1. Chapter 15

2. I. Prokaryotic Gene Control  A. Conserves Energy and Resources by  1. only activating proteins when necessary  a. don’t make tryptophan if it can be  absorbed from environment  2. only producing proteins when needed  a. don’t need lactose digesting enzymes  if no lactose is present 

3.  B. control enzymes already in cell : post- translation control  1. allosteric enzymes  a. activated  b. inhibited  2. feed back inhibition  a. end product of  anabolic path is  the inhibitor  3. adjustment to short  term changes

4.  C. control production of enzymes: transcription control  1. control transcription of genes  a. repressors block promoters to stop  transcription  b. enhancers bind to promotor to speed  transcription  2. slower/longer lasting effects: more stable environment

5.  D. Negative control  1. negative slows or stops function  2. feed back inhibition  3. repressors blocking transcription  E. positive control  1. SPEEDS up production  2. just allowing production does not count!!  3. Enhancers bound to promoter  4. allosteric enhancers

6.  F. Operon model : clusters of functionally related genes controlled as a group (3 parts)  1. DNA code for the genes  2. promotor – stretch of DNA before genes  a. attracts RNA polymerase  b. needed to start transcription  3. operator – DNA sequence near promotor  a. binding site for repressor protein

7.  G. Regulatory Genes : make repressors  found up stream from operon they regulate

8.  H. trp Operon : trp = tryptophan amino acid  1. Repressible operon bcs it is normally active  2. genes make trp  3. low trp level in cell : operon active  a. repressor is inactive  b. promoter is open to RNA polymerase  c. genes to make trp are copied

9.  4. High trp level in cell : operon repressed  a. trp repressor is allosteric  1. binding to trp activates repressor  2. active repressor binds to operator  3. blocks RNA polymerase  4. genes not transcribed

10.  I. Lac operon : lactose (galactose + glucose)  1. Inducible operon : usually off  a. repressor is active unless lactose bound to it  2. genes make  a. β-galactosidase cleaves lactose in 1/2  b. permiase membrane transport protein  c. third gene  unknown

11.  3. presence of lactose:  a. lactose binds to allosteric repressor  i. inactivates repressor  ii. Repressor releases operator  iii. RNA polymerase copies all 3 genes  http://biology-animations.blogspot.com/2007/11/lac-operon-animation.html http://biology-animations.blogspot.com/2007/11/lac-operon-animation.html

12. J. Positive control of lac operon  1. lac operon also has positive control  a. inducible = can be activated  b. positive control = can be speeded up  2. If glucose is present E. coli prefer to use it  a. lack of glucose causes E. coli to speed up  use of lactose  b. lack of glucose causes build up of cAMP  (cyclic AMP) = signal molecule

13.  c. cAMP binds to regulatory protein CAP  i. catabolite activator protein becomes active  ii. CAP binds to start of promotor  iii. Makes promotor more attractive to RNA  polymerase  iv. speeds up translation  d. build up of glucose in cell causes lack of cAMP so CAP becomes inactivated

14.  https://smartsite.ucdavis.edu/access/content/user/00002950/bis10v/flashvideo/lac_positive.html https://smartsite.ucdavis.edu/access/content/user/00002950/bis10v/flashvideo/lac_positive.html

15.  Repressible operons  a) repressor inactive w/o allosteric binding  b) normally on  c) usually anabolic  Inducible operons  a) repressor active unless bound  b) normally off  c) usually catabolic

16. II. Eukaryotic Gene Control  A. Gene expression regulated at many stages  1. Transcription control  a. chromatin structure regulation  b. transcription initiation control  2. Post-transcriptional control  a. RNA processing  b. mRNA degradation  c. Translation initiation  3. Post-translational control a. allosteric P, b. P processing, c. P degradation

18. B. Chromatin structure control  1. Heterochromatin – Chromatin that remains tightly compacted even in interphase  a. genes not transcribed  2. acetylation –  a. acetyl group (-COCH 3 ) bonded to histone  b. loosens up chromatin winding  c. promotes transcription  3. DNA methylation –  a. –CH 3 bonds to DNA blocking transcription  b. methylated regions passed on to daughter  cells (x - chromosome inactivation)

20. C. Initiation control  1. control elements : non-coding DNA up- stream from promotor that bind transcription factor proteins  a. distal control elements are far up-stream  i. often act as enhancers  b. proximal control elements : near promotor

21.  2. transcription factors: proteins  a. needed for transcription initiation  b. general transcription factors (GTF) needed for all transcription of genes  i. GTFs bind each other & RNA Polym. II to form initiation complex  ii. Initiation complex binds to control elements near promotor: start transcription

22.  iii. One protein of the GTF will bind to a section of promotor called the TATA box. (fig 14.9)  iv. General Transcription Factor complexes allow slow transcription of gene  f. Specific Transcription Factors needed for rapid transcription of gene 

23.  3. Vocabulary in order to have a clue on 15.2  a. Things that are part of the DNA  1. control elements : binding site for transcription factors  2. Enhancers : distal (far) control elements, can be activated or repressed by transcription factor proteins  3. TATA box : section of the promoter’s code  4. promoter : just upstream from start of gene, where RNA polymerase binds to start transcription

24.  b. Things that are proteins  1. Transcription factor : regulatory protein binds control elements on  a. general transcription factors allow transcription  b. activators speed transcription  c. repressors slow transcription  2. Mediator proteins : form link between regulatory proteins and DNA

25.  4. Distal control elements = enhancers  a. may be up or down stream  b. each gene can have many enhancers  i. each active under different conditions  ii. Or active in different cell types  iii. Each enhancer works with only one gene  c. transcription factors called activator proteins bind to enhancer control elements  i. fold DNA so that the activator protein/enhancer complex binds to initiation complex to speed up transcription

27.  d. repressor transcription factors interfere with the activator transcription factors to slow transcription  i. by binding to distal control elements and keeping activators out  ii. By binding to activator proteins

28.  5.coordination of functionally related genes  a. related genes have same control element sequences  b. bind same transcription factors  c. environmental signal triggers TF and the bind to all the matching control elements in the genome  d. activate all the related genes

29. D. Post-transcription Control  1. RNA processing  a. alternative splicing  b. poly A tail length  c. cap designation  2. mRNA degradation  a. specialized RNAs can degrade mRNA  3. Translation initiation control  a. proteins bond to mRNA prevent initiation  b. egg mRNA lack poly-A tail so no initiation  c. global control : lack of initiation factor (egg)

30. E. Protein Processing/degradation  1. Allosteric control or activation by phosphate  2. protein processing  a. inactive form cut to activate (pro-insulin)  b. glycoproteins, lipoproteins  3. selective degradation  a. ubiquitin = protein attached to proteins tags them for destruction

31. F. Non-coding RNA (ncRNA) 1)Don’t code for proteins/affect gene expression  a) microRNAs (miRNAs)  i. complexes w/ proteins  ii. binds to complementary mRNA  iii stops translation or trigger degradation  b) small interfering RNA (siRNA)  i. turn off gene expression  ii. Used in knock-out experiments  c) ncRNA affect heterochromatin formation

32. G. Monitoring gene expression  1) in situ hybridization (see if gene is transcribed)  a. fluorescent DNA probe added to solution around embryo  b. probe hybridizes & concentrates in cells that have complementary mRNA  2) reverse transcriptase – PCR (RT-PCR)  a. used to see how much mRNA is present  b. make cDNA  c. do PCR for genes of interest  d. run electrophoresis to see what cells have it  3) RNA sequencing : sequence cDNA

33. In situ hybridization fruit fly embryo