Regulation of gene expression Part II MOLECULAR BIOLOGY – Regulation of gene expression II

Figure 7-92 Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II There are MULTIPLE opportunities to regulated gene expression (derivation of functional protein) after the initiation of transcription !

MOLECULAR BIOLOGY – Regulation of gene expression II Riboswitch … short sequences of RNA that change their conformation upon binding small molecules such as metabolites, usually reside in 5’ UTR Mostly observed in bacteria but also documented in fungi and plants

Riboswitch can can activate ‘self cleaving’ ribozyme activity MOLECULAR BIOLOGY – Regulation of gene expression II 5’5’ AUG STOP H2NH2N COOH 3’3’ ON AUG H2NH2N COOH 3’3’ OFF 5’5’ small molecule Autolytic ribozyme activity STOP

Figure 7-106c Molecular Biology of the Cell (© Garland Science 2008) Riboswitch can affect the translation initiation from Shine-Dalgarno sequence MOLECULAR BIOLOGY – Regulation of gene expression II

Figure 7-106b Molecular Biology of the Cell (© Garland Science 2008) THERMOSENSOR MOLECULAR BIOLOGY – Regulation of gene expression II

Figure 7-106d Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II Antisense RNA

Figure 7-92 Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II

Figure 7-94 Molecular Biology of the Cell (© Garland Science 2008) ALTERNATIVE SPLICING MOLECULAR BIOLOGY – Regulation of gene expression II

e.g. Drosophila Dscam axon guidance receptors MOLECULAR BIOLOGY – Regulation of gene expression II Thousands of possible different functional protein sequence combinations

Figure 7-96 Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II Splicing is subject to tight regulatory control

Figure 7-92 Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II

Figure Molecular Biology of the Cell (© Garland Science 2008) RNA editing MOLECULAR BIOLOGY – Regulation of gene expression II - specific to eukaryotes (and viruses) Non-templated changes to the genetic information that affect the protein sequence and possibly function and are therefore not predictable from genomic DNA sequence Deamination reactions e.g. A - I/G in RNA duplexes by ADAR: Altered amino acid codons C - U deaminations also possible do not rely on RNA duplexes e.g. ApoB100 mRNA in liver and intestine

Figure Molecular Biology of the Cell (© Garland Science 2008) Extensive RNA editing in mitochondria of trypanosomes MOLECULAR BIOLOGY – Regulation of gene expression II Uracil insertions

Figure 7-92 Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II

Figure 6-80 Molecular Biology of the Cell (© Garland Science 2008) Nonsense-mediated mRNA decay MOLECULAR BIOLOGY – Regulation of gene expression II EJC ribonucleoprotein complexes are remnants of splicing DNA gene transcription EJC displacement during translation Premature termination of translation leaves ‘beacon’ EJC leading to abnormal mRNA degradation

Nanos protein Bicoid protein Some mRNAs are localized to specific regions of the cytoplasm bicoid mRNA nanos mRNA mRNAs for secreted proteins targeted to ER by SRP MOLECULAR BIOLOGY – Regulation of gene expression II Drosophila oocyte polarity and embryo patterning A P bicoid and nanos proteins inhibit translation of other homogenously distributed cell fate genes which in turn affects the expression of other cell fate mRNAs along the A-P axis as development proceeds

Figure 7-92 Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II

PROMOTER AATAAA exon 1exon 2exon 3 ATG TAA AAUAAA AUG UAA AAUAAA AUG UAA intron TRANSCRIPTION Pre-mRNA mRNA DNA RNA SPLICING protein TRANSLATION coding sequence MAPSSRGG….. OPEN READING FRAME 5’UTR3’UTR CAP AAAAA NUCLEUS CYTOPLASM MOLECULAR BIOLOGY – Regulation of gene expression II Central genetic dogma Regulation of gene expression via regulation of mature mRNAs

Figure Molecular Biology of the Cell (© Garland Science 2008) mRNA stability as a way to regulate gene expression in bacteria – half life couple minutes in eukaryotes – half life minutes to tens of hours MOLECULAR BIOLOGY – Regulation of gene expression II poly-A tail: participates in termination of transcription aids mRNA nuclear export protects from 3‘ - 5‘ exonuclease degradation in cytoplasm targets transcripts to ribosome for translation 3’ UTR contains regulatory elements that participate in transcript stability 5‘ Cap: aids mRNA nuclear export protects from 5‘ - 3‘ exonuclease degradation in cytoplasm targets transcripts to ribosome for translation

Figure Molecular Biology of the Cell (© Garland Science 2008) mRNA stability as a way to regulate gene expression in bacteria – half life couple minutes in eukaryotes – half life minutes to tens of hours MOLECULAR BIOLOGY – Regulation of gene expression II Cytoplasmic deadenylases e.g. PARN

Figure Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II PARN binds exposed 5‘ cap of non-translated mRNAs, leading to poly-A tail deadenylation and eventual degradation mRNA’s with longer poly-A tails more likely to be translated Regulatory potential to influence this equilibrium e.g. cytoplasmic polyadenylation of short poly-A tails e.g. fertilized mammalian oocytes and post-synaptic neurone sites

Figure Molecular Biology of the Cell (© Garland Science 2008) MOLECULAR BIOLOGY – Regulation of gene expression II mRNA stability mechanisms in action e.g. ferritin/ transferrin receptor system

PROMOTER AATAAA exon 1exon 2exon 3 ATG TAA AAUAAA AUG UAA AAUAAA AUG UAA intron TRANSCRIPTION Pre-mRNA mRNA DNA RNA SPLICING protein TRANSLATION coding sequence MAPSSRGG….. OPEN READING FRAME 5’UTR3’UTR CAP AAAAA NUCLEUS CYTOPLASM MOLECULAR BIOLOGY – Regulation of gene expression II

Figure Molecular Biology of the Cell (© Garland Science 2008) microRNA (miRNA)... small noncoding RNA regulators recognizing 3’UTRs ● one miRNA recognizes many mRNAs (even hundreds if they share common 3’UTRs) ● more than one miRNA can bind to one mRNA (combinatorial regulation) RNA-induced silencing complex MOLECULAR BIOLOGY – Regulation of gene expression II ● >1000 miRNA genes targeting approx 60% of genes ● estimated 16% of pre-miRNAs can be altered by ADAR mediated editing ● miRNA can be derived from spliced introns in non-mammalian species, known as mirtons Drosha & Prasha (DGCR8) Pri-miRNA Dicer Pre-miRNA RNA induced silencing complex (RISC) miRNA (imperfect duplex 22nts - guide strand incorporated, passenger strand degraded) 3’UTR sequence homology

Figure Molecular Biology of the Cell (© Garland Science 2008) RNA interference (RNAi) small interfering RNA (21-23nt) RISC re-used RNA-induced transcription silencing MOLECULAR BIOLOGY – Regulation of gene expression II Dicer mediated processing dsRNA viral genomes Pre-miRNA transcripts Experimentally introduced RNA n.b. sequence homology not limited to 3’UTR Argonaut mediated target mRNA degradation Co-transcriptional degradation of transcript

Origin of life - RNA world active RNAs involved in: RNA splicing Protein synthesis Expression regulation – siRNA, riboswitches MOLECULAR BIOLOGY – Regulation of gene expression II