation factors ensure accurate and efficient addition of amino acids to the growing polypeptide chain.

Termination of Translation

  • Termination is the final phase of translation when protein synthesis is completed.
  • It occurs when a stop codon is reached on mRNA.
  • Key components of termination:
    • Release factors (RFs): Recognize stop codons and promote the release of the completed polypeptide chain.
    • GTP hydrolysis: Accompanied by release factor binding, triggers the termination process.
    • Dissociation of ribosomal subunits: After peptide release, ribosomal subunits dissociate from mRNA.
  • Termination ensures the accurate and timely completion of protein synthesis.

Ribosome Profiling and Translation Efficiency

  • Ribosome profiling can provide insights into translation efficiency and transcriptome-wide protein synthesis.
  • It involves measuring the density of ribosomes along mRNA molecules.
  • Ribosome profiling allows for:
    • Quantification of translation dynamics.
    • Examination of codon usage bias and ribosome stalling.
    • Identification of open reading frames and translated regions.
  • Understanding translation efficiency can shed light on gene expression regulation and protein synthesis.

Translation and Gene Regulation

  • Translation is a critical step that can regulate gene expression post-transcriptionally.
  • Mechanisms for translational control:
    • Translational repression: Inhibits translation initiation or elongation.
    • Translational activation: Enhances translation initiation or elongation.
    • MicroRNAs (miRNAs): Bind to mRNA, leading to degradation or translational repression.
  • Translational regulation allows cells to fine-tune protein levels in response to various stimuli.

Co-Translational Protein Folding

  • Co-translational protein folding is the process by which nascent polypeptide chains fold into their functional structures during translation.
  • Factors influencing co-translational protein folding:
    • Ribosome-associated chaperones: Assist in proper folding and prevent misfolding.
    • Translation rate: Optimal translation speed allows for correct folding.
    • Ribosome tunnel environment: Provides space for folding and prevents misfolding.
  • Co-translational protein folding ensures the correct conformation and functionality of newly synthesized proteins.

Codon Bias and Translation Efficiency

  • Codon bias refers to the preference for certain codons encoding the same amino acid in different organisms or genes.
  • Codon usage can impact translation efficiency and protein expression levels.
  • Factors influencing codon bias:
    • tRNA availability: Abundance of specific tRNAs affects the speed and accuracy of translation.
    • Mutation bias: Accumulation of mutations in the genome over time can influence codon usage.
    • Selection pressure: Adapting to environmental conditions or optimizing protein production.
  • Codon bias plays a role in fine-tuning translation and protein synthesis.

Translational Control in Development and Disease

  • Translational control plays a crucial role in organismal development and is dysregulated in many diseases.
  • Examples of translational control in development:
    • Embryonic development: Temporally regulated translation of specific mRNAs guides different stages of development.
    • Neuronal development: Precise control of translation is essential for synapse formation and plasticity.
  • Dysregulated translational control is associated with various diseases, including cancer, neurodegenerative disorders, and metabolic diseases.

Summary

  • Translation is the process by which mRNA is decoded to synthesize proteins.
  • tRNA acts as an adapter, matching amino acids to their corresponding codons on mRNA.
  • Initiation, elongation, and termination are the main steps in translation.
  • Initiation factors assemble ribosomes at the start codon.
  • Elongation factors facilitate the addition of amino acids to the growing polypeptide chain.
  • Termination factors recognize stop codons and release the completed polypeptide.
  • Ribosome profiling provides insights into translation efficiency and dynamics.
  • Translation is regulated at multiple levels, influencing gene expression and protein synthesis.
  • Co-translational protein folding ensures proper protein conformation and functionality.
  • Codon usage and translational control play important roles in development and disease.