RNA Molecules and RNA Processing Benjamin A. Pierce GENETICS A Conceptual Approach SIXTH EDITION CHAPTER 14 RNA Molecules and RNA Processing © 2017 W. H. Freeman and Company

The family of Tsar Nicholas Romanov II of Russia.

Gene Organization The concept of colinearity and noncolinearity Number of nucleotides in a gene should be proportional to the number of amino acids in the encoded protein DNA much longer than mRNA; demonstrated through hybridization

14.1 The concept of colinearity suggests that a continuous sequence of nucleotides in DNA encodes a continuous sequence of amino acids in a protein.

14.2 The noncolinearity of eukaryotic genes was discovered by hybridizing DNA and mRNA.

Gene Organization Introns Exons

14.3 The coding sequences of many eukaryotic genes are disrupted by noncoding introns.

TABLE 14.1 Major types of introns Type of Intron Location Splicing Mechanism Group I Genes of bacteria, bacteriophages, and eukaryotes Self-splicing Group II Genes of bacteria, archaea, and eukaryotic organelles Nuclear pre- mRNA Protein-encoding genes in the nucleus of eukaryotes Spliceosomal tRNA tRNA genes of bacteria, archaea, and eukaryotes Enzymatic Note: There are also several types of minor introns, including group III introns, twintrons, and archaeal introns.

The Concept of the Gene The gene includes DNA sequences that code for all exons and introns Those sequences at the beginning and end of the RNA that are not translated into a protein, including the entire transcription unit The promoter The RNA coding sequence The terminator

The Structure of Messenger RNA A mature mRNA contains a 5′ untranslated region (5′ UTR, or leader sequence) Shine–Dalgarno sequence Protein-coding region 3′ untranslated region

14.5 Three primary regions of mature mRNA are the 5′ untranslated region, the protein-coding region, and the 3′ untranslated region.

Pre-mRNA Processing The addition of the 5′ cap: A nucleotide with 7-methylguanine; 5′-5′ bond is attached to the 5′ end of the RNA. The addition of the poly(A) tail: ~ 50-250 adenine nucleotides are added to the 3′ end of the mRNA.

Posttranscriptional modifications to eukaryotic pre-mRNA TABLE 14.2 Posttranscriptional modifications to eukaryotic pre-mRNA Modification Function Addition of 5' cap Facilitates binding of ribosome to 5' end of mRNA, increases mRNA 3' cleavage and addition of poly(A) tail Increases stability of mRNA, facilitates binding of ribosome to mRNA RNA splicing Removes noncoding introns from pre-mRNA, facilitates export of mRNA to cytoplasm, allows for multiple proteins to be produced through alternative splicing RNA editing Alters nucleotide sequence of mRNA

14. 6 Most eukaryotic mRNAs have a 5′ cap 14.6 Most eukaryotic mRNAs have a 5′ cap. The cap consists of a nucleotide with 7-methylguanine attached to the pre-mRNA by a unique 5′–5′ bond (shown in detail in the bottom box).

14.7 Most eukaryotic mRNAs have a 3′ poly(A) tail.

Pre-mRNA Processing RNA splicing: Consensus sequences: 5′ consensus sequence: GU A/G AGU: 5′ splice site 3′ consensus sequence: CAGG Branch point: the adenine “A”: ~18-40 nucleotides upstream of 3′-splicing site Spliceosome: five RNA molecules + 300 proteins The process of splicing

14. 8 Splicing of pre-mRNA requires consensus sequences 14.8 Splicing of pre-mRNA requires consensus sequences. Critical consensus sequences are present at the 5′ splice site and the 3′ splice site. A weak consensus sequence (not shown) exists at the branch point.

14.9 The splicing of nuclear introns requires a two-step process.

14. 10 RNA splicing takes place within the spliceosome 14.10 RNA splicing takes place within the spliceosome. The spliceosome assembles sequentially.

Concept Check 1 It would be shorter than normal. If a splice site were mutated so that splicing did not take place, what would the effect be on the protein encoded by the mRNA? It would be shorter than normal. It would be longer than normal. It would be the same length but would have different amino acids.

Concept Check 1 It would be shorter than normal. If a splice site were mutated so that splicing did not take place, what would the effect be on the protein encoded by the mRNA? It would be shorter than normal. It would be longer than normal. It would be the same length but would have different amino acids.

Pre-mRNA Processing Nuclear organization Intron removal, mRNA processing, and transcription take place at the same site in the nucleus. Minor splicing Self-splicing introns in some rRNA genes in protists and in mitochondria genes in fungi Alternative processing pathways for processing pre-mRNA

14.11 Group I and group II introns fold into characteristic secondary structures.

14.12 Eukaryotic cells have alternative pathways for processing pre-mRNA. (a) With alternative splicing, pre-mRNA can be spliced in different ways to produce different mRNAs. (b) With multiple 3′ cleavage sites, there are two or more potential sites for cleavage and polyadenylation; use of the different sites produces mRNAs of different lengths.

14.13 Pre-mRNA encoded by the gene for calcitonin undergoes alternative processing.

Concept Check 2 Alternative 3′ cleavage sites result in _____. multiple genes of different lengths multiple genes of pre-mRNA of different lengths multiple mRNAs of different lengths All of the above

Concept Check 2 Alternative 3′ cleavage sites result in _____. multiple genes of different lengths multiple genes of pre-mRNA of different lengths multiple mRNAs of different lengths All of the above

RNA Editing RNA editing: coding sequence altered after transcription. Guide RNAs play a role.

14. 14 Trypanosoma brucei causes African sleeping sickness 14.14 Trypanosoma brucei causes African sleeping sickness. Messenger RNA produced from mitochondrial genes of this parasite (in purple) undergoes extensive RNA editing. [David Spears/Last Refuge Ltd./Phototake.]

14. 15 RNA editing is carried out by guide RNAs 14.15 RNA editing is carried out by guide RNAs. The guide mRNA has sequences that are partly complementary to those of the preedited mRNA and pairs with it. After pairing, the mRNA undergoes cleavage and new nucleotides are added, with sequences in the gRNA serving as a template. The ends of the mRNA are then joined together.

14.16 Mature eukaryotic mRNA is produced when pre-mRNA is transcribed and undergoes several types of processing.

14.17 This representation of the nucleotide sequence of the gene for human interleukin 2 includes the TATA box, transcription start site, start and stop codons, introns, exons, poly(A) consensus sequence, and 3′ cleavage site.

The Structure of Transfer RNA Rare modified RNA nucleotide bases Ribothymine Pseudouridine Common secondary structure—the cloverleaf structure Anticodon

14. 18 Two of the modified bases found in tRNAs 14.18 Two of the modified bases found in tRNAs. All the modified bases in tRNAs are produced by the chemical alteration of the four standard RNA bases.

14.19 All tRNAs possess a common secondary structure, the cloverleaf structure. The base sequence in the flattened model is for tRNAAla.

14.20 Transfer RNAs are processed in both bacterial and eukaryotic cells. Different tRNAs are modified in different ways. One example is shown here.

Concept Check 3 Encoded by guide RNAs How are rare bases incorporated into tRNAs? Encoded by guide RNAs By chemical changes in one of the standard bases Encoded by rare bases in DNA Encoded by sequences in introns

Concept Check 3 Encoded by guide RNAs How are rare bases incorporated into tRNAs? Encoded by guide RNAs By chemical changes in one of the standard bases Encoded by rare bases in DNA Encoded by sequences in introns

The Structure of the Ribosome Large ribosome subunit Small ribosome subunit

Composition of ribosomes in bacterial and eukaryotic cells TABLE 14.3 Composition of ribosomes in bacterial and eukaryotic cells Cell Type Ribosome Size Subunits rRNA Component Proteins Bacterial 70S Large (50S) 23S (2900 nucleotides), 5S (120 nucleotides) 31 Small (30S) 16S (1500 nucleotides) 21 Eukaryotic 80S Large (60S) 28S (4700 nucleotides), 5.8S (160 nucleotides), 5S (120 nucleotides) 49 Small (40S) 18S (1900 nucleotides) 33 Note: The letter “S” stands for “Svedberg unit.”

Numbers of rRNA genes in different organisms TABLE 14.4 Numbers of rRNA genes in different organisms Species Number of Copies of rRNA Genes per Genome Escherichia coli 7 Yeast 100–200 Human 280 Frog 450

Ribosomal RNA is processed after transcription Ribosomal RNA is processed after transcription. Prokaryotic rRNA (a) and eukaryotic rRNA (b) are produced from precursor RNA transcripts that are methylated, cleaved, and processed to produce mature rRNAs. Eukaryotic 5S rRNA is transcribed separately from a different gene.

Concept Check 4 Methylation of bases Cleavage of bases What types of changes take place in rRNA processing? Methylation of bases Cleavage of bases Nucleotides trimmed from the ends of rRNAs All of the above

Concept Check 4 Methylation of bases Cleavage of bases What types of changes take place in rRNA processing? Methylation of bases Cleavage of bases Nucleotides trimmed from the ends of rRNAs All of the above

14.5 Small RNA Molecules Participate in a Variety of Functions RNA interference: limits the invasion of foreign genes and censors the expression of their own genes Types of small RNAs Processing and function of microRNAs

14.22 Small interfering RNAs and microRNAs are produced from double-stranded RNAs.

Differences between siRNAs and miRNAs TABLE 14.5 Differences between siRNAs and miRNAs Feature siRNA miRNA Origin mRNA, transposon, or virus RNA transcribed from distinct gene Cleavage of RNA duplex or single-stranded RNA that forms long hairpins Single-stranded RNA that forms short hairpins of double-stranded RNA Size 21–25 nucleotides Action Degradation of mRNA, inhibition of transcription, chromatin modification Degradation of mRNA, inhibition of translation, chromatin modification Target Genes from which they were transcribed Genes other than those from which they were transcribed

14.23 CRISPR RNAs function in defense against invasion of foreign DNAs, such as DNA from bacteriophage and plasmids.