Introns and Exons DNA is interrupted by short sequences that are not in the final mRNA Called introns Exons = RNA kept in the final sequence

Gene Regulation Ability of an organisms to control which genes are present in response to the environment

Prokaryotic cells turn genes on and off by controlling transcription. A promotor is a DNA segment that allows a gene to be transcribed. An operator is a part of DNA that turns a gene “on” or ”off.” An operon includes a promoter, an operator, and one or more structural genes that code for all the proteins needed to do a job. Operons are most common in prokaryotes. The lac operon was one of the first examples of gene regulation to be discovered. The lac operon has three genes that code for enzymes that break down lactose.

The lac operon acts like a switch. The lac operon is “off” when lactose is not present. The lac operon is “on” when lactose is present.

Trp operon lac operon

Points of control The control of gene expression can occur at any step in the pathway from gene to functional protein 1. packing/unpacking DNA 2. transcription 3. mRNA processing 4. mRNA transport 5. translation 6. protein processing 7. protein degradation

1. DNA packing as gene control Degree of packing of DNA regulates transcription tightly wrapped around histones no transcription genes turned off heterochromatin darker DNA (H) = tightly packed euchromatin lighter DNA (E) = loosely packed H E

DNA methylation Methylation of DNA blocks transcription factors no transcription  genes turned off attachment of methyl groups (–CH3) to cytosine C = cytosine nearly permanent inactivation of genes ex. inactivated mammalian X chromosome = Barr body Methylation results in a human disease called fragile X syndrome; FMR-1 gene is silenced by methylation.

Histone acetylation Acetylation of histones unwinds DNA loosely wrapped around histones enables transcription genes turned on attachment of acetyl groups (–COCH3) to histones conformational change in histone proteins transcription factors have easier access to genes

Fig. 18.13 Methylation of H19 inactivates transcription (involved in expression of insulin like growth factor)

Fig. 18.10a Chromatin remodeling Acetylation of histones enhances access to promoter region and facilitates transcription.

Epigenetic Inheritance Although the chromatin modifications just discussed do not alter DNA sequence, they may be passed to future generations of cells The inheritance of traits transmitted by mechanisms not directly involving the nucleotide sequence is called epigenetic inheritance http://learn.genetics.utah.edu/content/epigenetics/intro/

2. Transcription initiation Control regions on DNA promoter nearby control sequence on DNA (TATA box ) binding of RNA polymerase & transcription factors “base” rate of transcription enhancer distant control sequences on DNA binding of activator proteins “enhanced” rate (high level) of transcription

Model for Enhancer action Enhancer DNA sequences distant control sequences Activator proteins bind to enhancer sequence & stimulates transcription Silencer proteins bind to enhancer sequence & block gene transcription Much of molecular biology research is trying to understand this: the regulation of transcription. Silencer proteins are, in essence, blocking the positive effect of activator proteins, preventing high level of transcription. Turning on Gene movie

Transcription complex Activator Proteins • regulatory proteins bind to DNA at distant enhancer sites • increase the rate of transcription Enhancer Sites regulatory sites on DNA distant from gene Enhancer Activator Activator Activator Coactivator B F E RNA polymerase II A TFIID H Coding region T A T A Core promoter and initiation complex Initiation Complex at Promoter Site binding site of RNA polymerase

Promoter Activators Gene DNA Enhancer Fig. 18-9-3 Promoter Activators Gene DNA Distal control element Enhancer TATA box General transcription factors DNA-bending protein Group of mediator proteins RNA polymerase II Figure 18.9 A model for the action of enhancers and transcription activators RNA polymerase II Transcription initiation complex RNA synthesis

3. Post-transcriptional control Alternative RNA splicing variable processing of exons creates a family of proteins

RNA processing is also an important part of gene regulation in eukaryotes. mRNA processing includes three major steps. Introns are removed and exons are spliced together. A cap is added. A tail is added.

Regulation by alternative splicing Calcitonin gene-related peptide

4. Regulation of mRNA degradation Life span of mRNA determines amount of protein synthesis mRNA can last from hours to weeks RNA processing movie

5. Control of translation Block initiation of translation stage regulatory proteins attach to 5' end of mRNA prevent attachment of ribosomal subunits & initiator tRNA block translation of mRNA to protein Control of translation movie

6-7. Protein processing & degradation folding, cleaving, adding sugar groups, targeting for transport Protein degradation ubiquitin tagging proteasome degradation The cell limits the lifetimes of normal proteins by selective degradation. Many proteins, such as the cyclins involved in regulating the cell cycle, must be relatively short-lived. Protein processing movie

Concept 18.3: Noncoding RNAs play multiple roles in controlling gene expression Only a small fraction of DNA codes for proteins, rRNA, and tRNA A significant amount of the genome may be transcribed into noncoding RNAs Noncoding RNAs regulate gene expression at two points: mRNA translation and chromatin configuration

RNA interference NEW! Small interfering RNAs (siRNA) short segments of RNA (21-28 bases) bind to mRNA create sections of double-stranded mRNA “death” tag for mRNA triggers degradation of mRNA cause gene “silencing” post-transcriptional control turns off gene = no protein produced siRNA

Action of siRNA siRNA mRNA degraded functionally turns gene off Hot…Hot new topic in biology Action of siRNA dicer enzyme mRNA for translation siRNA double-stranded miRNA + siRNA breakdown enzyme (RISC) mRNA degraded functionally turns gene off

Gene Regulation 7 6 protein processing & degradation 1 & 2. transcription - DNA packing - transcription factors 3 & 4. post-transcription - mRNA processing - splicing - 5’ cap & poly-A tail - breakdown by siRNA 5. translation - block start of translation 6 & 7. post-translation - protein processing - protein degradation 5 4 initiation of translation mRNA processing 2 1 initiation of transcription mRNA protection mRNA splicing 4 3

Cancers result from a series of genetic changes in a cell lineage The incidence of cancer increases with age because multiple somatic mutations are required to produce a cancerous cell As in many cancers, the development of colon cancer is gradual

Eukaryotic Gene Regulation Hox genes – controls differentiation of genes Responsible for general body pattern of most animals Order of genes is order of body parts

Review questions What does DNA polymerase do? What does Helicase do? What does ligase do? Match the bases below. 5’ – A T C G T A – 3’ List 3 differences between RNA/DNA. What are the 3 types of RNA? Where does RNA go after it is made? Transcribe the DNA below. A T C G T A

What does RNA attach to when it leaves the nucleus What does RNA attach to when it leaves the nucleus? Amino Acids are the building block of ________. What type of RNA brings an amino acid? When does translation stop? Where is the codon located? Anticodon? What is a codon? Translate the mRNA strand below. A G C G A G Translate the DNA strand below ATG CTA TGCA