Transcript Gene Expression - Western Washington University

This Week
• Chapters 9, 10.1 and 10.4 - 10.7 for reference,
– exam material will be on lecture content for the above,
• Chapter 11.5 (Friday).
• Monday Paper,
Seminars
extra credit
• May 21 “Virgin Birth, Centrosomes, and
Microbes”. William Sullivan, Molecular, Cell, &
Developmental Biology, University of
California, Santa Cruz
• May 28 "Proliferation & Caste Formation in a
Polyembryonic Parasitoid Wasp". Laura
Corley, Department of Entomology,
Washington State University
• Wednesdays, 4:00 PM, BI 212
Gene Expression
…the processes by which information contained in
genes and genomes is decoded by cells,
...in order to produce molecules that determine the
phenotypes observed in organisms,
– transcription (post-transcriptional modifications),
– translation (post-translational modifications).
Transcription
Transcription
...the synthesis of mRNA from a DNA template,
- now it is important to understand when and where,
as well as how.
mRNA Synthesis
• Template (DNA) and Promoter,
• Nucleoside triphosphates (NTPs),
– N: A,U,G,or C,
• Enzymes (RNA polymerases),
• Energy (as in replication, from phosphate bonds).
E. coli RNA Polymerase
RNA Transcription
prokaryote
E. coli Promoter Sequences
concensus sequence alignment
Promoter Regions
Proximal ( ~ 100 - 200)
Core Promoter ( ~ 0 to ~ -40 bp)
Regulation of Transcription
(Prokaryotes)
• Regulation of gene expression is often at the
transcription level,
– Negative regulation,
• inducible,
• repressible,
– Positive regulation
Gene native state is “on”.
Gene native state is “off”.
Negative Regulation
(Inducible)
Protein!
Protein, or metabolite, etc.
…default state is “on”,
i.e., the gene is
transcribed.
Negative Regulation
(Repressible)
Protein!
Protein, or metabolite, etc.
…default state is “on”,
i.e., the gene is
transcribed.
Positive Regulation
…default state is “off”,
i.e., the gene is not
transcribed.
Eukaryotic Initiation
Transcription is
regulated by activators,
repressors, co-activators
and basal transcription
factors.
Promoter Bashing
Modulators of expression can act at great distances.
Drosophila Guts and Such
dpp locus
...(c)LE and (d) ID
enhancer driven,
give rise to fly
appenages.
Terms
• cis-acting elements;
– DNA sequences that
serve as attachments
sites for the DNAbinding proteins that
regulate the initiation
of transcription.
• trans-acting elements;
– the DNA-binding
proteins that regulate
the initiation of
transcription.
Chromatin
Remodeling
Expressed Proteins
Post-Transcriptional Events
• RNA processing,
– splicing,
– poly adenylation,
• Alternate RNA processing.
RNA can be Autocatalytic
• Group I and Group II introns,
– found in mitochondria, chloroplasts, and sometimes in bacteria,
Group II Introns
Eukaryotic Intron Excision
(not autocatalytic)
Eukaryotic Intron Excision
(sequence is important)
Spliceosomes
... small nuclear RNAs (snRNAs):
– RNA molecules that act as catalysts in spliceosomes,
• catalytic RNAs that have probably evolved from ancient RNA
enzymes (ribozymes).
…work in concert with > 100 proteins to facilitate
intron identification and removal,
– snRNPs: RNA/Protein structures.
Psuedouradine (y)
U1 and U2
U1 binds to the 5’ exon/intron junction.
U2 binds to the adenosine at the branch site.
Think about the required specificity for intron identification in cells.
mRNA Processing
splicing
Polyadenylation
AAUAA: concensus poly-A
recognition site.
…complicated biochemistry, lots
of sub-units.
Alternate mRNA Processing
…recognition of different poly-A sites.
…alternate splicing.
Complexity
Calcitonin gene.
Central Dogma
DNA
replication
transcription
RNA
(alternately processed)
translation
Protein
?
Expanded Central Dogma
Genome
Transcriptome
Proteome
•
Genome... the dynamic complement of genetic material in an individual,
•
Transcriptome... mRNA component in an individual,
– complexity increases resulting from transcription control and transcription and post-transcription
modification,
•
Proteome... the protein component of an individual,
– complexity increases due to post-translational modification, protein-protein interactions, etc.
Translation
…the synthesis of a polypeptide. This occurs
on ribosomes using the information encoded
on mRNA,
– tRNA molecules mediate the transfer of
information between mRNA and the growing
polypeptide.
Initiation
complexity
1. Recent reports of
“alternate” translation start
sites indicate that further
complexity in protein
production may occur at the
translational level.
2. Recent reports of
“alternate” small sub-unit
specificity affecting
translation,
- different eIs,
- different small sub- units.
What to Study?
• What regulates gene expression,
• How is gene expression modulated?
– Transcriptionally,
– Post-Transcriptionally,
– Translationally.