Lecture 4 Topic 2

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Lecture 4 Topic 2
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Gene Function Gene Expression
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Topic 2 Material and Chapter 3

• Not Covering
• Transcription
• Transcription Termination in Bacteria
• Already covered
• Gene Structure
• component parts of gene, template and
  non-template strand, mRNA-like strand, 5 and 3
  end of gene
• Will cover in detail later
• Transcription
• Initiation of transcription, promoter structure,
  RNA polymerase structure and function
• Will cover now
• steps in transcription, details of eukaryotic
  transcription termination, RNA processing,
  translation, protein structure and function as it
  relates to gene function and mutant phenotypes

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Transcription Termination and mRNA 5-End and
3-End-Formation
Steps of Transcription RNA processing during
transcription - 7mG capping 5 end - 3 end
formation 5 CAP 3 Poly A Tail
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RNA Splicing

• RNA splicing reactions
• Two transesterification reactions
• first cleaves at 5 end of intron (5 splice
  site)
• second cleaves at 3 end of intron (3 splice
  site)
• Self-Splicing RNAs (discovery of ribozymes)
• Group I and Group II introns remove themselves
• difference is in the first transesterification
  (see later)
• Group I introns use a free G nucleotide to
  catalyze reaction
• Group II splicing is similar reaction to that in
  pre-mRNA splicing
• pre-mRNA Splicing
• catalyzed by spliceosome
• multiple complexes of proteins and RNA
  (ribonucleoprotein particles called snRNPs
• snRNAs called U RNAs (U1,2,4,5 and 6)
• splicing proteins
• snRNPs bind to pre-mRNA in a specific sequence to
  catalyze splicing

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Transesterification Reactions in RNA Splicing

• Attack of the sugar-phosphate bond at 5 splice
  site
• Group 1 3 OH of free G nucleotide
• Group 2 and pre-mRNA 2 OH group of branch
  point A nucleotide
• Attack of the sugar phosphate bond at the 3
  splice site
• 3 OH of nucleotide at 3 end of exon 1
• Intron released
• Group 1 introns linear
• Group 2 and pre-mRNA introns lariat
• Base pairing between U snRNAs and intron
  sequences brings intron sequences together and
  catalyzes the transesterification reactions

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(No Transcript)
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Identification of Intron Sequences important for
Splicing
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U snRNApre-mRNA base pairing
2. Positioning of A nucleotide for
transesterification reaction 1
1. Identification of 5 splice site and Branch
Point
3. Positioning of 3 splice site near exon1 for
transesterification reaction 2
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Summary of terms and processes
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Mutations affecting Splicing

• Use mutation in globin genes to illustrate
  effects of mutation on gene function
• splicing mutations
• mutations of splice sites
• mutation in exons creating cryptic splice site
• reduction in protein levels
• production of abnormal protein
• mutations in coding sequence
• reduction in protein levels
• production of abnormal proteins

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Genetic Disorders of Hemoglobin
Hemoglobinopathies
hemoglobinopathy ltdisease, hematologygt Disorder
due to abnormalities in the hemoglobin molecule,
the best known being sickle cell anemia in which
there is a single amino acid substitution (valine
for glutamate) in position 6 of the beta chain.
In other cases one of the globin chains is
synthesised at a slower rate, despite being
normal in structure.
Hemolytic anemias (RBC lysis) Thalassemias
(decreased amount to to unbalanced synthesis of
one globins) Methemoglobinemias (reduced oxygen
transport due to defective hemoglobin)
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Genetic Disorders of Hemoglobin
Hemoglobinopathies
Clinical Importance 5 world population carry
alleles causing hemoglobinopathies! Model for
determining the molecular basis of disease One
of first protein structures determined First
disease related gene to be cloned Model for the
structure, regulation, and function of human genes
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Structure and Function of Hemoglobin
Tetramer 2 alpha-like globins 2 beta-like
globins 4 heme groups (1/globin) Require equal
amounts of alpha and beta globins for proper
Hb concentration in RBCs Mutations in beta
globin splice sites reduce amount of hemoglobin
resulting in beta thalassemia
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Mutations in beta thalassemia
Mutations in splice sites - reduce usage of
proper splice site - no splicing or use of a
cryptic splice site Mutations in introns or
exons - creates new splice site - competes with
proper site - reduces usage of proper splice site
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Splicing mutations causing beta thalassemia