GMC Exam #2: Lecture 1

Gene Expression

Signal to nucleus to regulate transcription

Metabolism

Glycogen to glucose (adrenaline response)

Cell growth

Growth factories trigger cell division

Immune Response 

antibodies bind to cell surface antigens 

How are stimuli translated into cellular behaviors

• gene expression

• metabolism 

• cell growth 

• immune response 

Signal transduction 

• can change due to gene expression 

• turn genes on/off 

• turns genes up/down 

• slow: minutes to hours

• transcription factors

Transcription factors

proteins that help find gene and modulation gene expression

2 ways of looking at a gene

1. as a unit encoding a phenotype ( gene expression) 

2. As a unit of inheritance (passed on to future cells/generations) 

Gene

unit of genetic information

genome

database of all genetic information

Features of DNA that make it a good building block for storing genetic information 

1. Universal: 4 nucleotides used across living systems 

2. Very Stable: half life of 521 years 

Structure and function of a DNA double helix (genome)

• Structure: DNA double helix

  • Stability 

  • Replicability 

• Function : stable database of genetic information that can be reliably reproduced 

Structure and function of nucleosome (genome)

• Structure:

  • DNA is wrapped around proteins called histones 

  • DNA + 8 histones = nucleosome 

• Function: 

  • DNA is accessible to inaccessible to proteins 

Structure and function of chromatin fiber (genome)

• Structure: 

  • Chromatosome = nucleosome + 1 linker histone (H1) 

  • folded into chromatin fiber 

• Function: 

  • DNA is tightly packaged — takes up less space 

Structure and function of chromosome (genome)

• Structure: 

  • chromatin fibers further folded and compared into chromosome 

• Function: 

  • tightly coiled to fit into a tight space of nucleus 

  • Completely inaccessible for transcription, ready for mitosis or meiosis 

How do you find genes in a genome?

human genome: 15 of the DNA are genes

• regulatory sequence: modulate gene expression 

• genes can be on either DNA strand 

Regulatory sequences

modulate gene expression

Exons 

sequences that stay in mRNA (coding sequence)

Introns

sequences that are removed from mRNA (not part of coding sequence)

Splicing

process that removes introns and links exons together

Promoter

DNA sequence where transcription machinery binds; tells which DNA strand is to be read and direction of transcription ( determines TSS) 

TSS

transcription start site: first nucleotide to be transcribed

Terminator

DNA sequences ends transcription; usually part of the RNA-coding sequence 

Transcription

the synthesis of an RNA molecule for a template —DNA

What is required for RNA transcription?

1. template — ssDNA 

2. Raw materials — ribonucleotide triphosphate 

3. enzyme and other proteins — transcription machinery: proteins necessary for catalyzing the synthesis of RNA 

Nucleic acid polymerization

Nucleotide addition and 3’ —OH end of growing chain, leaving 2 phosphates behind

Nucleotide addition

added one at a time to the 3’ —OH groups of the growing RNA molecule-thus direction of the transcription is 5’ to 3’ 

Bacterial RNA polymerase

synthesizes all classes of bacterial RNA

• 5 subunits + sigma factor 

Eukaryotic RNA polymerase

responsible for synthesizing different classes of RNA

• many accessory proteins necessary 

RNA polymerase 1

• Present in: all eukaryotes 

• Transcribes: large rRNAs 

RNA polymerase II

• Present in: all eukaryotes 

• Transcribes: pre-mRNA, some snRNAs, snoRNAs, some miRNAs 

RNA polymerase III

Present in: all eukaryotes

transcribes: tRNAs, small rRNAs, some snRNAs, some miRNAs 

Initiation

Transcription apparatus assembles on the promoter and begins the synthesis of RNA

1. promoter recognition 

2. formation of a transcription bubble 

3. creation of the first bonds between rNTPs 

4. escape of the transcription apparatus form the promoter 

Elongation

DNA is threaded though RNA polymerase; polymerase unwinds the DNA and adds new nucleotides, one at a time, to the 3’ end of the growing RNA strand; rewinds the DNA at the trailing edge 

Termination

Recognition of the end of the transcription unit and the separation of the RNA molecule for the DNA template — terminator is transcribed 

Initiation: Where do we begin? How does a cell know where a gene begins on the template?

• Promoter recognition

  • RNA polymerase sits down on the promoter

  • aided by transcription factors

  • DNA sequence is recognized and bounded by transcription apparatus

    • determines TSS: first nucleotide to be transcribed based on distance from consensus sequence

Prokaryotic Promoter

conserver -35 and -10 sequences

Eukaryotic Promoters

• Regulatory promoters

  • upstream of core 

  • mixed and matched consensus sequence 

    • regulate teh transcription rate e.g. enhancer 

• Core Promoter: 

  • immediately upstream of gene TSS 1

  • 1 or more cones sequences 

Promoter recognition

1. TFIID transcription factor and TBP bind to TATA box = 11-15bp DNA unwinding 

2. Other TFs bind to their consensus sequences and RNA pol  

3. More DNA unwinding = ssDNA template 

RNA Pol II

Transcription polymerase

TBP

TATA binding protein

General transcription factors (TFs)

bind to DNA and modulate level of transcription

Formation of transcription bubble

a short stretch (~18 bp) of unwound DNA

Creation of the first bonds between rNTPs

Phosphodiester bond formation polymerase synthesizes RNA molecule 9-12 bp in length = polymerase transitions to the elongation stage 

Escape of the transcription apparatus

change in its shape = can’t bind promoter

Elongation: how do we read DNA to create RNA?

• read/transcribe template strand by synthesizing mRNA in 5’ to 3’ (RNA Pol on template) 

• Phosphodiester bond formed 

  • reaction: 3’ OH and a phosphate 

• Transcription bubble keeps moving 

  • polymerase unwinds DNA and adds new nucleotides to the 3’ end of the growing RNA strand 

  • DNA rewinds behind bubble 

  • multiple bubbler ber gene 

Termination: where do we stop?

• Transcribe until terminator sequence 

• terminator sequence: forms hairpin loops in RNA 

• causes RNA polymerase to fall off DNA 

Transcription is selective

only certain parts of the DNA are transcribed at a time