Structure of Genes 2024 PDF
Document Details
Uploaded by Deleted User
2024
Tags
Related
Summary
These lecture notes cover the structure of genes, transcription, translation, and gene expression in prokaryotes and eukaryotes. The notes detail the central dogma, gene organization, and the differences in gene expression between prokaryotes and eukaryotes.
Full Transcript
Lecture 2 BIOL2004-5-6 The Structure of genes Learning Outcomes 1 – Recap on central dogma – transcription and translation 2 – Compare and contrast gene expression In Prokaryotes and Eukaryotes 3 – Consider intron removal and alternative spl...
Lecture 2 BIOL2004-5-6 The Structure of genes Learning Outcomes 1 – Recap on central dogma – transcription and translation 2 – Compare and contrast gene expression In Prokaryotes and Eukaryotes 3 – Consider intron removal and alternative splicing 4 – Understand how the above relates and implications for the practical ‘cloning of MLF1’). Transcription and translation : the central dogma Replication The central dogma DNA of DNA describes Double-stranded (ds) a flow of information in an (4 nucleotides: A, C, G, T)... A 5’ T G C C C C G T T G A...3’ organism... 3’ T A C G G G G C A A C A... 5’ Transcription of genes DNA RNA Single-stranded (ss)... A 5’ U G C C C C G U U G A...3’ (4 nucleotides: A, C, G, U) Stop codon RNA Translation of mRNA Protein Single-stranded +NH3 Met Pro Arg COO- Protein (20 amino acids) Gene organisation in bacteria In eukaryotes genes are transcribed as single units however in bacteria genes involved in the same pathways are often arranged linearly as an operon When the operon is transcribed all of the genes are transcribed in one continuous mRNA known as a polycistronic mRNA This increases the efficiency of gene regulation – for instance if the bacterial cell needs to make biotin it makes sense to transcribe all of the genes involved in this process together –means there can be a simple genetic switch to turn the process on. The structure of eukaryotic genes Quizlet.com Eukaryotic genes are more complex than those of prokaryotes Typically the open reading frame (ORF) which is composed from the exons within the gene is split by intervening non-coding DNA in the form of introns (not always) In order to be expressed a transcript from a eukaryotic gene has to be processed to remove the introns Overview of gene expression Despite the universality of the central dogma, there are important variations in the The central dogma way information flows from DNA to protein between prokaryotes and Eukaryotes. describes a flow of information in an organism DNA RNA Protein Overview of gene expression A - gene expression in bacteria B - gene expression in eukaryotic cells Coupled transcription and translation in prokaryotes Because prokaryotes have their genomic DNA naked in the cytoplasm (no nucleus) and doesn’t require polyadenylation or intron removal, transcription and translation are coupled Translation begins as the mRNA molecule is being transcribed, note that a number of genes in the same operon are being transcribed together from the same mRNA (polycistronic). On the right is a section of the bacterial genomic DNA the dark areas are ribosomes attached to mRNA molecules perpendicular to the genomic DNA Eukaryotic mRNA is transported out of the nucleus Because the eukaryotic cell has a nucleus – mRNA needs to be transported into the cytoplasm to be translated into protein by the ribosome mRNAs are marked for export by the interaction of specific proteins: Cap-binding protein Poly-A-binding protein Exon junction complex (EJC) Gene expression in eukaryotes Summarising this - the differences between gene expression in eukaryotes and prokaryotes include: 1. eukaryotic transcripts need to be transported from the nucleus where they are transcribed to the cytoplasm where they are translated – uncoupled 2. many eukaryotic genes contain untranslated sequences – introns – these need to be removed before translation 3. eukaryotic transcripts are polyadenylated at the 3’ end 4. mRNA transcripts are capped at 5’ end 5. eukaryotic transcripts are monocistronic – they do not contain the transcripts of several genes Gene structure – the core promoter Directs initiation of transcription concentrating on RNA Polymerase II – core promoters have conserved general features: 1. TATA Box – consensus 5’ TATAWAAR -3’ (W = A or T, R = A or G): -25 bases and/or 2. Inr (initiator) (mammalian consensus – 5’-YCANTYY-3’ (Y = C or T, N= any): ~ +1 bases 3. DPE (downstream promoter element) – variable sequence; +28 to +32 (binds TFIID) 4. 7bp GC Box – immediately upstream of TATA box (binds TFIIB) 5. PSE (only in snRNA genes) Transcription of ribosomal genes Transcribes all eukaryotic protein-coding genes transcribes DNA to synthesize ribosomal 5S rRNA, tRNA and other small RNAs Gene structure – the upstream promoter elements typical yeast promoter Elements located upstream of the core promoter. typical promoter Allow interaction from a ‘higher’ with RNA eukaryote polymerase The complexity of promoter elements varies hugely in eukaryotes Simple single-celled eukaryotes commonly have promoters that are simple and proximal to the gene itself whilst multicellular organisms may have regulatory elements many kilobases away from the gene itself Builds on lac operon from last year, simple in yeast then more complex in other eukaryotes, start with yeast the galactose regulon Introns are removed in the nucleus A mixture of small nuclear RNA molecules (snRNA) and proteins are responsible for the excising of introns and joining together the 3’ and 5’ ends of exons to form the mature transcript This only happens in eukaryotes Sometimes the exons of genes with multiple introns can be spliced together in different ways – (alternative splicing) – this means that the coding sequence of the transcripts can differ and results in different proteins which may be involved in different processes Alternative splicing of introns Allows a single gene to produce multiple proteins, which have different functions – see subunits in proteins Alternative splicing of introns Brining it back to your practical… Reminder - The aim is to amplify, clone, sequence and express in E. coli a protein called MLF1 (Myeloid Leukaemia Factor 1) DNA RNA Protein https://www.phosphosite.org/proteinAction?id=14020&showAllSites=true http://cancer.sanger.ac.uk/cosmic3d/protein/MLF1 In terms of the human genome and your practical… You can look at the position and structure of any gene using ENSEMBL (https://www.ensembl.org/index.html) From the whole chromosome view (top), regional view (middle) to view the structure of the gene and its transcripts (bottom) MLF1 and alternate splicing Problems: Cannot use genomic DNA (Why??) So what do we use instead? (https://www.ensembl.org/index.html) Todays practical Week 2 – Making RNA, quantifying RNA and designing primers Allows cDNA (without introns / modification / splicing issues To enable polymerase chain reaction – to amplify DNA Reminder - The aim is to amplify, clone, sequence and express in E. coli a protein called MLF1 (Myeloid Leukaemia Factor 1)