Chapter 12: Gene Transcription and RNA Modification

Gene

A segment of DNA used to make a functional product either an RNA or a polypeptide.

Transcription

The process of copying a segment of DNA sequence into RNA sequence.

Translation

The process by which the sequence of an mRNA is converted into the sequence of amino acids in a protein.

Gene expression

The process by which information from a gene is used to synthesize a functional gene product (usually a protein).

Protein-coding genes (or structural genes)

Genes that code the amino acid sequence of a polypeptide.

Polypeptide vs. Protein

Polypeptide is a structural unit, protein is the functional unit.

The synthesis of functional proteins determines an _____.

organism’s traits.

The Central Dogma of Genetics

(1) DNA replication of chromosomal DNA (2) Transcription to create mRNA (3) Translation producing a polypeptide from mRNA and it becomes a functional protein contributing to an organism’s traits.

DNA base sequences define __________.

The beginning and end of a gene and regulate the level of RNA synthesis.

Proteins must _____.

Recognize and act on DNA for transcription to occur.

Terminator

The DNA sequence where transcription ends.

Template Strand

The DNA strand that is transcribed into RNA. The RNA transcript is complementary to the template strand.

Coding Strand

The opposite DNA strand of the template strand. The base sequence in RNA is identical to the coding strand of DNA, except uracil is substituted for thymine.

Transcription factors

Proteins that bind to DNA and regulating the rate at which genetic information is transcribed into mRNA.

Ribosomal-binding site and codons

mRNA sequences that control translation

transcription factors can bind directly to _____ or _____.

the promoter and regulatory elements.

transcription factors can either _____ or ____ transcription.

facilitate or inhibit

the three stages of transcription:

initiation, elongation, temination

Initiation

The first step in transcription. The promoter functions as a recognition site for transcription factors. The factors enable RNAP to bind to the promoter. Following binding, the DNA is denatured into an open transcription bubble.

Elongation

RNAP slides along the DNA in an open complex to synthesize RNA.

Termination

The final step in transcription. A terminator is reached that causes RNA polymerase and the RNA transcript to dissociate from the DNA.

Promoter

The DNA sequence/site in DNA where transcription begins. They “promote” gene expression. They are typically located just upstream of the site where transcription begins.

The bases in a promoter sequence are numbered in relation to _____.

the transcription start site.

the first nucleotide that acts as a template for transcription is designated as ____.

+1

the numbering of nucleotides to the left of +1 _____.

negative. often referred to as upstream

the nucleotide to the immediate left of the transcription start site (+1) is designated ____.

-1

Is there a nucleotide numbered 0?

No

The numbering of nucleotides to the right of transcription start site (+1) are _____.

positive. often referred to as downstream.

-35 and -10 sequence in promoters

Key regions in bacterial promoters that are necessary to create a functional promoter.

Pribnow box

Another term for -10 sequence.

Consensus sequence

The most common sequence. It is likely to result in a high level of transcription.

RNAP

The enzyme that catalyzes the synthesis of RNA.

E. coli RNAP holoenzyme

Composed of the core enzyme and sigma factor

sigma factor

Recognizes the promoter and initiates transcription

Initiation of bacterial transcription

RNAP holoenzyme binds loosely to the DNA. It then scans along the DNA, until it encounters a promoter. When it does, sigma factor recognizes both the -35 (highly) and -10 sequences.

Alpha helices within the sigma factor fit into ____.

the major groove and hydrogen bond to nucleotide bases.

Initiation of bacterial transcription continued…

The binding of RNAP to the promoter forms the ____.

closed complex

Open complex

Formed when the TATAAT box in the -10 sequence unwinds. The A-T rich regions in the -10 sequence are easier to separate. After its formation, a short RNA strand is made, and sigma factor is released. The core enzyme now slides down the DNA to synthesize an RNA strand. This signifies the transition to the elongation phase.

Elongation in bacterial transcription - RNA synthesis

The RNA transcript is synthesized. DNA is unwound as RNAP moves along the double helix. RNAP uses the template strand to make complementary RNA strand (U/T). The DNA double helix reforms behind the RNAP open complex.

RNA synthesis

RNAP synthesizes RNA in the 5’ to 3’ direction. RNAP does not have proofreading capability, therefore, it makes more mistakes than DNAP.

A promoter specifies the direction of transcription

With regard to adjacent genes along a chromosome, some promoters direct transcription in one direction and others direct transcription in the opposite direction.

Termination of bacterial transcription

Termination is the end of RNA synthesis. It occurs when the short RNA-DNA hybrid of the open complex is forced to separate. This releases the newly made RNA as well as RNA polymerase. There are two different mechanisms for termination.

two different mechanisms for termination

rho-dependent termination, rho-independent

Rho-dependent termination

A type of transcription termination that requires the rho protein.

Rho-independent termination

A type of transcription termination that is facilitated by two sequences: (1) a uracil-rich sequence located at the 3’ end of the RNA. (2) A stem-loop structure upstream of the uracil-rich sequence

transcription in eukaryotes

many of the basic features of gene transcription are very similar in bacteria and eukaryotes. However, gene transcription in eukaryotes is more complex.

Gene transcription in eukaryotes is more complex due to ____,____,____.

Larger, more complex cells (organelles); added cellular complexity means more genes that code proteins are required; multicellularity adds another level of regulation.

In eukaryotic transcription, nuclear DNA is transcribed by three different RNA polymerases:

RNAP l, RNAP ll, RNAP lll

RNAP l

transcribes all rRNA genes

RNAP ll

transcribes all mRNAs and some snRNA.

RNAP lll

transcribes tRNA, 5s rRNA, and microRNA

All three are very similar structurally and are composed of many subunits.

There is also a remarkable similarity between the bacterial RNAP and eukaryotic DNAP.

eukaryotic gens have a ____ and ____ that influence the core promoter.

core promoter and regulatory elements

core promoter

relatively short. It typically consists of the TATA box, a transcriptional start site, and one or more downstream promoter elements (DPEs)

In core promoters, what is not always present?

the TATA box. When it is present, it is important in determining the precise start point for transcription

the core promoter by itself produces a low level of ____.

a low level of transcription. Different from bacterial genes because its core promoter produces a high level of transcription.

the core promoter by itself produces ____.

basal transcription.

basal transcription

the core promoter by itself at low levels of transcription without additional regulatory elements

regulatory elements

short DNA sequences that control gene expression and are the primary regulatory components of the genome: enhancers, activators, and repressors

enhancers

Segments of DNA that are usually 50 bp to 1000 bp and contain one or more regulatory elements. They are recognized by regulatory transcription factors (RTFs). The role of them are to increase transcription rates.

regulatory transcription factors (RFTs)

Proteins that recognize and bind to enhancers.

activators

stimulate (increase) transcription

repressors

inhibit (decrease) transcription

eukaryotes vs bacteria enhancers

eukaryotes enhancers vary widely in their locations but are often found in the -50 to -100 region. bacteria enhancers are very close to the core promoter.

factors that regulate gene transcription can be divided into two general types:

cis-acting elements and trans-acting factors.

cis-acting elements

DNA sequences that exert their effect only over a particular gene. Examples: TATA box, enhancers containing regulatory elements.

trans-acting factors

general and regulatory transcription factor proteins that bind to cis-acting elements.

three categories of proteins are required for transcription to occur at the promoter:

RNAP ll, general transcription factors (GTFs), and mediator

general transcription factors (GTFs)

six different proteins

mediator

a protein complex