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MCB 3020, Spring 2005 Chapter 7: Molecular Genetics. Molecular Genetics I: Replication I. Heredity and Genetics II. Genomes III. DNA structure IV. Bacterial DNA replication V. Replication at the ends of linear DNA. DNA carries the information. - PowerPoint PPT Presentation
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1
MCB 3020, Spring 2005
Chapter 7:Molecular Genetics
2Molecular Genetics I: ReplicationI. Heredity and GeneticsII. GenomesIII. DNA structureIV. Bacterial DNA replicationV. Replication at the ends of linear DNA
3I. HeredityThe transmission of characters to progeny.DNA carries the information necessary for the transmission of characters.The biological information is encoded in the sequence of bases.
TB
4
the study of the mechanisms of heredity and variation in organisms
Genetics:
TB
5
Flow of information replication DNA DNAtranscription RNAtranslation protein
6
Genome = all the DNA of a cell (or all the genetic
material of a virus)
II. Genome
7
one circular double-stranded DNA chromosome
oftenplasmid(s)
Typical bacterial genome
500-12,000 genes TB
8typical viral genome
DNAor
RNA
4-200 genes TB
9Typical eukaryotic genome
4-224, linear chromosomes
5,000 - 125,000 genes TB
10III. DNA structuredeoxyneucleotidesphosphodiester bonds5' and 3' endsantiparallelcomplementarydouble helix
TB
11
N
N N
N
NH2
Deoxyadenosine (purine)
HOCH2
TB
HO H
12
HO
NH
N
deoxythymidine (pyrimidine)
O
OHOCH2
TB
H
H3C
13
O
-P-O-C
OP
OC
-O
O-
phosphodiester bond
ssDNATB
14
O
-P-O-C
OP
OC
HO
O-
3’ end ssDNA
5' end
5’
2’
1’
3’
4’
ring numbering system for
deoxyribose
-C
TB
15dsDNA
5’5’3’3’
antiparallel
dsDNA is always antiparallel
TB
16complementary
GGATGCGT
3’-CCTACGCA-5’
Two ssDNA molecules joined bystandard base-pairing rulesIn dsDNA, the strands are alwayscomplementary.
TB
5’- -3’
17
double helix
right handed
TB
18Supercoiling
relaxed DNA supercoiled DNA
TB
Within cells DNA is supercoiled
19IV. Bacterial DNA replication
DNA synthesis using a DNA template
Complementary base pairing (A=T, GC) determines the sequence of the newly synthesized strand.
DNA replication always proceeds from 5’ to 3’ end.
TB
20
Flow of information replication DNA DNAtranscription RNAtranslation protein
21Overview of bacterial DNA replication
single origin (in bacteria)bidirectionaltheta structuresreplication forksemi-conservative
TB
22bacterial DNA replication
bacterialchromosome
origin (start point) bidirectional
TB
23
two replication forks
thetastructure
TB
24semi-conservative
+
**
*
* TB
25
Key EnzymeshelicasessDNA binding proteinprimaseDNA polymerase IIIDNA polymerase IDNA ligase
TB
IV. Bacterial DNA replication
26
All DNA polymerases require a primer
DNA is synthesized 5' to 3'
Important facts
TB
27helicase
Unwinds duplex DNA
TB
28ssDNA binding protein
binds to and stabilizes ssDNA
prevents base pairing
ssDNA binding proteinTB
29primase
synthesizes a short RNA primerusing a DNA template
RNA primer(a short starting sequence made of RNA)
primase
TB
30DNA polymerase III
Synthesizes DNA from a DNAtemplate and proofreads
TB
31DNA polymerase I
Synthesizes DNA from a DNAtemplate and removes RNA primers.
TB
32DNA ligaseJoins DNA strands together by forming phosphodiester bonds
DNA ligase
TB
33replication fork
5'
5'
3'
3'
template strands
lagging strand
leading strand
TB
34
helicasessDNA binding proteins
RNA primer
3'
5'Leading strand synthesis
TB
35
helicase
ssDNA binding proteins
DNA polymerase
3'
5'
TB
36
helicase
ssDNA binding proteins
DNA pol III
3'
5'Leading strand synthesis
TBDNA
37
helicasessDNA binding proteins
(primase)pol III
3'
Lagging strand synthesis (discontinuous) Okazaki
fragment(~1000 bases)
TB
3'
5'
38Primer removalpol III
pol I
pol I
3'
5'
5’ to 3’exonucleaseactivity
TB
39
DNA ligase
Ligation
TB
40Proofreading
Pol III removes misincorporated basesusing 3' to 5' exonuclease activity
This decreases the error rate to about10-10 per base pair inserted
TB
41
Since all known DNA polymerasesneed a primer, how are the ends oflinear DNA replicated in eukaryotes?
5' 3'
RNA primer
template
newly synthesized DNA
TB
V. Replication of the ends of linear DNA
42
repetitive DNA at the end of lineareukaryotic chromosomes
Telomeres
(GGGGTT)n
Example
n = 20 - 200
GGGGTT GGGGTT GGGGTT
5' TB
43Telomerases are enzymes that add DNA repeats to the 3' end of DNA.
Telomerases are composed of protein and an RNA molecule that functions as the template for telomere synthesis.
AACCCCAAC
telomerase
44
AACCCCAAC
5'GGGGTTGGGGTT
5'
telomerase
45
AACCCCAAC
5'
5'GGGGTTGGGGTT GGGGTT
primase
GGGGTT GGGGTT GGGGTT
46pol III
pol I5'
ligase
telomeric repeats
47For most cells, telomeres are added during development. Later telomerase becomes inactive.
Hence, as cells divide the DNA becomes shorter.
TB
Note that telomerase is reactivated in many types of cancer cells.
48Study objectives1. Compare and contrast bacterial, viral and eukaryotic genomes.2. What are the 4 bases in DNA? Which are purines? Which are pyrimidines? What is the sugar? I will not ask you to recognize the structures of individual bases, but note that deoxythymidine has a methyl group in the pyrimidine ring.3. Understand how the following terms apply to DNA structure: phosphodiester bonds, 5' and 3' ends, antiparallel, complementary, double helix. What parts of the nucleotides are joined in the phosphodiester bond? 4. Understand how the following terms apply to DNA replication: template, complementary base pairing, origin, bi-directional, theta structures, replication fork, semi-conservative.5. Know how the following enzymes function in leading and lagging strand replication: helicase, ssDNA binding protein, primase, DNA polymerase III, DNA polymerase I. What is an Okazaki fragment?6. What is proofreading?7. Understand the problem of replicating the ends of linear DNA. Understand how telomerase solves that problem for eukaryotic chromosomes.
49Molecular Genetics II: TranscriptionI. RNA II. Gene expressionIII. Prokaryotic Transcription
50
Flow of information replication DNA DNAtranscription RNAtranslation protein
51I. RNA (ribonucleic acid)
A polymer of nucleosides held together by phosphodiester bonds.
RNA plays a key role in decodingthe information in DNA.
RNA is usually single-stranded.
TB
52A. Functions of the major RNAs
1. messenger RNAs (mRNA) contain genetic information to encode a protein
3. ribosomal RNAs (rRNA) are structural and catalytic component of ribosomes
2. transfer RNAs (tRNA) act as adapters between the mRNA nucleotide code and amino acids during protein synthesis
phe
53B. RNA structure
1. RNA nucleosides2. phosphodiester bonds3. 5' and 3' ends4. complementary base pairing5. stem-loops
TB
54
The RNA nucleosides have 2'-hydroxyl groups which arenot found in DNA.
1. The RNA nucleosides
"U" is found in RNA (in place of "T")
Guanosine (G)Adenosine (A)
Cytidine (C)Uridine (U) TB
55
2. The phosphodiester bonds of RNA are analogous to those of DNA.
3. The 5' and 3' ends of RNA are analogous to those of DNA.
TB
56
O
P-O-C5' end
ring numbering system for ribose
O
5'
2’
1’
3’
4’-C O
OP
OC
HO
O-
3’ end RNA
O
OH
OHphosphodiester
bond
TB
574. Complementary base pairing
CCCUUUGGGAAA
GGGAAACCCUUU RNA
RNA
GGGAAACCCUUU RNA
CCCTTTGGGAAA DNA
TB
hydrogenbonding
585. RNA stem loops
complementarybase pairing(helical)ssRNA
A common RNA secondary structure
TB
59II. Gene expressionDifferent scientists define the termgene expression differently. Most commonly, gene expression refers tothe decoding of genes into proteinsor RNAs.
1 gene encodes 1 polypeptide, 1 tRNA, 1 rRNA, or 1other RNA TB
60A. Gene numbers
virusesprokaryotes eukaryotes
groupapproximategene number
4-200500-12,000 5,000-125,000
TB
61Any given species has a unique setof genes that confers a unique set of properties.
Proteins and RNAs determine all of thecharacteristics of organisms and cells.
Example: Escherichia coli has 4405 genes
~117 encode RNAs (tRNA, rRNA) ~4288 encode proteins TB
62
1 gene
1 mRNA
transcription
1 polypeptide
translation
1. Expression of single genesEx.1: a single gene that encodes a protein
C. Gene expression in prokaryotes
TB
63
1 genetranscription
1 RNA
degraded 1 tRNA etc.
RNA processing
Ex. 2: a single gene that encodes one rRNA or tRNA
TB
64
operontwo or more genes transcribed together
a single RNA molecule that represents more than one gene
polycistronic message
2. Expression of operons
TB
A B CDNA
transcription
polycistronicmRNA
65a. Operons can encode several polypeptides or proteins.
TB
1 operonA B C
2 or more polypeptides
translation
AB
C
1 polycistronic mRNA
transcription
66
1 operon
processing
rRNArRNA
degraded2 or more rRNAs
b. Operons can encode several rRNA molecules.
1 polycistronic RNA
TB
673. Important points
Most prokaryotes use operons.Operons are used to coordinategene expression and often containgenes of related function.
The details of organization, processing and degradation are different for different RNAs.
TB
68
The expression of rRNA and tRNA is similar in eukaryotes and prokaryotes.
1. Expression of eukaryotic rRNA and tRNA genes
D. Eukaryotic gene expression
TB
69
geneE I I IE E E
E = exon = coding sequences I = intron = intervening, noncoding sequences
2. Eukaryotic protein expressiona. Typical eukaryotic genes have exons and introns.
Eukaryotes do NOT have operons TB
701 gene with exons and intronsE I I IE E E
transcription
1 RNA representing exons and introns(primary transcript)
TB
71
primary transcript
1 polypeptide
1 mRNA
processing
TB
b. Primary transcripts
72c. Processing of primary transcripts
i. cappingii. splicingiii. tailing
TB
73i. CappingAddition of a 5' cap
CAP
Capping usually occurs beforetranscription is finished.
TB
74
OCH2
HO
N
N N
N NH2
O
OH
CH3Typical 5' CAP
7-methylguanosine
PP
P
5' carbonof RNA chain
5' to 5' linkage
O
TBKnow the name (methylguanosine cap, 5' cap), but don't memorize structure.
75ii. SplicingThe removal of introns.
primary transcriptsplicing
RNA without intronsTB
76
Addition of a poly-A tail
iii. Tailing
A1A2...A~200
TB
773. Notes on eukaryotic RNA processing
Processing occurs in the nucleus
The exact order of capping, tailing and splicing varies for different genes.
Poly-A tails are added by poly-Apolymerase, NOT during transcription.
TB
78E. Comparison of eukaryotic and prokaryotic gene expression.
Eukaryotic mRNAs are usuallyspliced,capped and tailed.
Eukaryotes do NOT have operons.
tRNA and rRNA expression are generally similar
Prokaryotic genes very very rarely have introns TB
79III. Prokaryotic transcription
A. overviewB. transcribed regionsC. RNA polymeraseD. promotersE. terminatorsF. sigma factor
TB
80
Flow of information replication DNA DNAtranscription RNAtranslation protein
81A. Overview of prokaryotic transcription
RNA polymerase
primary transcript complementary to one strand of the coding region
RNA synthesis from a DNA template
typicalgene dsDNA
TB
82B. Defined regions are transcribed
upstream region
transcribedregion
downstream region
promoter(RNA polymerase
binding site)
transcriptionstart site
terminationsite
gene dsDNA
TB
83
gene,or operon
RNA polymerase
C. RNA polymerase is the enzyme that synthesizes RNA from a DNA template.
complementary RNA
DNA template
TB
84
++ completed
transcriptTB
85D. PromotersSites on DNA where RNA polymerase binds to start transcription
promoter
upstream region
transcribedregion
downstream region
transcriptionstart site
terminationsite
gene dsDNA
TB
861. Typical bacterial 70 promoter
TTGACA TATAATAACTGT ATATTA
TATAAT = -10 consensus
sequence
TTGACA = -35 consensus
sequence
TB
*also called Pribnow box; ~ 10 bases before start
site of transcription
87A more common way to draw a promoter
TTGACA TTAACT
-10-35
Note:The - 10 and -35 sequences can vary somewhat.
5' 3'
TB
88E. Transcriptional terminatorsDNA region that mediates the termination of transcription.
gene dsDNA
region whereterminators areusually found
terminationsite
TB
891. Intrinsic terminator
DNA encoding an RNA that formsa stem loop followed by a run of "U"sthat is used for transcriptional termination.
UUUURNA
3' end of RNATB
90
The RNA stem loop binds to RNA pol and causes termination
Intrinsic terminator function
Important fact: Intrinsic terminators must betranscribed in order to function. TB
912. Rho-dependent terminatorA DNA site where RNA polymerasepauses and transcription is terminated by Rho protein
TB
92
Rho protein
Rho protein binds RNA then moves along RNA until it contacts RNA pol and terminates transcription
RNA polpauses at
Rho termination siteTB
93F. The sigma factor cycle
Sigma factors are needed for promoter binding, but after transcription starts they dissociate.
Sigma factors ( ) are a subunit of RNA polymerase.
TB
941. Subunit structure of bacterial RNA polymerase
'
core enzyme
The holoenzyme includes one of several sigma factors. TB
'
holoenzyme
95RNA pol holoenzyme (core + sigma)
sigmafactor
RNA (~10 nucleotides)
sigma factor
core enzyme
TB
96
+core enzyme
termination
RNA
sigma
holoenzyme TB
97
TAATGTGAGTTAGCTCACTCATTA
GGCACCCCAGGCTTGACATTTATG
CTTCCGGCTCGTATGTTGTGTGGA
AATTGTGAGCGGATAACAATTTCA
CACAGGAAAGAGCTATGACC...
Upstream region of the lactose operon
-35 region
-10 region (Pribnow)
Shine-dalgarno (RBS)
Translation start site
Transcription start site
98Study objectivesYou will need to know ALL the concepts and details in this lecture.1. What are the three main types of RNA and what are their functions?2. Understand how the following terms apply to RNA structure: phosphodiester bonds, 5' and 3 ends, nucleosides, complementary base pairing, stem loops.3. Compare and contrast DNA and RNA structure.4. What is a gene? What is gene expression? *Understand transcription, translation, and RNA processing in both prokaryotes and eukaryotes. 5. Define operons and polycistronic messages. How do they function in prokaryotic gene expression? 6. *Compare and contrast the features of prokaryotic and eukaryotic gene expression. Do eukaryotes have operons? What are exons, introns, primary transcripts, capping, tailing, and splicing. What is the 5' cap (methylguanosine cap)? How and when is the poly-A tail added to the transcript? Where does eukaryotic RNA processing occur? 7. Understand the structure and function of promoters and terminators in transcription. Contrast intrinsic terminators and rho-dependent terminators.8. Know the subunit structure of bacterial RNA polymerase and the sigma cycle.
99Molecular Genetics III: Prokaryotic translation
I. Key components of translation II. Steps in translationIII. The genetic code
100Overview of prokaryotic translationProtein synthesis from an mRNA template.
mRNA
translated region
translation
protein of specific amino acid sequenceTB
phe
101I. Key components of translation
A. mRNAB. tRNAC. ribosomes and rRNA
102
translated regionseries of codons
(usually ~300 codons)
mRNA
start codon
A. mRNA
stop codon
Shine-Dalgarno sequence
TB
RNA template for protein synthesis
1031. Shine-Dalgarno sequence~AGGAGG, ribosome binding sequence, critical for ribosome binding
2. start codonsAUG, GUG, or UUG
TB
3. stop codons (nonsense codons)
UAA, UGA, or UAG
1044. Translated region (coding sequence)• Series of codons that determines the amino acid sequence of the encoded protein.
• Coding sequences have an average of about 300 codons.
• Except for the stop codon, each codon specifies a particular amino acid. TB
105
AUGCAUUGUUCU...codons
protein fMet1 2
- His3
- Cys4
- Ser ...
5. Codons consist of 3 bases
TB
1 2 3 4
startcodon
106B. tRNAThe adapter molecule for translation
1. Particular tRNAs carry particular amino acids.
TBtRNA-f-Met
f-Met
tRNA-His
His
His
107
AUGCAUUGUUCU...
codons
AA1 AA2
tRNAs
2. Particular tRNAs recognize particular codons.
amino acid (AA)
This allows amino acids to be brought together in a particular order. TB
1083. tRNA structureAll tRNAs are generally similar in structure.
TB
a. 1o structure
ssRNA 73-93 nucleotides long
5' 3'UAC
109b. 2o structureclover leaf
anticodon loop
TC armD-arm
acceptor arm
extra arm
TB
110c. 3o structure inverted "L"
TB
111d. AnticodonA 3 base sequence in tRNA complementary to a specific codon.
anticodonBase pairing between an anticodon and a codon allows a tRNA to recognize a specific codon. TB
112e. codon-anticodon interactions
AAU5' 3' mRNA
codon1 2 3
UUA
anticodon123
5'3'
tRNA
TB
1134. tRNA charging (adding amino acid)
3'
tRNA(uncharged)
3'H2N-CH-C-OR
O
aminoacyl-tRNA(charged)
tRNA charging uses the energy of ATP TB
114Aminoacyl-tRNA synthetases
amino acidATP
tRNAaminoacyl-AMP
AMP PPiaminoacyl-tRNA
AMP = adenosine monophosphate PPi = inorganic pyrophosphate
enzymes that attach amino acids to tRNA
TB
enzyme
1155. tRNA facts
tRNAs contain many modified bases.
Prokaryotes have about 60 differenttRNAs.
TB
116C. Ribosomes and rRNA
Ribosomesribonucleoprotein complexes that
catalyze protein synthesis.
rRNAs have structural and catalytic roles
TB
1171. Prokaryotic 70s ribosome
23s rRNA5s rRNA34 proteins
16s RNA21 proteins
50ssubunit
30ssubunit
TB
118
A
2. Ribosomal sites where tRNAs bind
E = exit
PP = peptidyl
A = aminoacyl
E
TB
1193. 16S rRNA
The 3' end of the 16s rRNA is complementary to the Shine-Dalgarno sequence (ribosome binding sequence of mRNAs)
120
AUG
P-site f-met
Shine-Dalgarno(AGGAGG on mRNA)
II. Steps in translation
mRNA
A. initiation 30s subunitof ribosome
50s subunit
GTP hydrolysis
f-met
30s subunit TB
AGGAGG-----AUG
1211. f-met tRNA (formyl-methionine tRNA)
In Bacteria, different met-tRNAs are used forelongation and initiation.
initiation, formyl-methioninetRNAmetf
elongation, methioninetRNAmetm
TB
122
In Eukarya, the ribosome recognizes the 7-methylguanosine cap at the 5’ end ofmRNA and initiates at the first AUG.
In Eukarya and Archaea, initiation begins with methionine rather than f-met.
In Bacteria, the formyl group of the initiator formylmethionine (f-met) is later removed.
TB
2. Initiation in different domains
123
AA
P-site A-site
AA1. AA-tRNA binding
AA AA
mRNA
B. Elongation
TB
124
AA AA
AAAA
(peptidyl transferase)
2. peptide bond synthesis
TB
1253. translocation
AAAA
GTPhydrolysis
TB
AAAA
126
AAAA
C. Termination
AAAAAA
UAA
AAAA
AAAAAA
termination
stop codon
TB
127
"Polysomes" are mRNAs with several ribosomes attached.
mRNA
mRNAs can be translated by 5-10 ribosomes simultaneously.
1. Ribosomes move along the mRNA.
D. Additional notes on translation
TB
1282. In prokaryotes only, transcription and translation are coupled.
Translation begins before transcription ends.
DNA
mRNA TB
1293. Protein folding into the active form can occur spontaneously or with the help of a large protein complex called a molecular chaperone.
ATP
ADP
properly folded protein
improperly folded protein molecular
chaperone
130III. The genetic code
A. universal codeB. degenerate code
1. synonyms2. codon families3. codon pairs
C. wobble base pairing
131III. The genetic code 8 codon families, 14 codon pairs, 3 stop codons
(Do not memorize)
132A. The genetic code is almost universal.
Most organisms use the same genetic code.
TB
133B. The genetic code is degenerate.
more than one codon can code for the same amino acid
TB
UUU phenylalanineUUC phenylalanine
1341. synonymscodons that code for the sameamino acid
Not all synonyms are used with equal frequency. This is called "codon usage bias".
UUU phenylalanineUUC phenylalanine
1352. codon families
CUUCUCCUACUG
leucine
any nucleotide in the 3rd positions
TB
1363. codon pairs
UUUUUC phenylalanine
any pyrimidine in the 3rd position
CAACAG glutamine
any purine inthe 3rd position
TB
137C. Wobble base pairing
UUUAAG
codon (mRNA)
anticodon (tRNA)
5'3'
3'5'
U-G and G-U base pairs are allowed inthe 3rd position of the codon.
TB
138
Flow of information replication DNA DNAtranscription RNAtranslation protein
139Study objectives1. Know the DETAILS of the structure and function of mRNA, tRNA, rRNA, and ribosomes in translation. Memorize the start and stop codons. You do NOT need to memorize codons other than the start and stop codons. 2. What reaction is catalyzed by aminoacyl-tRNA synthetases? 3. For the process of translation, know the details of initiation, elongation peptide bond formation, translocation and termination. 4. Compare and contrast Bacterial, Archaeal and Eukaryal initiation.5. What are polysomes?6. What is meant when it is said that transcription and translation are coupled in prokaryotes? 7. Some proteins fold spontaneously while others require assistance. What are molecular chaperones?8. How do the following terms apply to the genetic code: synonyms, codon pairs, codon families, wobble, codon usage bias.
140
MCB 3020, Spring 2004
Chapter 7:Regulation of
Gene Expression
141
Note: put allosteric regulation in protein / enzyme section
Regulation of Gene Expression I:I. Regulation of gene expressionII. Transcriptional regulationIII. Examples of gene repressionIV. Example of gene induction
142
Not all genes are turned on (expressed) all the time
In general, they are turned ononly when needed.
I. Regulation of gene expression
TB
143Cells can respond to environmental changes by regulating gene expression.
glucose
maltose
lactose
arginine
tryptophan
144Different genes are expressed when cells grow on different compounds.
maltoseglucose
TCA
lactose
P O lacZ lacY lacA
lac permease (transport protein)-galactosidase
e.g. Growth on lactose requires expression of at least three additional genes.
(galactose--1,4-glucose)
145A. Why regulate gene expression?
Regulation allows cells to respond to environmental conditions by synthesizing selected gene products only when they are needed.
146B. Gene expression synthesis of a gene product
1. constitutive2. regulated
1471. Constitutive gene expression
e.g. "housekeeping genes" like primase ssDNA binding proteins
expression of genes at about the same level under all environmental conditions
TB
1482. Regulated gene expressionControl of the rate of protein or RNA synthesis as an adaptive response to stimuli.
induction: increase in gene expression
repression: decrease in gene expression
149a. gene induction increase in gene expression
amount of gene product
time
inducer
TB
150
maltosecatabolicenzymes(molecules/cell)
time
maltose absent
e.g. genes that encode maltose-utilizing enzymes are induced by maltose.
maltose added
lag phase
151b. gene repression decrease in gene expression
amount of gene product
timeTB
152
enzymes for tryptophanbiosynthesis(molecules/cell)
time
tryptophan absent tryptophan present
e.g. genes that encode enzymes for tryptophan biosynthesis are repressed by tryptophan.
153Important general principle
• catabolic substrates (e.g. maltose and lactose) induce the genes required for their catabolism
• biosynthetic molecules (e.g. amino acids and purines) repress the genes required for the biosynthesis
154II. Transcriptional regulation
• regulation of RNA synthesis• the most common method of gene regulation in all cells
A. Regulatory proteinsB. Regulatory protein binding sitesC. Effector molecules
TB
155A. Regulatory proteins
• Cells have many different regulatory proteins.• Specific regulatory proteins control the transcription of specific groups of genes.
• Transcriptional regulation is mediated by regulatory proteins.
TB
• Examples of regulatory proteins are "repressor proteins" and "activator proteins."
156
DNA
RNA polymeraseP
Promoter
Repressor protein (dimer)
Repressor proteins decrease transcription when bound to DNA by interfering with the activity or binding of RNA polymerase.
1. Repressor proteins
TB
157
RNA polymerase
2. Activator proteins
DNA
P
"weak" promoter
Activator protein
Activator proteins increase transcription when bound to DNA by helping RNA polymerase bind to weak promoters. TB
158B. Regulatory protein binding sites
Regulatory proteins bind to specific DNA sequences.
A particular regulatory protein will only control the expression of genes having appropriate binding sites.
TB
1591. Operator sites
Imperfect palindrome
GTGTAAACGATTCCAC
CACATTTGCTAAGGTG
binding sites for repressor proteins
Usually found near promoters.
lac repressor binding site
TB
1602. Activator binding sites
GTGAGTTAGCTCAC
CACTCAATCGAGTG
Imperfect palindrome
Binding sites for activator proteins
Usually found near promoters.
crp binding
site
TB
161C. Effector molecules
Small molecules from the environment (or made inside cells) that signal specific changes in gene expression.
TB
162
e.g. catabolic substrates: sugars, amino acids, fatty acids
a. inducerssmall molecules that mediate gene induction
1. Classes of effectors
lactose
maltose
TB
163
e.g. biosynthetic products:amino acids, purines, pyrimidines, fatty acids etc.
b. corepressorssmall molecules that mediate gene repression
tryptophanarginineTB
1642. How effectors work
conformational change (change in 3-D structure)
regulatory protein effector
Effectors change the DNA binding affinityof regulatory proteins for their binding sites.
TB
165
DNA
conformational change(change in 3-D structure)
regulatory protein
effector
A. Some effectors increase DNA binding affinity
TB
166
DNA
regulatory protein
conformational change(change in 3-D structure)
B. Some effectors decrease DNA binding affinity
effector TB
167Since most regulatory proteins influence transcription when bound to DNA, the binding of effectors to regulatory proteins changes gene expression.
TB
effector
regulatory protein
168
TB
III. Examples of gene repression
A. Regulation of the trp operonB. Regulation of the arg operon
169
The trp operon
polycistronic mRNA
E D C B A
Five enzymes for tryptophan biosynthesis
trp genespromoter
TB
A. The trp operon is a group of genes used for biosynthesis of the amino acid tryptophan (Trp).
170
2. When Trp is available, E. coli takes up Trp from the environment and represses the trp operon.
1. When Trp is NOT available in the environment, expression of the trp operon allows Escherchia coli to make Trp needed for protein synthesis.
TB
171trp promoter
operatorinactiverepressor
genes on TB
RNA polymerase
tryptophan
activerepressor
genes off
172Note: Repression of the trp operon by tryptophan involves a repressor protein.
• When tryptophan binds to the repressor protein, the repressor protein binds to DNA. • Transcription is blocked.
Result: VERY low amounts of tryptophan are synthesized when the cell can get tryptophan from the environment .
173
P
B. Regulation of the arg operon for arginine biosynthesis
operator arg biosynthetic genesargC argB argH
If arginine is present in large amounts • arg biosynthetic enzymes NOT needed
• RNA polymerase can't bind to promoter
• arg binds repressor • arg-repressor binds DNA
transcription rate decreases
174
Poperator arg biosynthetic genes
argC argB argH
If arg is absent, the cell needs to make arg • repressor doesn't bind DNA
• RNA polymerase can bind • transcription of arg genes occurs
175IV. Example of gene induction: Regulation of the lac operon
A. The lac operon is a group of genes used for catabolism of the sugar lactose.
Z Y A
lac genespromoter
operatorTB
176
• When lactose is available, E. coli induces expression of lac operon.
• When lactose is unavailable, the catabolic enzymes are NOT needed.
The lac operon isexpressed at only very low levels.
TB
177B. Lactose unavailable
In the ABSENCE of lactose, the lac repressor protein binds DNA.
Z Y A
lac promoter
genesoff
Note: the role of crp/cAMP in control of thelac operon is not considered here. TB
178
Z Y A
C. Lactose available lac promoter
geneson
RNA polymerase
lactose allolactose
repressor does not bind DNA TB
179Important points
Repressor proteins can mediate gene repression (e.g. trp operon) or gene induction (lac operon).
Activator proteins can mediate both gene induction and gene repression.
TB
180
Lactose (a sugar) can be an energy source.If lactose is absent, • enzymes for using lactose are not needed • lac repressor binds to the lac operator • the lac genes are not expressed
P O lacZ lacY lacACAPsite
Some repressor proteins mediate gene induction.Example: the lac repressor
181
P O lacZ lacY lacA
Lactose ( ) induces the expression of lac genes. If lactose is present, • enzymes for using lactose are needed • (allo)lactose binds to the lac repressor and causes a conformational change • repressor-lac does NOT bind to DNA • expression of lac genes is possible
CAPsite
Some repressor proteins mediate gene induction.
+
182Study objectives: Please study all the concepts and details of Regulation.1. Why do cells control gene expression? What is constitutive gene expression?2. What is gene induction? gene repression?3. Are catabolic genes more likely to be repressed or induced? Why?4. Are anabolic (biosynthetic) genes more likely to be repressed or induced? Why?5. What are the functions of the following in the regulation of transcription: repressor protein, activator protein, effector, co-repressor, inducer, activator binding site, operator, palindromic sequences, protein conformational changes? Understand the concepts and details. Do NOT memorize the specific palindromic sequences.6. Describe regulation of the trp operon and arg operon by repressor proteins.7. Describe the effect of lactose on the induction of the lac operon.8. Explain how repressor proteins can mediate gene repression. Explain how repressor proteins can mediate gene induction.9. Know that some activator proteins can mediate gene induction, while other activator proteins mediate gene repression
183Regulation of Gene Expression II:I. Activator proteinsII. Global regulationIII. Two-component regulatory systemsIV. Attenuation
184I. Activator proteins
eg. maltose activator proteincatabolite activator protein (CAP) cyclic AMP receptor protein (crp)
Proteins that activate transcription when bound to activator binding sites.
185
RNA polymerase
A. Typical activator protein
DNA
activator binding site
P
unusual promoter
TB
186B. Typical activator binding site
Imperfect palindrome
GTGAGTTAGCTCACCACTCAATCGAGTG
P O
TB
187C. Unusual promoters are involved in control by activator proteins.
-10consensus
(Pribnow box)
No-35
consensus
TB
188
1. In the lac operon, the activator protein is called the catabolite activator protein (CAP) or the cyclic AMP receptor protein (crp).
2. When cyclic AMP (cAMP) is present, the cAMP/CAP (crp) complex binds DNA and activates transcription.
CAP (crp)
cAMP
cAMP/CAPcomplex
binds DNA
D. The catabolite activator protein
TB
189
O
N
N N
N
NH2
CH2
OH
HOP=O
O
cyclic AMP (cAMP)cyclic adenosine monophosphate
(Don't memorize)
TB
190
P O lacZ lacY lacAcrp
P O lacZ lacY lacAcrpbinding site
Without activator protein, RNA polymerase binds weakly and the transcription rate is low.
With activator protein (crp), RNA polymerasebinds well and the transcription rate is higher.
Role of CAP (crp) in the lac operon
1913. MANY operons that encode catabolic enzymes have the same crp binding site ( ) and are controlled by the same regulatory protein (CAP or crp).
bacterial chromosome
crp binding site
192
Bacterial chromosome
operator oractivator binding sitesof similar DNAsequence
II. Global regulation A. Control of many genes by a single regulatory protein
TB
193B. Example: catabolite repressionA global regulatory system that allows glucose to be consumed in preference to a variety of other carbon sources.
TB
1941. Catabolite repression enables Escherichia coli to use glucose in preference to other carbon sources.
maltoseglucose
TCA
P O lacZ lacY lacAcrpbinding site
lac permease (transport protein)
lactose
-galactosidase
Lactose utilization requires additional proteins.
(galactose--1,4-glucose)
195
a. cAMP (cyclic AMP)an effector molecule that increases the DNA binding affinity of the catabolite activator protein
b. CAP (or crp) Catabolite activator protein, a transcriptional regulatory protein; also called crp (cAMP receptor protein)
2. Key components of catabolite repression
TB
196
CAP (or crp)
bacterialchromosome
CAP (or crp) binding sites
cAMP
cAMP/CAPcomplex
3. CAP/cAMP binds to DNA and regulates transcription.
TB
197
c. Without cAMP/CAP, genes required to catabolize nonglucose energy sources are transcribed at very low rates.
4. How does catabolite repression work? a. Genes needed for the catabolism of many carbon and energy sources require cAMP/CAP for expression.
*b. Glucose decreases cellular cAMP levels.
d. Therefore, glucose is preferentially used as a carbon and energy source.
198C. Global regulation is often used together with other more specific regulatory systems.
Example: the lactose operonrequires both lactose andcAMP/CAP for induction.
199
P O lacZ lacY lacAcrpbinding site
Both lactose and cAMP/CAP are needed for high induction of lac operon.
P O lacZ lacY lacAcrp
glucose absent, lactose present
glucose present, lactose absent
lac repressor binds DNA in absence of lactose
glucose decreases cAMP
200III. Two-component regulatory systems
Transcriptional regulatory systems composed of a sensor kinase andresponse regulator.
TB
201A. Sensor kinaseIntegral membrane proteins thatsense environmental conditions andphosphorylate proteins
B. Response regulator
Cytoplasmic transcriptional regulatoryproteins controlled by sensor kinasesthrough phosphorylation TB
202
cytoplasmic membrane
sensorkinase
effector
P
response regulator
PPdephosphorylation
phosphorylation
TB
203
Phosphorylation changes the DNA binding affinity of the response regulator.
C. Transcriptional control
When response regulators are bound to DNA, they induce or repress gene expression.
TB
204IV. Attenuation
A "fine tuning" system for regulating gene expression by control of transcriptional termination.
1 2 3 4UUUUU
conformation 2 intrinsictranscriptional
terminator
TB
205A. The trp operon is regulated at two levels.
P O E D C B A
1. repression by trp repressor (on/off)
genes encoding the enzymes used for tryptophan biosynthesis
2. attenuation (fine tuning by transcriptional termination)
R
R
206B. Leader region and leader peptide
P O E D C B A
leaderregion
tryptophan biosynthesis genes "structural genes"
tryptophan biosynthetic enzymesleader peptide
translation
transcription mRNA
207
leaderregion
coding region for trp enzymes
1. leader region of trp mRNAmRNA region upstream of the coding region for the trp biosynthesis enzymes
trp mRNATB
208
A short peptide encoded by the leader region of the trp mRNA.
trp leader mRNA (has 2 trp codons)
translation
leader peptide
2. leader peptide
met-lys-arg-ile-phe-val-leu-lys-gly-trp-trp-arg-thr-ser(Don't memorize sequence) TB
209
1. The leader region of the trp mRNA has four segments that can fold into 2 mutually exclusive conformations by complementary base pairing.
trp mRNA leader region
1 2 3 4
C. The trick to attenuation
TB
210mRNA leader region
1 2 3 4
conformation 1
2 31 4
1 2 3 4UUUUU
conformation 2 intrinsictranscriptional
terminator(3:4 loop) TB
2112. Conformation 2 of the trp mRNA leader is an intrinsic terminator.
Plentiful tryptophan favors conformation 2 and termination. Energy is not wasted making tryptophan when it is plentiful.
If conformation 2 is formed, transcription of the trp operon is terminated before the remainder of the trp mRNA is made.
1 2 3 4UUUUU
TB
212
1 2 3 42:3 stem loop mRNA
1 2 3 4UUUU
Intrinsicterminator(3:4 stem-loop)
3. The rate of TRANSLATION of the leader peptide determines which conformation (stem-loop) will form.
TB
213
a. When tryptophan is plentiful, translation of the trp leader peptide is FAST (i.e. ribosomes move fast).
4. The leader region encodes two tryptophans in a row.
Fast translation favors formation of the intrinsic terminator (the 3:4 loop). Transcription terminates before the structural genes are transcribed.
214b. When tryptophan levels are LOW, translation of the trp leader peptide is SLOW. The ribosome PAUSES at the trp codons, waiting for tryptophan-tRNA.
When the ribosome pauses, the 2:3stem-loop forms. The 3:4 intrinsic terminator stem-loop CANNOT form.Transcription of the trp biosynthesis genes continues.
215C. Transcription of the trp operon when tryptophan is plentiful.
1 Ribosome begins translation immediately after RNA synthesis occurs.
1 2 Ribosome finishes translation of the leader peptide and leavesthe mRNA.
21 Stem loop 1:2 forms. TB
2161 2 3 4 Transcription continues.
Stem loop 3:4 forms.
1 2 3 4UUUU
Stem loop 3:4 is an intrinsic terminator thatprevents furthertranscription.
UUUUU
P O E D C B A
leaderregion
tryptophan biosynthesis genes are NOT transcribed
X X X X X1 2 3 4
TB
217D. Transcription of the trp operon when tryptophan is low.
1 Ribosome begins translation immediately after RNA synthesis occurs.
1 2Because tryptophan is low, the ribosome pauses at tryptophancodons of the leader peptide and remains attached to the mRNA.
1 2 3Transcription continues.
TB
218
no terminator is formed
2 31The ribosome blocksbase pairing betweensegments 1 and 2.
42 31Segments 2 and 3 pairblocking the pairing of3 and 4.
Note that the alternative conformations of the trpleader mRNA are mutually exclusive. TB
219
no terminator is formed
P O E D C B A
tryptophan biosynthesis genes are transcribed
and translated
42 31
End result: When tryptophan levels are low, the genes for the tryptophan biosynthesis are expressed.
mRNA
DNA
proteins
220Study objectives: Please study both the concepts and details of Regulation.
1. What is an activator protein? How does it work? What is the catabolite activator protein or cAMP receptor protein (crp)? 2. What are the roles of cAMP, CAP and glucose in catabolite repression? 3. What is global regulation? Describe the example presented in class.4. How do the lac repressor system and cAMP/CAP system regulate expression of the lac operon? Understand (in detail) the effects of both lactose and glucose on the expression of the lac operon. 5. Describe how sensor kinases and response regulators function in two-component regulatory systems. 6. Understand the CONCEPTS and DETAILS of attenuation. What is the role of tryptophan, the leader peptide, the ribosome, and alternative leader mRNA conformations in trp operon attenuation?7. Compare and contrast (i) transcriptional regulation by regulatory proteins (ii) two-component regulatory systems and (iii) attenuation.
221
MCB 3020, Spring 2004
Chapter 8:Viruses
222Viruses:I. General properties of virusesII. Examples of virusesIII. Viral structureIV. Phage reproductionV. Reproduction of lysogenic phageVI. Overview of animal viruses
TB
223Typical viruses (30-200 nm)nucleic acid
helicalcapsid
envelope
icosahedral capsids
viralspecificproteins TB
224I. General properties of viruses A. small (~30-200 nm) B. non-cellular
TB
C. replicate within host cells and take over the host machinery D. released from the host cell and infect other cells virion = extracellular state of virus.E. often damage or kill the host
225
A. Human wart virusII. Some examples of viruses
Icosahedral symmetry (20 regular faces)TB
Picture18
226B. Tobacco mosaic virus
RNA virus
Helical symmetry
TB
Picture19
227C. Flu virus
enveloped virusTB
Picture20
228D. Lambda virus
host = a bacteriumbacterial viruses are also called
bacteriophage ("bacteria eaters") or phage
229III. Viral structure
A. genomesB. capsidsC. envelopesD. packaged enzymes
TB
230A. Viral genomes
All the hereditary material of a virus
dsDNAssDNAdsRNAssRNA
4 - 200 genes
TB
231B. Viral capsidsProtein shell that surrounds the genome
Protects the viral genomeOften needed for attachment to the host cellsUsually helical or icosahedral
capsid (protein coat)cross-section of
icosahedral capsidgenome
TB
232C. Viral envelopes
Composed of host lipids and viral proteins Often used for attachment to the host cell
lipids from host
viral proteins
TB
Outermost layer of enveloped viruses
233D. Packaged proteinsProteins found within the capsidDifferent functions in different viruses
viral proteine.g. reverse transcriptase RNA-dependent RNA polymerase TB
2341. Reverse transcriptase
enzyme that synthesizes DNA from an RNA template
2. RNA-dependent RNA polymerase
enzyme that synthesizes RNA froman RNA template
TB
235IV. Reproduction of phageA. AttachmentB. PenetrationC. Expression of viral genesD. Genome replicationE. Capsid formationF. PackagingG. Release
TB
236A. AttachmentBinding of a capsid or envelope protein to a host receptor.
host receptor
host cell
(usually a specific protein,lipid, or polysaccharide)
Specificity for the host receptordetermines virus host range TB
237Attachment and penetration
Virus tail fibers interacting with core polysaccharides
238B. penetration
injection ofviral nucleic acidand packaged
proteins.
TB
239C. Expression of viral genes
Viral proteins
viral genome
host machinery
TB
240
capsid proteinsproteins that block host gene expressionproteins that block restriction systems
Typical viral proteins
TB
proteins for genome replicationproteins for assembly of viral particles
241D. Genome replication
various methods: for example,host enzymes onlyviral enzymes onlyhost and viral enzymes
TB
242E. capsid formation self-assembly of capsid proteins
TB
243F. packaging
Insertion of the nucleic acid intothe capsid
Method varies
The "headfull" method is common
TB
244
1. Lysis G. Release
TB
2452. Budding (enveloped viruses)
viralproteins
host lipids
TB
246V. Reproduction of lysogenic phageA. lysisB. lysogenyC. prophage induction
TB
247A. LysisThe most frequent method of reproduction
Occurs as described above TB
248
bacterialchromosome
lysogen(cell with integrated virus)
integrationprophage
(integrated virus)
1. Prophage integration B. Lysogeny
TB
249
host replication
2. Prophage replication
TB
250C. Prophage induction
Excision of the prophagefollowed by lytic replication.
UV light and other DNA damaging agents cause prophage induction.
TB
251
binding to host receptor and uptake by endocytosis
1. Attachment and penetrationVI. Overview of animal viruses
animal celluncoatingTB
2522. Gene expression and genome replication for animal viruses must follow (or adapt to) eukaryotic rules
eukaryotic RNA processingcompartmentation (nucleus vs. cytoplasm)
TB
What features of transcription and translation would differ between phage and animal viruses?
253B. Host interactions
1. lysis2. persistent infection3. latent infection4. transformation
TB
2541. lysisdestruction of the host cell
2. persistent infectionviruses bud from host over a longperiod of time.
3. latent infectioninfections that reoccur periodically
4. transformationincreased growth rate of host cells TB
255Study objectivesPlease understand ALL the CONCEPTS and DETAILS presented in this lecture.
1. Describe the general properties of viruses.2. Define virions, bacteriophage, phage.3. Describe viral genomes, capsids (protein coats or shells), envelopes, and packaged proteins. What are the functions of these molecules? Know the specific examples presented in class.4. Compare and contrast the details of the reproductive cycles of phage and animal viruses. Thought question: What features of transcription and translation would differ between phage and animal viruses?5. How do phage reproduce by lysogeny?6. What is a lysogen? a prophage? 7. What effects can animal viruses have on their hosts?
256
I. Polio virusII. Flu virusIII. HIV virusIV. HIV replicationV. HIV treatmentVI. ViroidsVII. Prions
Eukaryotic viruses, viroids, and prions:
257I. Polio virus
A. Basic properties
nonenvelopedinfects nerve cells
+ssRNAicosahedral
258+ RNA (plus strand RNA) means that the RNA genome reads the same as the mRNA
mRNA 5' G G U U C C A A 3'
+ RNA 5' G G U U C C A A 3'
259
nerve cell
1. penetration and uncoating
nucleus of cell
uncoating+ssRNA
B. Life cycle
cytoplasm
2602. Genome replication
+RNA (genome)
-RNA
viral RNA-dependentRNA polymerase
2613. Gene expression
+RNA (mRNA) poliogenome
translation (host machinery)
polyproteinauto-proteolysis and proteolysis
coat proteins, proteases, RNA polymerase etc.
262a. Gene expression facts
Polio mRNA can be translated without a eukaryotic 5' cap (methylguanosine cap).
Polio inactivates translation of host mRNAs by destroying thehost protein that recognizes themethylguanosine cap.
2634. Assembly and release
+ strand RNAs are assembled intocapsids and the host cell is lysed.
264II. Flu virus
A. Basic properties
helical capsidinfects mucus membrane cells of the respiratory tract
-ssRNAsegmented genomeenveloped
265
neuraminidase
segmented genome(-RNA)
B. Structure
hemagglutinin
viral envelope
266
- RNA (minus strand RNA) is complementary to the mRNA
mRNA 5' G G U U C C A A 3'
- RNA 3' C C A A G G U U 5'
267C. Key proteins
1. Hemagglutininmediates fusion of the viral envelope to the host cell membrane
2. NeuraminidaseBreaks down sialic acid and assists in budding
268D. Antigenic shift
This can cause dramatic changesin surface antigens and produce new virulent strains.
Major changes in viral proteins dueto mixing of genome segments from different viruses.Occurs when two different viruses infect the same host.
269III. HIV (AIDS) virusHuman immunodeficiency virus
Causes AIDS
HIV kills CD4+ cells of the immune system
Healthy adults have about 800 CD4+ T-cells/cubic millimeter of blood.
HIV patients are said to have AIDS whenthey develop opportunistic infections or when their CD4+ T-cell count falls below 200.
270A. HIV infection
No cure
Almost always fatal
Usually acquired by sexual intercourse
No vaccine
In the US,~1/250 people are infected.
In the US,~1/3,000 people contract HIV each year
On average, 8-10 years pass between HIV infection and the development of AIDS.
271B. PreventionCelibacyInsistence on condoms Clean needles
4-weeks of treatment with possible side effects of headache, nausea, fatigue and anemia.
Post-exposure drug treatment within 24 h ???
272C. HIV replicationHIV is a retrovirus = an RNA virus that replicates through a DNA intermediate.
reversetranscriptase
HIV genome+ ssRNA(2 copies)
DNA
273D. HIV structure
envelope protein
reverse transcriptase
integrase
protease
+ssRNA
274
gag pol env other genes
LTR = long terminal repeat LTR
Genetic map of typical retrovirus
gag: encodes internal structural proteinspol: encodes reverse transcriptaseenv: encodes envelope proteinsThere are also other genes specific to different retroviruses.
275A. HIV proteins
1. Envelope protein: mediates binding to CD4 receptor
2. Reverse transcriptase: synthesizes DNA from an RNA template
2763. Integrase:
splices viral DNA into the host genome
4. Protease cleaves the viral polyprotein into active parts
277E. HIV reproductive cycle
CD4 receptor
cell membrane
HIV provirus
b
c
e
f
nuclearmembrane
ga
d
278
a. penetration and uncoatingb. reverse transcriptionc. integrationd. gene expression e. replication f. polyprotein cleavage by HIV proteaseg. assembly and budding
Steps in the HIV reproductive cycle
279F. HIV Treatment
http://www.hivatis.org/trtgdlns.html(The latest information on HIV treatment)
In general, two reverse transcriptaseinhibitors are used in combinationwith a protease inhibitor; however,treatment is complex and rapidly changing.
A. Reverse transcriptase inhibitorsB. protease inhibitors
280G. HIV drug resistance
HIV protease
proteaseinhibitor
inhibitor binding to the active siteinactivates the protease
mutation inhibitor nolonger binds butprotease stillfunctions
drugresistantprotease
281VI. Viroids
circular single stranded RNA molecules that cause plant diseases
viroids are "naked" RNA(no proteins associated with RNA)
viroid genomes do NOT encode proteins
282VII. Prions
Infectious proteins
Prion proteins appear to transmit disease without DNA or RNA.
283A. Prion diseases (spongiform encephalopathies)
Scrapie, sheep and goatsMad cow disease, cowsCreutzfeldt-Jacob, humans
284Mad cow disease (BSE)• Bovine spongiform encephalopathy (BSE)• source of infection appears to be feeding
cows with "meat-and-bone meal" remains of infected sheep or cows, especially infected brain tissue
• prion is not destroyed by cooking
285"new variant" Creutzfeldt-Jacob syndrome• human disease thought to be caused by eating BSE-infected beef• about 92 cases, most victims have died• unusual in that many victims are < 30 years old• incubation time is 10 to 15 years
286
normal PrP (prion protein)
disease causing PrP
Disease-causing PrP catalyzesa conformational change that turns normal PrP into disease causing PrP.
Over time, disease causing PrP accumulates and symptoms result.
B. How prions cause disease
287
If a protein transmits the disease, where is its gene?
The prion gene (prp) turned out to be anormal gene found in animals.
Unusual forms of the gene (mutants)are thought to cause disease.
C. The prion gene
288Study objectives1. Describe the structure of the polio virus. Explain polio virus replication and gene expression. What are polyproteins?2. Distinguish between plus strand and minus strand RNA genomes.3. Describe the structure of the flu virus. What is the relationship of flu virus genome structure to antigenic shift. 4. What are the functions of hemagglutinin and neuraminidase.5. How is HIV transmitted? 6. How is HIV infection prevented?7. How is HIV infection treated?8. Describe the structure of HIV. What is a retrovirus? Describe the general structure of a retroviral genome and the proteins encoded.9. Describe the HIV reproductive cycle. Know the functions of the HIV proteins.10. What are viroids? How do viroids differ from viruses and prions?11. What are prions?12. What diseases do prions cause?13. How are prions thought to cause disease? 14. Where are prions genes found?