Control of Eukaryotic Genes (Ch. 19)

1. Control of Eukaryotic Genes (Ch. 19)

2. The BIG Questions… • How are genes turned on & off in eukaryotes? • How do cells with the same genes differentiate to perform completely different, specialized functions?

3. Evolution of gene regulation • Prokaryotes • single-celled • evolved to grow & divide rapidly • must respond quickly to changes in external environment • exploit transient resources • Gene regulation- Operons • turn genes on & off rapidly • flexibility & reversibility • adjust levels of enzymes for synthesis & digestion

4. Evolution of gene regulation • Eukaryotes • multicellular • evolved to maintain homeostasis • regulate body as a whole • specialization • turn on & off large number of genes • must coordinate the body as a whole rather than serve the needs of individual cells

5. Points of control • The control of gene expression can occur at any step in the pathway from gene to functional protein

6. from DNA double helix to condensed chromosome 1. DNA packing

7. Nucleosomes 8 histone molecules • “Beads on a string” • 1st level of DNA packing • histone proteins • 8 protein molecules • positively charged amino acids • bind tightly to negatively charged DNA

8. 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

9. DNA methylation • Methylation of DNA blocks transcription factors • attachment of methyl groups (–CH3) to cytosine • nearly permanent inactivation of genes • ex. inactivated mammalian X chromosome = Barr body

10. Histone acetylation Chromatin changes Transcription RNA processing Translation mRNA degradation Protein processing and degradation Histone tails DNA double helix Amino acids available for chemical modification (a) Histone tails protrude outward from a nucleosome Unacetylated histones Acetylated histones (b) Acetylation of histone tails promotes loose chromatin structure that permits transcription • Acetylation of histones unwinds DNA • Loose histone wrapping • enables transcription • genes turned on • attachment of acetyl groups (–COCH3) to histones • conformational change in histone proteins • transcription factors have easier access to genes

11. 2. Transcription initiation • Control regions on DNA • promoter • nearby control sequence on DNA • binding of RNA polymerase & transcription factors • enhancer • distant control sequences on DNA • binding of activator proteins

12. 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

13. 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 E F B RNA polymerase II Coding region TFIID H A T A T A Core promoter and initiation complex Initiation Complex at Promoter Site binding site of RNA polymerase

14. Many control elements for different genes are the same It is the combination of control elements that provides specificity

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

16. 4. Regulation of mRNA degradation • Life span of mRNA determines amount of protein synthesis • mRNA can last from hours to weeks

17. RNA interference NEW! The micro- RNA (miRNA) precursor folds back on itself, held together by hydrogen bonds. The bound miRNA can base-pair with any target mRNA that contains the complementary sequence. One strand of each short double- stranded RNA is degraded; the other strand (miRNA) then associates with a complex of proteins. 3 The miRNA-protein complex prevents gene expression either by degrading the target mRNA or by blocking its translation. 4 1 5 2 2 An enzyme called Dicer moves along the double- stranded RNA, cutting it into shorter segments. Chromatin changes Transcription RNA processing mRNA degradation Translation Protein complex Protein processing and degradation Dicer Degradation of mRNA OR miRNA Target mRNA Blockage of translation Hydrogen bond

18. A Brief, RNAi Cartoon

19. miRNA’s • Scientists discovery video • Animation of miRNA function

20. RNA interference 1990s | 2006 “for their discovery of RNA interference —gene silencing by double-stranded RNA” Andrew Fire Stanford Craig Mello U Mass

21. 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

22. 6-7. Protein processing & degradation • Protein processing • folding, cleaving, adding sugar groups, targeting for transport • Protein degradation • ubiquitin tagging • proteasome degradation

23. Ubiquitin 1980s | 2004 • “Death tag” • mark unwanted proteins with a label • 76 amino acid polypeptide, ubiquitin • labeled proteins are broken down rapidly in "waste disposers" • proteasomes Aaron Ciechanover Israel Avram Hershko Israel Irwin Rose UC Riverside

24. Proteasome • Protein-degrading “machine” • cell’s waste disposer • breaks down any proteins into 7-9 amino acid fragments • cellular recycling

25. 1 2 3 6 5 7 4 4 Gene Regulation 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 initiation of translation mRNAprocessing initiation of transcription mRNA protection mRNA splicing

26. Concept Check • Do miRNA’s increase or decrease gene expression? • Does histone de-acetylation increase or decrease gene expression?

27. Genome Sizes and Estimated Numbers of Genes*

28. Exons (regions of genes coding for protein, rRNA, tRNA) (1.5%) Types of DNA sequences in the human genome Repetitive DNA that includes transposable elements and related sequences (44%) Introns and regulatory sequences (24%) Unique noncoding DNA (15%) Repetitive DNA unrelated to transposable elements (about 15%) Alu elements (10%) Simple sequence DNA (3%) Large-segment duplications (5–6%)

29. Just how little of out Genome Codes for Genes?

30. Cancer results from genetic changes in cell cycle control

31. Proto-oncogene --> oncogene

33. Multistep Model of cancer

34. Genome Evolution How did our genome get so big?

37. The similarity in the amino acid sequences of the various globin proteins • Supports this model of gene duplication and mutation Table 19.1

38. Evolution of Genes with Novel Functions • The copies of some duplicated genes • Have diverged so much during evolutionary time that the functions of their encoded proteins are now substantially different

40. Concept check • How much of the human genome consists of exons? • How can exon shuffling lead to the evolution of a new gene

41. Turn yourQuestion Genes on!

42. A larger portion of the DNA in a eukaryotic cell is transcribed than would be predicted by the proteins made by the cell. What is being transcribed and what is its function? • Multiple enhancer regions are being transcribed to amplify the transcription of protein-coding genes. • These transcriptions are of non-coding “junk” DNA that is a remnant of mutated protein coding segments. The transcripts are degraded by nuclear enzymes. • The additional DNA that is transcribed is introns that are excised fro the primary transcript in the produciton of mRNA. • Many RNA coding genes are transcribed. Precursor RNAs fold into hairpin structures, which are cut and processed into miRNAs that regulate translation of mRNAs.

43. How is the coordinated transcription of genes involved in the same pathway regulated? • The genes are transcribed in one transcription unit, although each gene has its own promoter. • The genes are located in the same region of the chromosome, and enzymes de-acetylate the entire region so that transcription may begin. • A steroid hormone selectively binds to the promoters for all the genes. • The genes have the same combination of control elements in the enhancer that bind with the particular activators present in the cell.

44. Histones are • Small positively charged proteins that bind tightly to DNA • Small bodies in the nucleus involved in rRNA synthesis • Basic units of DNA packing consisting of DNA wound around a protein core. • DNA bending proteins that facilitate formation of transcription initiation complexes. • Proteins responsible for producing repeating sequences at telomeres.

45. Heterochromatin • Has a higher degree of packing than does euchromatin. • Is visible with a light microscope during interphase. • Is not actively involved in transcription • Makes up metaphase chromosomes • Is all of the above.

46. What is the main reason that prokaryotic genes average 1,000 nucleotride base pairs whereas human genes average about 27.000 base pairs? • Prokaryotes have smaller, bue many more individual genes. • Prokaryotes are more ancient organisms, longer genes arose later in evolution. • Prokaryotes are much simpler organism humans have many types of differentiated cells. • Prokaryotic genes do not have introns, human genes have many • Human proteins are much larger and more complex than prokaryotic proteins.

47. A eukaryotic gene typically has all of the following associated with it except • A promoter • An operator • Enhancers • Introns and exons • Control elements.