MASTERING BIOLOGY
A researcher wishes to increase the half-life of a particular mRNA that she is transfecting into a eukaryotic cell line. How should this researcher accomplish this goal?
Increase the length of the poly(A) tail of the mRNA prior to transfection.
RNA plays important roles in many cellular processes, particularly those associated with protein synthesis: transcription, RNA processing, and translation. transcription/RNA processing
snRNA pre-mRNA mRNA
Indicate processing events that you would also expect to find for the same species of RNARNA from a bacterial cell.
rRNA degradation of transcribed spacers generation of several molecules from one large precursor tRNA addition of CCA sequence if necessary methylation of bases Not found in bacterial cells addition of poly(A) tail on 3' end capping of 5' end removal of leader sequence at 5' end
What can you conclude about the number of introns present in gene Y?
At least one intron must be present in gene Y. It is also possible that more than one intron is present.
What can you conclude about the number of introns present in gene Z?
It is impossible to determine whether there are any introns in gene Z.
not used in protein synthesis
RNA primers
An exon within a coding region of a gene contains the following sequence: 5';-ATTGCACCTG-3'. Exposure of cells to a mutagen results in this sequence changing to 5'-ATTCGCACCTG-3'. Which of the following is true?
This gene has experienced a frameshift
Suppose that a portion of double-stranded DNA in the middle of a large gene is being transcribed by an RNA polymerase. As the polymerase moves through the sequence of six bases shown in the diagram below, what is the corresponding sequence of bases in the RNA that is produced?
UGAGCC There are three principles to keep in mind when predicting the sequence of the mRNA produced by transcription of a particular DNA sequence.The RNA polymerase reads the sequence of DNA bases from only one of the two strands of DNA: the template strand. The RNA polymerase reads the code from the template strand in the 3' to 5' direction and thus produces the mRNA strand in the 5' to 3' direction.In RNA, the base uracil (U) replaces the DNA base thymine (T). Thus the base-pairing rules in transcription are A→U, T→A, C→G, and G→C, where the first base is the coding base in the template strand of the DNA and the second base is the base that is added to the growing mRNA strand.
The 5'; cap and poly(A) tail of eukaryotic mRNAs __________.
have no template DNA sequence
Which of the following events signals the end of the initiation phase of bacterial transcription?
loss of the sigma factor from the RNA polymerase
After transcription begins, several steps must be completed before the fully processed mRNA is ready to be used as a template for protein synthesis on the ribosomes.
1. A poly-A tail (50-250 adenine nucleotides) is added to the 3' end of the pre-mRNA. 2. Noncoding sequences called introns are spliced out by molecular complexes called spliceosomes. 3. A cap consisting of a modified guanine nucleotide is added to the 5' end of the pre-mRNA. Once RNA polymerase II is bound to the promoter region of a gene, transcription of the template strand begins. As transcription proceeds, three key steps occur on the RNA transcript: Early in transcription, when the growing transcript is about 20 to 40 nucleotides long, a modified guanine nucleotide is added to the 5' end of the transcript, creating a 5' cap. Introns are spliced out of the RNA transcript by spliceosomes, and the exons are joined together, producing a continuous coding region. A poly-A tail (between 50 and 250 adenine nucleotides) is added to the 3' end of the RNA transcript. Only after all these steps have taken place is the mRNA complete and capable of exiting the nucleus. Once in the cytoplasm, the mRNA can participate in translation.
Life as we know it depends on the genetic code: a set of codons, each made up of three bases in a DNA sequence and corresponding mRNA sequence, that specifies which of the 20 amino acids will be added to the protein during translation. Imagine that a prokaryote-like organism has been discovered in the polar ice on Mars. Interestingly, these Martian organisms use the same DNA → RNA → protein system as life on Earth, except that there are only 2 bases (A and T) in the Martian DNA, and there are only 17 amino acids found in Martian proteins.
5 bases In the most general case of x bases and y bases per codon, the total number of possible codons is equal to xy . In the case of the hypothetical Martian life-forms, is the minimum codon length needed to specify 17 amino acids is 5 (25 = 32), with some redundancy (meaning that more than one codon could code for the same amino acid). For life on Earth, x = 4 and y = 3; thus the number of codons is 43, or 64. Because there are only 20 amino acids, there is a lot of redundancy in the code (there are several codons for each amino acid).
To investigate the possible presence of introns in three newly discovered genes (X, Y, and Z), you perform an experiment in which the restriction enzyme HaeIII is used to cleave either the DNADNA of each gene or the cDNA made by copying its mRNAmRNA with reverse transcriptase. The resulting DNA fragments are separated by gel electrophoresis, and the presence of fragments in the gels is detected by hybridizing to a radioactive DNA probe made by copying the intact gene with DNA polymerase in the presence of radioactive substrates. The following results are obtained: What can you conclude about the number of introns present in gene X?
At least one intron must be present in gene X. It is also possible that more than one intron is present.
During transcription, RNA polymerase synthesizes RNA from a DNA template with the help of accessory proteins. In this tutorial, you will review the steps of transcription in eukaryotes and bacteria and investigate splicing of mRNAs in eukaryotes. The diagram below shows a length of DNA containing a bacterial gene.
Bacterial transcription is a four-stage process. 1.Promoter recognition: RNA polymerase is a holoenzyme composed of a five-subunit core enzyme and a sigma (σσ) subunit. Different types of σσ subunits aid in the recognition of different forms of bacterial promoters. The bacterial promoter is located immediately upstream of the starting point of transcription (identified as the +1 nucleotide of the gene). The promoter includes two short sequences, the -10 and -35 consensus sequences, which are recognized by the σσ subunit. 2. Chain initiation: The RNA polymerase holoenzyme first binds loosely to the promoter sequence and then binds tightly to it to form the closed promoter complex. An open promoter complex is formed once approximately 18 bp of DNA around the -10 consensus sequence are unwound. The holoenzyme then initiates RNA synthesis at the +1 nucleotide of the template strand. 3. Chain elongation: The RNA-coding region is the portion of the gene that is transcribed into RNA. RNA polymerase synthesizes RNA in the 5′ → 3′ direction as it moves along the template strand of DNA. The nucleotide sequence of the RNA transcript is complementary to that of the template strand and the same as that of the coding (nontemplate) strand, except that the transcript contains U instead of T. 4. Chain termination: Most bacterial genes have a pair of inverted repeats and a polyadenine sequence located downstream of the RNA-coding region. Transcription of the inverted repeats produces an RNA transcript that folds into a stem-loop structure. Transcription of the polyadenine sequence produces a poly-U sequence in the RNA transcript, which facilitates release of the transcript from the DNA.
The following eukaryotic structural gene contains two introns and three exons. The table below shows four possible mRNA products of this gene. Use the labels to explain what may have caused each mRNA.
If a mutation alters a splicing signal sequence of an intron, that intron will not be removed accurately during the splicing reaction. This will result in the production of an abnormally spliced mature mRNA.Mutations in promoter sequences will affect transcription initiation and are likely to result in no mRNA being produced.
Flowchart to show the order of events as they are thought to occur during eukaryotic transcription involving RNA polymerase II (RNA pol II).
Transcription by RNA pol II in eukaryotes begins when TFIID recognizes and binds to the TATA box. The bound TFIID helps recruit TFIIA, TFIIB, TFIIF, and RNA pol II. Once those subunits of the minimal initiation complex are bound, TFIIE and TFIIH bind to form the complete initiation complex. Assembly of the complete initiation complex releases RNA pol II, which begins synthesizing the RNA transcript in the 5′ → 3′ direction. After the first 20-30 nucleotides have been synthesized, a cap consisting of a methylated guanine is added to the 5′ end of the pre-mRNA. Intron removal occurs as RNA pol II continues to elongate the pre-mRNA. When the polyadenylation signal has been transcribed, a poly-A tail is added to the 3′ end of the pre-mRNA. Polyadenylation is usually coupled with the termination of transcription.
If the nucleotides 5'-GAT-3' are paired with the nucleotides 3'-CUA-5', the pairing must be __________.
during transcription
During transcription in eukaryotes, a type of RNA polymerase called RNA polymerase II moves along the template strand of the DNA in the 3'→5' direction. However, for any given gene, either strand of the double-stranded DNA may function as the template strand.
the specific sequence of bases along the DNA strands In eukaryotes, binding of RNA polymerase II to DNA involves several other proteins known as transcription factors. Many of these transcription factors bind to the DNA in the promoter region (shown below in green), located at the 3' end of the sequence on the template strand. Although some transcription factors bind to both strands of the DNA, others bind specifically to only one of the strands. Transcription factors do not bind randomly to the DNA. Information about where each transcription factor binds originates in the base sequence to which each transcription factor binds. The positioning of the transcription factors in the promoter region determines how the RNA polymerase II binds to the DNA and in which direction transcription will occur.
In eukaryotes, pre-mRNA is produced by the direct transcription of the DNA sequence of a gene into a sequence of RNA nucleotides. Before this RNA transcript can be used as a template for protein synthesis, it is processed by modification of both the 5' and 3' ends. In addition, introns are removed from the pre-mRNA by a splicing process that is catalyzed by snRNAs (small nuclear RNAs) complexed with proteins. The product of RNA processing, mRNA (messenger RNA), exits the nucleus. Outside the nucleus, the mRNA serves as a template for protein synthesis on the ribosomes, which consist of catalytic rRNA (ribosomal RNA) molecules bound to ribosomal proteins. During translation, tRNA (transfer RNA) molecules match a sequence of three nucleotides in the mRNA to a specific amino acid, which is added to the growing polypeptide chain. RNA primers are not used in protein synthesis. RNA primers are only needed to initiate a new strand of DNA during DNA replication.
transcription/RNA processing snRNA pre-mRNA mRNA translation rRNA tRNA not used in protein synthesis RNA primers
RNA plays important roles in many cellular processes, particularly those associated with protein synthesis: transcription, RNA processing, and translation.
translation rRNA tRNA
