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Flashcards in Exam 3 Deck (420):
1

Chromosomes have a versatile, modular structure for packaging DNA that supports flexibility of

form and function

2

__ is the generic term for any complex of DNA and protein found in a nucleus of a cell

chromatin

3

__ are the separate pieces of chromatin that behave as a unit during cell division

chromosomes

4

Chromatin is 1/3 __, 1/3 __, and 1/3 __

DNA, histones, nonhistone proteins

5

DNA interaction with __ and __ proteins produces sufficient level of compaction to fit into a cell nucleus

histones; nonhistone proteins

6

What are histones?

proteins that interact directly with DNA

7

How do histones interact with DNA?

histones neutralize DNA in the first level of compaction

8

The core histone complex makes up the

nucleosome

9

What are the five types of histones?

H1, H2A, H2B, H3, and H4

10

Of the five types of histones, which ones are core histones?

H2A, H2B, H3, and H4

11

160 base pairs of DNA wraps twice around a

nucleosome core

12

40 base pairs of linker DNA connect

adjacent nucleosomes

13

Which histone associates with linker DNA as it enters and leaves the nucleosome core?

H1

14

Diameter of DNA helix is

20 A

15

Diameter of nucleosome core is

100 A

16

Histones make up __ of all chromatin protein by weight

half

17

There are about 200-200,000 molecules of each kind of __ protein in chromatin

nonhistone

18

What are the functions of nonhistone proteins?

-structural role: chromosome scaffold
-chromosome replication: e.g. DNA polymerases
-chromosome segregation: e.g. kinetochore proteins
-transcription: largest group

19

The __ is the fundamental unit of chromosomal packaging

nucleosome

20

When DNA wraps twice around nucleosome core octamer, what does that result in?

a 7-fold compaction of DNA

21

How does spacing and structure of nucleosomes affect genetic function?

-determines whether DNA between nucleosomes is accessible for proteins to initiate transcription, replication, and further compaction
-arrangement along chromatin is highly defined and transmitted from parent to daughter cells during DNA replication

22

DNA must be condensed __ 7-fold

more than

23

What does the nucleosome do?

condenses naked DNA 7-fold to a 100 A fiber

24

What does supercoiling do?

causes additional 6-fold compaction of DNA, achieving a 40-50-fold condensation relative to naked DNA

25

What does the radical loop-scaffold do?

through progressive compaction of 300 A fiber, condenses DNA to rod-like mitotic chromosome that is 10,000 times more compact than naked DNA

26

100 A fiber is compacted into 300 A fiber by

supercoiling

27

What does the radical loop-scaffold model for higher levels of compaction state?

-several nonhistone proteins (NHPs) bind to chromatin every 60-100 kb and tether the 300 A fiber into structural loops
-other NHPs gather several loops together into daisy like rosettes

28

What is heterochromatin?

chromatin that is highly condensed, and usually inactive transcriptionally. Genes near heterochromatin have reduced expression or are "silenced"

29

When heterochromatin is constitutive what does that mean?

chromatin is condensed in all cells (e.g. most of the Y chromosome and all pericentromeric)

30

When heterochromatin is facultative what does that mean?

chromatin is condensed in only some cells and relaxed in other cells (e.g. position effect variegation, X chromosome in female mammals)

31

What is euchromatin and what is found in it?

relaxed chromatin that is usually transcriptionally active; housekeeping genes are found in this region (e.g. proteins that maintain cell function and are always expressed)

32

Transcription is controlled by

chromatin structure and nucleosome position

33

The more compacted DNA is,

the less transcription takes place

34

What are the three major mechanisms that can regulate chromatin patterns?

-histone modifications
-remodeling complexes
-histone variants

35

What are histone modifications?

the addition of methyl or acetyl groups

36

What do remodeling complexes do and how do they do it?

remodeling complexes can alter nucleosome patterns; they do it by
-changing accessibility of promoter sequences
-remove or reposition promoter-blocking nucleosomes

37

What can histone variants do?

they can cause different nucleosomal structures (e.g. CENP-A at centromeres)

38

Promoters of transcribed genes are located in

nucleosome free regions

39

Promoters of non-transcribed genes are wrapped in

nucleosomes

40

Origins of replication are also

devoid of nucleosomes

41

When transcription is required, promoters are exposed by

removing or repositioning nucleosomes

42

In histone modification and chromatin remodeling, the histone tails can undergo __ __ with chemical groups

covalent modification

43

In histone modification and chromatin remodeling, enzymes can add

chemical groups (methyl groups, phosphate groups, ubiquitin, etc.)

44

In histone modification and chromatin remodeling, modified tails can alter __ and bind __ __ __

nucleosomes; chromatin modifier proteins

45

In histone modification and chromatin remodeling, what does acetylation of lysines do?

-prevents close packing of nucleosomes
-favors expression of genes in euchromatin
-de-acetylation results in reduced transcription

46

In histone modification and chromatin remodeling, what does methylation of lysines and arginines do?

-can either close or open chromatin, depending on specific amino acid modified
-ex: adding methyl group to H3 lysine 9 favors heterochromatin formation
-de-methylation reverses

47

What is the rate of DNA synthesis in human cells?

about 50 nt/sec

48

Most mammalian cells have about __ origins

10,000

49

The human genome has about __ base pairs

3.2 billion (avg 70 million per chromosome)

50

It would take __ hours to replicate the human genome if there was only one origin of replication

800

51

T or F? Many origins are active at the same time

T

52

Nucleosomes are __ and __ during DNA replication

disassembled; reformed

53

DNA is packaged in nucleosomes within __ of synthesis

minutes

54

Chromatin fiber unwinds __ to synthesis

prior

55

Synthesis of histones (in cytoplasm) and transport into nucleus is tightly correlated with

synthesis of DNA

56

Newly synthesized DNA associates with

new histones

57

In very early embryo, both __ __ are active

X chromosomes

58

In humans, random X-inactivation occurs about __ __ after fertilization

2 weeks

59

Some cells have __ X inactivated, other cells have __ X inactivated

maternal; paternal

60

All cell descendants have the same

inactive X

61

Adult female calico cates are __ at X-linked genes

mosaic

62

In female calico cats heterozygous for X linked mutation:

-some cells have wild-type allele inactivated
-some cells have mutant allele inactivated

63

What is an example of facultative heterochromatin?

-dosage compensation in mammals so that X-linked genes in XX and XY individuals are expressed at same level
-random inactivation of all except one X chromosome in XX

64

What are Barr bodies?

darkly stained heterochromatin masses observed in somatic cells at interphase

65

An XX person has how many Barr bodies?

one

66

An XXX person has how many Barr bodies?

two

67

An XXY person has how many Barr bodies?

one

68

Chromosomes support the __, __, __, and __ of genetic info

packaging, replication, segregation, expression

69

What are chromosomal abnormalities characterized by a change in the number of chromosomes?

-aberrant euploidy
-aneuploidy

70

What are chromosomal abnormalities characterized by a change in the structure of chromosomes?

-deletion
-duplication
-translocation
-inversion

71

Chromosomes have distinct "banding patterns" from staining that can be used as

physical markers for locations of genes

72

On a chromosome, the short arm is called the __ arm

p

73

On a chromosome, the long arm is called the __ arm

q

74

What are the types of chromosomal rearrangements?

-deletion
-duplication
-inversion
-translocation

75

What is deletion?

the loss of a segment of a chromosome

76

What is duplication?

the gain of a segment of a chromosome

77

What is inversion?

the reversal of a region of a chromosome

78

What is translocation?

the movement of a segment of a chromosome among chromosomes

79

What is ploidy?

the basic number of chromosomes sets

80

What is euploidy?

the normal number of chromosomes within a cell for a species
-for ex., the euploid number of chromosomes in a human somatic cell is 46

81

What does haploid (n) mean? and what is an example of a type of cell that is haploid?

one chromosome set; this is the normal state for some cell type/organisms
-ex: human germ cells

82

What does diploid (2n) mean? and what is an example of a type of cell that is diploid?

two of the same chromosome set; this is the normal state for many organisms
-ex: human somatic cells

83

What does polyploid (>2n) mean?

more than two sets of chromosomes

84

What is transposition?

a type of sequence rearrangement with a significant genomic impact

85

What are transposable elements?

small segments of DNA that move from one position of DNA to another

86

Who discovered transposable elements?

Barbara McClintock with her study of mottling of corn color

87

What do retrotransposons do?

transpose (move their DNA) via reverse transcription of an RNA intermediate

88

What do transposons (a.k.a DNA transposons) do?

move their DNA directly without an RNA intermediate

89

What is a common mechanism retrotransposons use?

transcription by RNA polymerase into an RNA that encodes a reverse transcriptase-like enzyme.

90

The transcriptase-like enzyme can

copy RNA into a single strand of cDNA and then use that single DNA strand as a template for producing double stranded cDNA

91

Some retrotransposons have a __ __ at the 3' end of the RNA-like DNA strand, which is similar to mRNA molecules

poly-A

92

What are the retrotransposons in humans?

-LINES (long interspersed elements
-SINES (short interspersed elements)

93

What is the hallmark of DNA transposons?

that their ends are inverted repeats (mirror images) of each other
-these repeats are 10-200 bp long

94

DNA between the transposon's inverted repeats commonly contains a gene encoding __, a protein that catalyzes transposition through its recognition of those repeats

transposase

95

What is the DNA transposon mechanism?

1. excision of the transposon from its original genomic position
2. integration into a new location
3. the double-stranded break at the transposon's excision site is either
-repaired accurately
-the transposon will be lost from the original genomic site after transposition

96

__% of the human genome consists of transposable elements

44%

97

Approximately 90% of the transposable elements in the human genome are

retrotransposons

98

Of the 90% of the transposable elements in the human genome that are retrotransposons, 20% are __ and 13% are __

LINES; SINES

99

__% of the human genome consists of DNA transposons

3

100

Most of the transposable elements in the human genome are __ and cannot __

defective; move anymore

101

Insertion of a transposable element near or within a gene can affect __ and change __

expression; phenotype

102

Retrotransposon insertion mutations have been shown to cause about 100 know human diseases, including,

forms of hemophilia A, hemophilia B, cystic fibrosis, and muscular dystrophy

103

What is aneuploidy?

the loss or gain of one or more chromosomes

104

What are aneuploids?

individuals whose chromosome number is not an exact multiple of the haploid number (n) for that species

105

__ for any autosome is generally lethal

monosomy

106

__ for most autosomes is usually lethal, with a few exceptions

trisomy

107

Most organisms tolerate aneuploidy for

sex chromosomes

108

What are monosomic individuals?

individuals that lack one chromosome from the normal haploid number (2n-1)

109

What are trisomic individuals?

individuals that have one chromosome in addition to the normal diploid number (2n+1)

110

What are tetrasomic individuals?

organisms with four copies of a particular chromosome (2n+2)

111

How does aneuploidy occur?

chromosomal nondisjunction in meiosis

112

What is chromosomal nondisjunction in meiosis?

a process by which chromosomes or chromatids fail to separate during meiosis that results in gametes with an abnormal number of chromosomes

113

Chromosomal nondisjunction in meiosis usually results in the addition of loss of a __ chromosome

single
-resulting organism will have either 45 (one less) or 47 (one more) chromosome in its cells

114

T or F? Nondisjunction can occur during meiosis I or meiosis II

T

115

What happens in nondisjunction during meiosis I?

homologous pairs fail to separate during anaphase

116

What happens in nondisjunction during meiosis II?

sister chromatids fail to separate during anaphase

117

What are syntenic blocks?

colored segments that contain at least two genes whose order is conserved in the mouse genome

118

The human genome has about __ genes

25,000

119

The part of the genome corresponding to exons is the

exome

120

Most of a genome is non-coding DNA, what is it made of?

-exome (expressed regions) = about 2%
-introns
-centromeres, telomeres, transposable elements
-simple repeating sequences

121

What are gene-rich regions?

chromosomal regions that have many more genes than expected from average gene density over entire genome
-ex in humans: class III region of major histocompatibility complex (60 genes within 700 kb region)

122

What are gene deserts?

regions of >1 Mb that have no identifiable genes
-3% of human genomes is comprised of gene deserts

123

T or F? Biological significance of gene-rich regions and gene deserts is not known

T

124

Exons often encode __ __

protein domains (sequence of amino acids that fold into functional units)

125

Shuffling, addition, and deletion of domain regions can produce new __ in cells and organisms

functions

126

Reorganization of domain provides raw material for

evolution

127

After exon shuffling, protein products have novel

domain architectures

128

Gene families can evolve by __ followed by __

duplication; divergence

129

What are gene families?

groups of genes that are closely related in sequence and function
-ex: hemoglobin genes (alpha and beta globes), immunoglobins (antibodies)

130

Changes in number and arrangement of exons can alter

functions

131

Duplication and divergence of genes can create genes with both __ and __ functions

new; old

132

Rearrangements and duplications create many possibilities for

novel functions

133

Virtually all knowledge of gene structure, expression, and regulation came from studies of

bacteria and bacteriophages

134

The advent of recombinant DNA technology depended on understanding of

bacterial genes, chromosomes, and restriction enzymes

135

All bacteria are __, which lack a defined nuclear membrane

prokaryotes

136

All bacteria lack

membrane-bound organelles

137

Most bacteria have a cell wall made of __ that surrounds the cell membrane

carbohydrate and peptide polymers

138

Bacteria have a single

chromosome

139

Bacteria divide __

rapidly (1 hour in minimal medium, 20 min. in high nutrient conditions)

140

__ is the most studied and best understood bacterial species

E.coli

141

E.coli inhabits the intestines of

warm-blooded animals

142

E.coli can grow in

complete absence of oxygen or in air

143

E.coli are phototrophic, meaning

they can grow in minimal media
-single carbon and energy source (e.g. glucose)
-inorganic salts

144

The E.coli genome is tightly packed with

genes

145

Describe the genome of the K12 strain of E.coli that was sequenced

-4.6 Mb
-about 90% of genome encodes protein
-4288 genes, but function known for only 60%
-on average, 1 gene per kb
-no introns
-very little repetitive DNA
-small intergenic regions

146

Individual E.coli strains contain a subset of the E.coli __

pangenome

147

What is the core genome of E.coli?

about 1000 genes that are found in all strains

148

What is the pangenome of E.coli?

the core genome plus all genes that are found in some strains but not others (about 15,000 genes)

149

The typical bacterial genome is composed of one circular __

chromosome

150

In bacteria, the DNA molecule condenses by

supercoiling and looping

151

Each bacterium replicates and then divides by __ __ into two daughter cells

binary fission

152

__ __ elements dot the genomes of many types of bacteria

insertion sequences (IS)

153

What are insertion sequences and what do they do in bacteria?

small transposable elements
-inverted repeats (IRs) at ends
-carry transposase gene, which initiates transposition by recognizing IRs
-can move to other locations in genome
-can disrupt genes by insertion into coding regions (cause of many spontaneous mutations)

154

Tn elements in bacteria are __ __ __

composite transposable elements

155

Tn elements contain

transposase gene and genes conferring resistance to antibiotics or toxic metals

156

What are plasmids?

smaller circles of DNA that carry genes beneficial to the host cell

157

Plasmids don't carry genes essential to the host, but may

benefit the host under certain conditions

158

What are some examples of genes that are beneficial to the host

-genes that protect host against toxic chemicals (e.g. mercury) and metabolize environmental pollutants (e.g. toluene, petroleum products)
-pathogenic genes (e.g. toxins produced by S. dysenteriae)
-genes encoding resistance to antibiotics
-multiple antibiotic resistance often due to composite IS/Tn elements on a plasmid

159

Movement of antibiotic resistance genes TO the plasmid was facilitated by

transposons

160

Multiple antibiotic resistance genes can be transposed from the plasmid as a

unit

161

Bacteria must be grown and studied in

cultures

162

What are some examples of mutant variation in bacteria?

-altered colony morphology
>large or small; shiny or dull; round or irregular
-resistance to bactericides
>antibiotics, bacteriophages (e.g. MRSA!)
-Auxotrophs: unable to reproduce in minimal media
-defective in using complex chemicals from the env
>ex: breaking down lactose into glucose and galactose
-defective in proteins essential for growth
>conditional lethal mutations, e.g. temp-sensitive (ts)

163

Rapid bacterial multiplication allows for detection of

very rare genetic events

164

What does effectively haploid mean?

straightforward relationship b/w mutation and phenotypic variation

165

What happens in selection?

est conditions in which only the desired mutant will grow

166

What happens in a genetic screening?

examine each colony for a particular phenotype using a technique called replica plating

167

Genomic analysis has revealed widespread occurrence of __ __ __ in many bacterial species

gene transfer mechanisms

168

Gene transfer is an important mechanism for __ __ __ __ __ and to development of pathogenic strains of bacteria

rapid adaptation to environmental changes

169

Describe lateral (or horizontal) gene transfer

traits are introduced from unrelated individuals or from different species

170

Describe vertical gene transfer

-occurs in sexually reproducing organisms
-traits are transferred from parent to offspring

171

In the three mechanisms for gene transfer in bacteria,

-donor bacterium provides the DNA that is transferred
-recipient bacterium receives the DNA, which can result in altered phenotype

172

In conjugation, the F plasmid contains genes for

synthesizing connections between donor and recipient cells

173

What is conjugation?

direct transfer of DNA from donor cell to connected recipient cell

174

Donors for conjugation are __

F+ (carry a special F plasmid)

175

Recipients for conjugation are __

F- (don't carry an F plasmid)

176

F plasmid has three

IS elements

177

__ __ __ cells are formed when an F plasmid integrates into the bacterial chromosome through recombination between IS elements

high frequency recombinant (Hfr)

178

20-30 different Hfr strains can be generated that differ in the __ and __ of the integrated F plasmids

location; orientation

179

Integrated F plasmid replicates with __ during cell division

chromosome

180

Her strains retain all __ __ __ and can be a donor for conjugation with an F- strain

F plasmid functions

181

Transfer of DNA starts in the F plasmid at the

origin of transfer

182

Chromosomal genes located next to F plasmid sequences are transferred to the

recipient

183

Transferred chromosomal DNA recombines into __ __ in recipient

homologous DNA

184

Usually conjugation terminates before

entire chromosome transfers

185

How are F' plasmids created?

-an Hfr plasmid comes out of a bacterial chromosome
-a few chromosomal genes will be removed with it, generating an F' episome (plasmid)

186

What are merodiploids?

partial diploids in which two gene copies are identical

187

What are merodiploids useful for?

complementation testing

188

To select for Trp+ transformants, plate on minimal media with

histidine and no tryptophan

189

To select for His+ transformants, plate on minimal media with

tryptophan and no histidine

190

To screen for His+ Trp+ co-transformants, test Trp+ individual
transformants and His+ individual transformants for growth on
minimal media with

neither tryptophan nor histidine

191

What are the questions that represent the challenges of gene regulation?

-what should be expressed?
-when should a gene be expressed?
-where should a given gene be expressed?
-how much of a protein is needed, so what level of expression is needed?

192

RNA polymerase participates in all three phases of

transcription

193

During initiation, what is present?

core RNA polymerase plus sigma (σ) factor

194

During elongation, what is present?

core RNA polymerase without σ factor

195

Most promoters are __ to the transcription start point

upstream

196

RNA polymerase makes strong contacts at __ and __

-10, -35

197

Core RNA has four subunits, what are they?

two alpha (α), one beta
(β), one beta prime (β')

198

What happens during initiation?

DNA is unwound and polymerization begins

199

Elongation continues until

RNA polymerase recognizes termination signal

200

What are the two kinds of transcription termination in bacteria?

Rho-dependent and Rho-independent

201

Describe Rho-dependent termination

Rho protein binds to RNA polymerase and removes it from RNA

202

Describe Rho-independent termination

20 nt sequence in RNA forms stem-loop

203

What are examples of transcriptional control of gene expression?

-binding of RNA polymerase to promoter
-Shift from initiation to elongation
-Release of mRNA at termination

204

Binding of RNA polymerase to promoter is the most critical step in

regulation of most prokaryotic genes

205

What are some examples of post transcriptional control of gene expression?

-stability of mRNA
-efficiency of translation initiation
-stability of polypeptide

206

Regulation of transcription requires __ that __

regulatory proteins; modify ability of RNA polymerase to recognize and bind promoter

207

Proteins bind to __ to regulate transcription

DNA

208

Gene regulation changes with __ context

environmental

209

Positive regulation __ transcription

enhances

210

Positive regulatory proteins help __ transcription

activate

211

Negative regulation __ transcription

inhibits

212

Negative regulatory proteins help __ transcription

block

213

Utilization of lactose by E.coli provides a __ __ of gene regulation

model system

214

What are the two enzymes required for lactose utilization?

permease and β-Galactosidase

215

What does permease do in lactose utilization?

transports lactose into cell

216

What does β-Galactosidase do in lactose utilization?

splits lactose into glucose and galactose

217

In the absence of lactose, both permease and β-Galactosidase are present at __ __ levels

very low

218

Cells prefer to use __ as an energy source

glucose

219

In a medium with both lactose and glucose, bacteria will choose __ first, until it's gone

glucose

220

Lactose is the __ of the genes encoding permease and β-Galactosidase

inducer

221

What is induction?

stimulation of synthesis of a specific protein

222

What is an inducer?

molecule responsible for induction

223

What are the advantages of using lactose utilization by E.coli as a model for understanding gene regulation?

-Lac- mutants survive and can be maintained on media with glucose and so lac genes are not essential for survival (you can keep your mutants alive)
-simple says for lac expression
-lactose induces a 1000-fold increase in β-Galactosidase activity
-lots of progeny

224

What does the operon theory state?

one signal can simultaneously regular expression of several clustered genes

225

Jacques Monod and Francois Jacob hypothesized that lac genes are transcribed together as a single mRNA (polycistronic) from a single

promoter

226

What are the three structural genes of the lactose operon?

lacZ, lacY, and lacA

227

What is a promoter?

site to which RNA polymerase binds

228

What does the cis-acting operator site do?

controls transcription initiation

229

What does the trans-acting repressor do?

binds to the operator (encoded by lacI gene)

230

What does an inducer do?

prevents repressor from binding to operator

231

What is the lac-operon?

a cluster of genes transcribed simultaneously (lacZ, lacY, lacA)

232

Describe lac-operon induction when lactose is present

1. inducer binds repressor
2. repressor changes shape and cannot bind to operator
3. RNA polymerase binds to the promoter and initiates transcription of the polycistronic lac mRNA

233

Repression of lac gene expression occurs in the __ of lactose

absence

234

Lac repressor is a __ regulatory element

negative

235

lacZ encodes

β-Galactosidase

236

lacY encodes

permease

237

lacA encodes

transacetylase

238

lacA is not necessary for

lactose catabolism

239

lacZ and lacY mutations led to new phenotype, what is it?

inability to utilize lactose

240

If cells grow in lactose medium, the mutations

complement

241

If cells do not grow in lactose medium, mutations

fail to complement and are in the same region

242

What is characteristic of lacZ-?

-inability to produce β-Galactosidase
-no induction of β-Galactosidase activity by lactose inducer

243

What is characteristic of lacY-?

-inability to produce permease
-no induction of permease activity by lactose inducer

244

What is characteristic of lacI-?

constitutive expression of β-Gal and permease

245

__ __ express the enzymes in the absence and presence of inducer

constitutive mutants

246

Why must lac I be a repressor?

cells require lac I protein to prevent expression of lac Z and lac Y in absence of an inducer

247

The PaJaMo experiment provided evidence that lacI encodes a

repressor

248

Jacob and Monod proposed that lacI encodes a repressor that binds to an operator site near the __ promoter

lac

249

What does the binding of an inducer to a repressor do?

changes the shape of the repressor so that it can no longer bind to DNA

250

When there is no inducer present, the repressor is able to

bind to DNA

251

Repressor is an allosteric protein, meaning that

it undergoes reversible changes in conformation when bound to another molecule

252

lacI- mutants have a __ __ that cannot bind to operator

mutant repressor

253

lacI(s) mutants have a __ that binds to operator but can't bind to the inducer

superrepressor

254

In lacI(s) mutants, lac genes are __ in the absence and the presence of inducer

repressed

255

lacO(c) mutants have a mutant operator that can't bind the __

repressor

256

In lacO(c) mutants, lac genes are __ in the absence and the presence of inducer

expressed

257

What are the 2 ways to get constitutive expression?

1. mutation in lacI (lacI-) that prevents the repressor from binding the operator whether lactose is present or not
2. Mutation in operator DNA sequence (lacO(c)) the prevents repressor from recognizing and binding

258

How can you distinguish between the two ways to get constitutive expression?

using the cis/trans test with merodiploids

259

Describe trans-acting elements

can diffuse through the cytoplasm and act at target DNA sites on any DNA molecule in the cell

260

Describe cis-acting elements

can only influence expression of adjacent genes on the same DNA molecule (operator, promoter, etc)

261

Cis DNA elements need to be on same chromosome as

genes they regulate

262

Trans elements are proteins produced on one molecule that can interact with

DNA on either molecule

263

If lacI(s) is cis-acting, then lacZ+ chromosome will be

inducible

264

If lacI(s) is trans-acting, then lacZ+ will be

non-inducible

265

lacI(s) protein acts in

trans

266

lacO(c) acts in

cis

267

The lacO(c) mutation affects expression of genes only on

the DNA that it is located on

268

Why does the O+ operator have no effect in the lacO(c) mutation?

because it is adjacent to a mutant lacZ on the plasmid and can't impact the lacZ+ on chromosome

269

Initiation of transcription under control of regulatory genes whose protein products bind to DNA near promoter alter

RNA polymerase

270

What is the biochemical evidence for lac repressor binding to lacO?

-lac repressor has two separate domains
-lac repressor has a helix-turn-helix (HTH) motif
-most DNA-binding regulatory proteins are oligomeric (one domain), with two to four subunits

271

Mutated DNA sequences in many different lacI- mutants are clustered in the __ __ domain of lac repressor

DNA-binding

272

Mutated sequences in many different lacI(S) mutants are clustered in the __ __ domain of the lac repressor

inducer-binding

273

A protein with an HTH (helix-turn-helix) motif has two __ __ separated by a turn in the protein

α-helical regions

274

The HTH motif fits into the major groove of

DNA

275

One of the α-helical regions of a protein with an HTH motif has amino acids that 'recognize' a specific

DNA sequence

276

HTH motifs are found in many __ __ proteins

DNA-binding

277

lac repressor tetramer binds to __ sites

two

278

lac repressor is a tetramer, with each subunit containing a DNA-binding __ __

HTH motif

279

Two repressor subunits of lac repressor bind to

O1

280

Two repressor subunits of lac repressor bind to either __ or __

O2 or O3

281

The binding of the repressor subunits of lac repressor to O1, O2, and O3 causes the formation of a loop that leads to

highly efficient repression

282

When lac repressor is bound to lac operator, functional binding of RNA polymerase to the promoter is

blocked

283

Many negative regulators (e.g. lac repressor) prevent transcription initiation by blocking

the functional binding of RNA polymerase

284

Many positive regulators (e.g. CRP-cAMP) establish contact with RNA polymerase that

enhances transcription initiation

285

The lac operon of E.coli is regulated by both __ and __

lactose; glucose

286

When both glucose and lactose are present, only __ is utilized

glucose

287

Lactose induces __ __ __, but only in the absence of glucose, even if lactose is present

lac mRNA expression

288

Positive regulation increases transcription of lacZ, lacY, and lacA only when

lactose is present and glucose is absent

289

Lactose prevents repressor from binding to

lacO

290

lac repressor is a __ regulator of lac transcription

negative

291

lac mRNA expression cannot be induced if

glucose is present

292

Glucose controls the levels of

cAMP

293

cAMP binds to __ __ __

cAMP receptor protein (CRP)

294

CRP-cAMP is a __ regulator of lac transcription, but ONLY in absence of glucose

positive

295

Describe catabolite repression

overall effect of glucose is to prevent lac gene expression by limiting availability of cAMP

296

CRP-binding sites have a two-fold __ __

rotational symmetry

297

CPR protein binds as a

dimer

298

CRP-binding site consists of two recognition sequences, one for each

subunit of the CRP dimer

299

CRP-cAMP complex makes direct contact with

RNA polymerase

300

Without interaction with CRP-cAMP, RNA polymerase can bind to the promoter but is less likely to __ __ __ __ __

unwind DNA and initiate transcription

301

What is a reporter gene?

protein-encodind gene whose expression in the cell is quantifiable by sensitive and reliable techniques

302

How do you measure gene expression?

-fuse coding region of lacZ to cis-acting regulatory regions from other genes
-conditions that induce expression of gene of interest will generate β-gal

303

How do you control gene expression?

-fuse the lac regulatory sequences to the coding region of a foreign gene
-inducible expression of the foreign gene controlled by IPTG

304

lacZ fusion is used to perform genetic studies of the regulatory region of

gene X

305

Conditions that regular expression of the test regions from gene X will alter the levels of

β-galactosidase

306

Specific regulatory sites can be identified by constructing and testing mutations in the test regions of

gene X

307

Expression of gene X is under the control of the

lac regulatory system

308

Expression of human growth hormone in E.coli is controlled by

lac control region

309

How do eukaryotes use complex sets of interactions?

-regulated interactions of large networks of genes
-each gene has multiple points of regulation
-turning on and off genes in right place ad time

310

What are the themes of gene regulation in eukaryotes?

-environmental adaptation, growth, and division in prokaryotes
-maintenance of homeostasis in multicellular eukaryotes
-genes are turned on and off in right place and time
-differentiation and precise positioning of tissues and organs during embryonic development

311

Eukaryotic genomes are __ than prokaryotic genomes

larger

312

Compared to prokaryotes, eukaryotes have additional

levels of complexity for controlling gene expression

313

__ __ in eukaryotes makes DNA unavailable to transcription machinery

chromatin structure

314

Additional __ __ events occur in eukaryotes

RNA processing

315

In eukaryotes, transcription takes place in __ and translation takes place in __

the nucleus; the cytoplasm

316

Both eukaryotes and prokaryotes utilize DNA binding proteins for

transcriptional regulation

317

There are multiple steps where production of the final gene product can be regulated in

eukaryotes

318

Eukaryotes can regulate these steps to control __ __ in different tissues

cell differentiation

319

What does RNA polymerase I do?

transcribes genes that are the major RNA components of ribosomes (rRNAs)

320

What does RNA polymerase II do?

transcribes genes that encode all proteins

321

What does RNA polymerase III do?

transcribes genes that encode the tRNAs and certain other small RNA molecules

322

RNA pol II catalyzes synthesis of the __ __, which is complementary to the template strand of the gene

primary transcript

323

Most RNA pol II transcripts undergo further processing to generate __ __

mature mRNA

324

What does RNA splicing do?

removes introns

325

What does addition of 5' GTP cap do?

protects RNA from degradation

326

What are the further processes that RNA pol II transcripts must undergo to generate mature mRNA?

-RNA splicing
-addition of 5' GTP cap
-cleavage of 3' end by ribonuclease, and addition of 3' polyA tail (poly-A polymerase)

327

Promoters are usually adjacent to

the protein-coding gene

328

Promoters include the transcription initiation site and often have

about 7 base pair TATA box

329

Binding of RNA pol II allows __ __ of transcription

basal level

330

Describe enhancers

can be distant (10,000s of bps) from gene, or even within gene introns or reversed in orientation

331

Binding of proteins can augment or repress __ __

basal transcription

332

Trans-acting factors interact with cis-acting elements to control rates of

transcription initiation

333

How do the direct effects of transcription factors come about?

-through binding to DNA
-basal factors
-activators and repressors

334

How does the indirect effect of transcription factors come about?

through protein-protein interactions

335

Basal transcription factors assist the binding of RNA pol II to

promoters

336

What are the key components of the basal factor complex?

-TATA box-binding protein (TBP)
-binds to TATA box
-first of several proteins to assemble at promoter
-TBP-associated factors (TAFs)
-bind to TBP assembled at TATA box

337

Basal factors bind to promoters of all __ __ __

protein-encoding genes

338

What is the ordered pathway of assembly at promoter?

1. TBP binds to TATA box: produces "bend" in DNA of TATA box
2. TAFs bind to TBP
3. RNA pol II binds to TAFs

339

RNA pol II associates with basal complex and initiates

basal level of transcription

340

Activators are transcription factors that bind to

enhancers

341

What is the significance of activators?

binding of different activators to enhancers is responsible for much of the variation in levels of transcription of different genes in different cell types

342

How do activators increase levels of transcription?

by interacting directly or indirectly with basal factors at the promoter

343

Binding of activators to enhancers increases

transcriptional levels

344

Low level transcription occurs when only __ __ are bound to promoter

basal factors

345

When basal factors AND activators are bound to DNA, rate of transcription __

increases

346

What are the two functional domains of activator proteins?

1. sequence-specific DNA binding domain (bind enhancer)
2. transcription-activator domain (which binds other trans-factors)

347

__ __ leads to physical interaction of distant DNA regions

DNA looping

348

What are the mechanisms of activator effects of transcription?

-stimulate recruitment of basal factors and RNA pol II to promoters
-recruit coactivators to open chromatin structure

349

What are the effects of activators on transcriptions?

-stimulate recruitment of basal factors and RNA pol II
-stimulate activity of basal factors
-facilitate changes in chromatin structure

350

What do the DNA-binding domains of activator proteins do?

interact with major groove of DNA
-certain amino acids have high-affinity binding to specific nucleotide sequence

351

What are the three best-characterizes motifs?

1. helix-loop-helix (HLH)
2. helix-turn-helix (HTH)
3. zinc finger

352

__ __ __ are activators, but only in the presence of specific hormones

steroid hormone receptors

353

Steroid hormones don't bind directly to DNA but are __ of steroid hormone receptors

coactivators

354

In the absence of hormone, steroid hormone receptors can't bind to DNA and can't

activate transcription

355

In the presence of hormone, steroid hormone receptors bind to enhancers for specific genes and activate

expression

356

T or F? Not all transcription factors activate gene expression

T

357

When a repressor binds to the same enhancer sequence as the activator, what effect does it have on the basal transcription level?

no effect

358

Describe quenchers

bind to the activator but do not bind to DNA; the repressor blocks the activator from functioning

359

Some repressors eliminate virtually all basal transcription from a promoter by blocking

promoter access

360

In humans, about __ genes or __% of genes encode transcriptional regulatory proteins

2000; 10%

361

Each regulatory protein can act on __ genes

many

362

Each regulatory protein can have __ of enhancers

dozens

363

The same transcription factors can be an activator or a repressor depending on

-the cell that it's in
-which cis-regulatory element it binds to

364

Regulatory proteins have vast complexity for precise control of

gene expression

365

What are the two methods that cells use to regulate transcription?

-binding of transcription factors to enhancers
-DNA methylation

366

What does binding transcription factors to enhancers do?

modulates the spatial and temporal expression of many genes that are expressed only in particular tissues at specific times during development

367

Describe methylation

(a biochemical modification of DNA itself)
-a methyl (CH3) group is added to the 5th carbon of the cytosine base in a 5' CpG 3' dinucleotide pair on one strand of the double helix

368

DNA methylation is important for controlling expression of

"housekeeping genes"

369

DNA methylation also regulates some

cell-type specific genes

370

Why can DNA methylation alter gene expression heritability without changing the base sequence of DNA

because methylation affects transcription levels, and methylation patterns are copied during DNA replication (this is called the epigenetic phenomenon)

371

Methylation is key to epigenetic in mammals and is called

genomic imprinting

372

What organisms have no DNA methylation?

C. elegans and yeast

373

What organisms have very little DNA methylation?

other invertebrates and lower euks

374

DNA methylation at CpG islands __ gene expression

silences

375

DNA methylation usually __ the transcription of eukaryotic genes

inhibits

376

DNA methylation usually inhibits the transcription of eukaryotic genes particularly when it occurs in the vicinity of the

promoter

377

In vertebrates and plants, CpG islands occur near many

promoters of genes

378

CpG islands are commonly __ in length and contain a high number of __

1000 to 2000 bp; CpG sites

379

DNA methylation is thought to play an important role in the silencing of tissue-specific genes to prevent

them from being expressed in the wrong tissue

380

Nucleosomes can make promoters __

inaccessible

381

What are epigenetic changes?

changes in chromatin structure that are inherited from one generation to the next
-DNA sequence is not altered
-but particular cells with altered chromatin will have altered gene expression, which can be inherited from one cell generation to the next

382

Chromatin reduces binding to basal factors and RNA pol II to

very low levels

383

Nucleosomes can be __ or __ by chromatin remodeling complexes

repositioned or removed

384

After remodeling, DNA at promoters and enhancers becomes __ __ to transcription factors

more accessible

385

Transcription is active near __ CpG islands

unmethylated

386

CpG islands are regions with a high concentration of

CpG dinucleotides

387

Near genes, CpG islands are usually unmethylated because

an activator binds and blocks access by DNMTs
-the chromatin is open and transcription is activated

388

DNA methylation at CpG islands __ gene expression

silences

389

In the absence of activators, the CpG islands become

methylated

390

Methyl-CpG-binding proteins (MeCPs) binds and close the

chromatin structure

391

Gene expression repression is often long-term because

the methylation pattern is maintained through numerous cell divisions

392

Long-term repression through DNA methylation is called

silencing

393

DNA methylation is an __ __ because it can heritably alter gene expression without changing DNA sequence

epigenetic phenomenon

394

Cytosine methylation pattern is copied during

DNA replication

395

DNA methylation patterns are copies during DNA replication by

a special DNMT present at the replication fork
-this DNMT recognizes semi-methylated DNA (on the parent strand) and methylates the newly synthesized strand

396

Sex-specific DNA methylation is responsible for

genomic imprinting

397

What is the Mendelian rule?

parental origin of allele does not affect F1 phenotype (usually!)

398

Describe genomic imprinting

expression of a gene depends on whether it was inherited from the mother or father
-epigenetic effect (no change in DNA sequence)

399

__ __ __ is transcriptionally silenced if it was transmitted from the father and the maternally inherited allele is expressed

paternally imprinted gene

400

__ __ __ is transcriptionally silenced if it was transmitted from the mother and the paternally inherited allele is expressed

maternally imprinted gene

401

imprinted =

silenced

402

Clinical geneticists realized that genomic imprinting existed long before molecular biology techniques made it possible to confirm its existence, how?

pedigree analysis of rare diseases
-the patterns were clear in certain rare cases where the condition was caused by a deletion that removed the imprinted gene
-the inheritance of the deletion and the disease could be followed in karyotypes

403

When examining a pedigree chart of an imprinted gene, the sex of the parent carrying a mutant allele determines offspring __, not sex of the offspring!

phenotype

404

If there is a deletion of a paternally imprinted autosomal gene.. fathers can pass the deletion to their sons and daughters and they will not be affected because

the child's wild-type maternal allele would be expressed

405

The genomic imprint is ___ during mitosis

maintained

406

Patterns of DNA methylation must be __ during meiosis before being passed on to the next generation

reset

407

Imprinting is

sex-specific

408

Methylation is removed in

germ-line cells

409

Imprinting occurs in the __ __ and is accompanied by heavy __

germ line; methylation

410

Epigenetic imprints are erased during __ __ __ and reset by __ __ __

germ-line development; sex-specific patterns

411

Complementary base pairing between a small RNA and mRNA can prevent

gene expression

412

What are the 3 classes of small regulatory RNAs that have been identified

1. micro-RNAs (miRNAs)
2. small interfering RNAs (siRNAs)
3. Piwi-interacting RNAs (piRNAs)

413

Each small RNA class leads to the production of

single-stranded RNAs of slightly different lengths

414

Each small RNA class is in the range of __ nucleotides

21-30

415

What are the targets of miRNAs?

mRNAs

416

What are the effects of miRNAs?

-block mRNA translocation/ destabilize mRNAs

417

What are the targets of siRNAs?

mRNAs

418

What are the effects of siRNAs?

block translation/ destabilize mRNAs

419

In order to prevent gene expression, small RNAs work with proteins in the __ family

Argonaute
-form ribonucleoprotein complexes

420

The small RNA in each ribonucleoprotein complex guides the complex to

particular nucleic acid
-this nucleic acid target will have perfect or partial complementarity with the small RNA
-the mRNA will not be expressed in most cases
-this is a type of post transcriptional RNA modulation