RNA: Posttranscriptional

Modifications

By

Amr S. Moustafa, M.D.; Ph.D.

Enhancers and Silencers

Cis-acting DNA sequencesEnhancers: stimulate; Silencers: inhibitUpstream or downstreamClose or thousands bp from promoter

DNA response elements: activators-binding sitesOn either strand of DNA

Bending of enhancer DNA allows activators(specific TF) to interact with:

General TF and RNA polymerase II

Enhancers and Silencers

Cis-acting DNA sequencesEnhancers: stimulate; Silencers: inhibitUpstream or downstreamClose or thousands bp from promoter

DNA response elements: activators-binding sitesOn either strand of DNA

Bending of enhancer DNA allows activators(specific TF) to interact with:

General TF and RNA polymerase II

Transcription

Factors

General:Binds to promoter

Activators:Binds to Enhancer

The Lac OperonOperon:

Coordinately expressed genesStructural and their regulatory genes

Lac operon: (For E. coli catabolism of lactose)

Structural genes: code for 3 enzymes:Lac Z gene: β-galactosidaseLac Y: permeaseLac A: thiogalactoside transacetylase

Regulatory Sequences: Promoter (P) region Operator (O) site CAP-binding site LacI gene: repressor protein

The Lac OperonOperon:

Coordinately expressed genesStructural and their regulatory genes

Lac operon: (For E. coli catabolism of lactose)

Structural genes: code for 3 enzymes:Lac Z gene: β-galactosidaseLac Y: permeaseLac A: thiogalactoside transacetylase

Regulatory Sequences: Promoter (P) region Operator (O) site CAP-binding site LacI gene: repressor protein

The Lac Operon: Off

The Lac Operon: On

The Lac Operon: Off

Posttranscriptional Modifications

The rRNA and tRNA:Both pro- and eu-karyotes

The mRNA:Only eukaryotes

Posttranscriptional

Modifications: The rRNA

Ribonucleases

Site: Nucleolus

Protein association: Before and during modifications

e 5 S rRNA:

Separate modification

Posttranscriptional Modifications: The

tRNA - 11. Ribonucleolytic cleavage:

16-Nucleotides at 5’-end2 Uracil at 3’-end14 Nucleotides (intron) at anticodon loop

2. Addition of CCA at 3’-endNucleotidyltransferase

3. Base modifications

Posttranscriptional Modifications: The

tRNA - 2

Posttranscriptional Modifications: The

euokaryotic mRNA - 1

1. 5’- Capping: 7-Methyl-guanosine

2. 3’- Poly-A tail

3. Removal of introns and splicing of exons

Posttranscriptional Modifications: The

euokaryotic mRNA - 2Cap: 7-Methylguanosine triphosphate

Bond: Unusual 5’-5’ triphosphate

Site: Nucleus & cytoplasm

Enzymes: Nuclear guanylyltransferase Cytoplasmic methyl transferase Methyl donor: SAM

Functions: Initiation of translation Stabilizes mRNA

Posttranscriptional Modifications: The

euokaryotic mRNA - 33’- Poly-A tail: 40 – 200Exception: mRNA for histones and interferons

Enzyme: Nuclear polyadenylate polymerase

Site: Nucleus & cytoplasm

Signal: AAUAAA

Functions:Stabilizes mRNAFacilitates its exit from nucleus

Poly-A tail: Shortened after mRNA enters cytosol

Posttranscriptional Modifications: The

euokaryotic mRNA - 4

Posttranscriptional Modifications: The

euokaryotic mRNA - 5Removal of introns and splicing of exons:

Exons: Coding sequences for proteins

Spliceosome: Binding of snRNPs & primary transcriptCorrect alignment of neighboring exonsallows splicing of exons

The snRNPs: snRNAs + proteins

Introns: Sequences not coding for proteinsNo, few or many introns/mRNA

Posttranscriptional

Modifications: The

euokaryotic mRNA - 5

Systemic lupus Erythematosus:Auto Abs against snRNPs

β-Thalassemia:Splice site mutations

Posttranscriptional

Modifications: The euokaryotic

mRNA - 6

Alternative splicing:A diverse set of proteins from a small set of genes

Selective tissue expressione.g., tissue-specific tropomycin