CONTROL OF GENE EXPRESSION © 2012 Pearson Education, Inc.

 Gene regulation is the turning on and off of genes.  Gene expression is the overall process of information flow from genes to proteins. Proteins interacting with DNA turn prokaryotic genes on or off in response to environmental changes © 2012 Pearson Education, Inc.

Proteins interacting with DNA turn prokaryotic genes on or off in response to environmental changes  A cluster of genes with related functions, along with the control sequences, is called an operon.  Found mainly in prokaryotes. © 2012 Pearson Education, Inc.

Proteins interacting with DNA turn prokaryotic genes on or off in response to environmental changes  When an E. coli encounters lactose, all enzymes needed for its metabolism are made at once using the lactose operon.  The lactose (lac) operon includes 1.Three adjacent lactose-utilization genes 2.A promoter sequence where RNA polymerase binds and initiates transcription of all three lactose genes 3.An operator sequence where a repressor can bind and block RNA polymerase action. © 2012 Pearson Education, Inc.

Proteins interacting with DNA turn prokaryotic genes on or off in response to environmental changes  Regulation of the lac operon –A regulatory gene, located outside the operon, codes for a repressor protein, that is always being transcribed & translated. –In the absence of lactose, the repressor binds to the operator and prevents RNA polymerase action. –Lactose inactivates the repressor, so –The operator is unblocked –RNA polymerase can bind to the promoter, and –all three genes of the operon are transcribed. © 2012 Pearson Education, Inc.

Operon turned off (lactose is absent): OPERON Regulatory gene Promoter Operator Lactose-utilization genes RNA polymerase cannot attach to the promoter Active repressor Protein mRNA DNA Protein mRNA DNA Operon turned on (lactose inactivates the repressor): RNA polymerase is bound to the promoter Lactose Inactive repressor Translation Enzymes for lactose utilization

Multiple mechanisms regulate gene expression in eukaryotes  Multiple control points exist in Eukaryotic gene expression  Genes can be turned on or off, or sped up, or slowed down. © 2012 Pearson Education, Inc.

Chromosome DNA unpacking Other changes to the DNA Gene DNA Transcription Gene Exon Intron Tail Cap Addition of a cap and tail RNA transcript Splicing mRNA in nucleus NUCLEUS Flow through nuclear envelope CYTOPLASM

Figure 11.7_2 mRNA in cytoplasm CYTOPLASM Breakdown of mRNA Translation Polypeptide Broken- down mRNA Cleavage, modification, activation Active protein Amino acids Breakdown of protein

Multiple mechanisms regulate gene expression in eukaryotes  These control points include: 1.Chromosome changes and DNA unpacking 2.Control of transcription 3.Control of RNA processing including the –addition of a cap and tail and –splicing 4.Flow through the nuclear envelope 5.Breakdown of mRNA 6.Control of translation 7.Control after translation –cleavage/modification/activation of proteins –protein degradation © 2012 Pearson Education, Inc.

Chromosome structure and chemical modifications can affect gene expression  Eukaryotic chromosomes undergo multiple levels of folding and coiling, called DNA packing. –Nucleosomes are formed when DNA is wrapped around histone proteins. –Nucleosomes appear as “beads on a string”. –Each nucleosome bead includes DNA plus eight histones. –At the next level of packing, the beaded string is wrapped into a tight helical fiber (30nm). –This fiber coils further into a thick supercoil (300nm). –Looping and folding further compacts DNA into a metaphase chromosome © 2012 Pearson Education, Inc.

DNA double helix (2-nm diameter) “Beads on a string” Linker Histones Supercoil (300-nm diameter) Nucleosome (10-nm diameter) Tight helical fiber (30-nm diameter) Metaphase chromosome 700 nm

Chromosome structure and chemical modifications can affect gene expression  DNA packing can prevent gene expression by preventing RNA polymerase & other proteins from contacting the DNA.  Cells seem to use higher levels of packing for long-term inactivation of genes.  Highly compacted chromatin is generally not expressed © 2012 Pearson Education, Inc.

Chromosome structure and chemical modifications can affect gene expression  Epigenetic inheritance –Inheritance of traits transmitted by mechanisms that do not alter the sequence of nucleotides in DNA –Chemical modification of DNA bases or histone proteins can result in epigenetic inheritance –Ex. Enzymatic addition of a methyl group (CH 3 ) to DNA –Genes are not expressed when methylated –Removal of the extra methyl groups can turn on some of these genes © 2012 Pearson Education, Inc.

Chromosome structure and chemical modifications can affect gene expression  X-chromosome inactivation –In female mammals either the maternal or paternal chromosome is randomly inactivated. –occurs early in embryonic development; all cellular descendants have the same inactivated chromosome. –An inactivated X chromosome = Barr body. –Tortoiseshell fur coloration is due to inactivation of X chromosomes in heterozygous female cats. © 2012 Pearson Education, Inc.

Cell division and random X chromosome inactivation Early Embryo Adult X chromo- somes Allele for orange fur Allele for black fur Two cell populations Active X Inactive X Active X Inactive X Black fur Orange fur

Complex assemblies of proteins control eukaryotic transcription  Prokaryotes and eukaryotes employ regulatory proteins (activators and repressors) that –bind to specific segments of DNA –either promote or block the binding of RNA polymerase, turning the transcription of genes on and off. © 2012 Pearson Education, Inc.

Complex assemblies of proteins control eukaryotic transcription  Eukaryotic RNA polymerase requires the assistance of proteins = transcription factors.  Transcription Factors Include: –Activator proteins bind to DNA sequences called enhancers. –A DNA bending protein bends DNA, bringing bound activators closer to promoter. –Once bent activators interact with other proteins, allowing RNA pol to bind the promoter, leading to transcription. © 2012 Pearson Education, Inc.

Enhancers DNA Promoter Gene Transcription factors Activator proteins Other proteins RNA polymerase Bending of DNA Transcription

Small RNAs play multiple roles in controlling gene expression  A significant amount of the genome codes for microRNAs  microRNAs (miRNAs) can bind to complementary sequences on mRNA molecules either –degrading the target mRNA or –blocking its translation  RNA interference (RNAi) is the use of miRNA to artificially control gene expression by injecting miRNAs into a cell to turn off a specific gene sequence. © 2012 Pearson Education, Inc.

miRNA Target mRNA mRNA degraded or Translation blocked miRNA- protein complex Protein 3214

Folding of the polypeptide and the formation of S—S linkages Initial polypeptide (inactive) Folded polypeptide (inactive) Active form of insulin Cleavage S S S S S S S S S S S S SH Protein Activation: The role of polypeptide cleavage

Later Stages of Gene Expression are Subject to Regulation  Breakdown of mRNA –Enzymes in the cytoplasm destroy mRNA –mRNA’s of eukaryotes have lifetimes from hours to weeks  Protein Breakdown –Final control mechanism –Cells can adjust the kinds and amounts of its proteins in response to environmental changes –Damaged proteins are usually broken down right away

CLONING OF PLANTS AND ANIMALS © 2012 Pearson Education, Inc.

Plant cloning shows that differentiated cells may retain all of their genetic potential  Most differentiated cells retain a full set of genes, even though only a subset may be expressed. Evidence is available from –plant cloning, in which a root cell can divide to form an adult plant –salamander limb regeneration, in which the cells in the leg stump dedifferentiate, divide, and then redifferentiate, giving rise to a new leg. © 2012 Pearson Education, Inc.

Root of carrot plant Root cells cultured in growth medium Cell division in culture Single cell Plantlet Adult plant

Nuclear transplantation can be used to clone animals  Animal cloning can be achieved using nuclear transplantation: the nucleus of an egg cell or zygote is replaced with a nucleus from an adult somatic cell.  Using nuclear transplantation to produce new organisms is called reproductive cloning (first used in mammals in 1997 to produce Dolly) © 2012 Pearson Education, Inc.

Nuclear transplantation can be used to clone animals  Another way to clone uses embryonic stem (ES) cells harvested from a blastocyst. This procedure can be used to produce –cell cultures for research –stem cells for therapeutic treatments. © 2012 Pearson Education, Inc.

Figure 11.13_1 The nucleus is removed from an egg cell. Donor cell A somatic cell from an adult donor is added. Nucleus from the donor cell Blastocyst The cell grows in culture to produce an early embryo (blastocyst).

Figure 11.13_2 Reproductive cloning Blastocyst The blastocyst is implanted in a surrogate mother. A clone of the donor is born. Therapeutic cloning Embryonic stem cells are removed from the blastocyst and grown in culture. The stem cells are induced to form specialized cells.

Reproductive cloning has valuable applications, but human reproductive cloning raises ethical issues © 2012 Pearson Education, Inc.  Since Dolly’s landmark birth in 1997, researchers have cloned many other mammals, including mice, cats, horses, cows, mules, pigs, rabbits, ferrets, and dogs.  Cloned animals can show differences in anatomy and behavior due to –environmental influences and –random phenomena.

Reproductive cloning has valuable applications, but human reproductive cloning raises ethical issues  Reproductive cloning is used to produce animals with desirable traits to –produce better agricultural products –produce therapeutic agents –restock populations of endangered animals © 2012 Pearson Education, Inc.

Therapeutic cloning can produce stem cells with great medical potential  When grown in laboratory culture, stem cells can –divide indefinitely –give rise to many types of differentiated cells  Adult stem cells can give rise to many, but not all, types of cells © 2012 Pearson Education, Inc.

Therapeutic cloning can produce stem cells with great medical potential  Embryonic stem cells are considered more promising than adult stem cells for medical applications.  The ultimate aim of therapeutic cloning is to supply cells for the repair of damaged or diseased organs. © 2012 Pearson Education, Inc.

Figure Blood cells Nerve cells Heart muscle cells Different types of differentiated cells Different culture conditions Cultured embryonic stem cells Adult stem cells in bone marrow

THE GENETIC BASIS OF CANCER © 2012 Pearson Education, Inc.

Cancer results from mutations in genes that control cell division  Mutations in two types of genes can cause cancer. 1.Oncogenes –Proto-oncogenes are normal genes that promote cell division. –Mutations to proto-oncogenes create cancer-causing oncogenes that often stimulate cell division. 2.Tumor-suppressor genes –Tumor-suppressor genes normally inhibit cell division or function in the repair of DNA damage. –Mutations inactivate the genes and allow uncontrolled division to occur. © 2012 Pearson Education, Inc.

Oncogene Hyperactive growth- stimulating protein in a normal amount Normal growth- stimulating protein in excess New promoter The gene is moved to a new DNA locus, under new controls Multiple copies of the gene A mutation within the gene Proto-oncogene (for a protein that stimulates cell division) DNA

Tumor-suppressor gene Mutated tumor-suppressor gene Normal growth- inhibiting protein Cell division under control Defective, nonfunctioning protein Cell division not under control

Colon wall DNA changes: Cellular changes: Growth of a polyp An oncogene is activated Increased cell division A tumor-suppressor gene is inactivated A second tumor- suppressor gene is inactivated Growth of a malignant tumor 123

Figure 11.17B Chromosomes 1 mutation 2 mutations 3 mutations 4 mutations Normal cell Malignant cell

Lifestyle choices can reduce the risk of cancer  After heart disease, cancer is the second-leading cause of death in most industrialized nations.  Cancer can run in families if an individual inherits an oncogene or a mutant allele of a tumor-suppressor gene that makes cancer one step closer.  But most cancers cannot be associated with an inherited mutation. © 2012 Pearson Education, Inc.

Lifestyle choices can reduce the risk of cancer  Carcinogens are cancer-causing agents that alter DNA.  Most mutagens (substances that promote mutations) are carcinogens.  Two of the most potent carcinogens (mutagens) are –X-rays –ultraviolet radiation in sunlight. © 2012 Pearson Education, Inc.

Lifestyle choices can reduce the risk of cancer  The one substance known to cause more cases and types of cancer than any other single agent is tobacco smoke. © 2012 Pearson Education, Inc.