Chapter 3 The Interrupted Gene

3.1 Introduction In eukaryotes, a gene may include additional sequences that lie within the coding region and interrupt the sequence that codes for the protein. Figure 3.01: Introns are removed to make mRNA from an interrupted gene.

3.2 An Interrupted Gene Consists of Exons and Introns Introns are removed by the process of RNA splicing. Only mutations in exons can affect polypeptide sequence. Mutations in introns can affect processing of the RNA and prevent production of polypeptide.

3.2 An Interrupted Gene Consists of Exons and Introns Figure 3.02: The order of exons does not change between DNA and RNA.

3.3 Organization of Interrupted Genes May Be Conserved Introns can be detected by the presence of additional regions. Genes are compared with their RNA products by restriction mapping or electron microscopy. The ultimate determination is based on comparison of sequences.

Figure 3.03: RNA hybridizes with the template strand of the gene.

3.3 Organization of Interrupted Genes May Be Conserved Figure 3.04: An intron does not hybridize with mRNA.

Figure 3.05A: Multiple introns form loops in hybridization. Photo reproduced from Berget, S. M., Moore, C., and Sharp, P.A., Proc. Natl. Acad. Sci. USA 74 (1977): 3171-3175. Used with permission of Philip Sharp, Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology.

Figure 3.05B: Multiple introns form loops in hybridization.

Figure 3.06: Restriction sites in introns are missing from the cDNA.

3.3 Organization of Interrupted Genes May Be Conserved The positions of introns are usually conserved when homologous genes are compared between different organisms. The lengths of the corresponding introns may vary greatly. Introns usually do not code for polypeptides.

3.3 Organization of Interrupted Genes May Be Conserved Figure 3.07: Continuous open reading frames are created when introns are removed from RNA.

Figure 3.08: Globin genes vary in intron lengths but have the same structure.

3.4 Exon Sequences Are Conserved but Introns Vary Comparisons of related genes in different species show that the sequences of the corresponding exons are usually conserved The sequences of the introns are much less well related. Introns evolve much more rapidly than exons. This is due to the lack of selective pressure to produce a protein with a useful sequence.

3.4 Exon Sequences Are Conserved but Introns Vary Figure 3.09: Related genes diverge in the introns.

Figure 3.10: Interrupted genes predominate in higher eukaryotes. 3.5 Genes Show a Wide Distribution of Sizes Primarily Due to Intron Size and Number Variation Most genes are uninterrupted in yeasts, but are interrupted in higher eukaryotes. Figure 3.10: Interrupted genes predominate in higher eukaryotes.

Figure 3.11: Genes have a wide range of sizes.

3.5 Genes Show a Wide Distribution of Sizes Primarily Due to Intron Size and Number Variation Introns are short in lower eukaryotes, but range up to several 10s of kb in length in higher eukaryotes. The overall length of a gene is determined largely by its introns.

Figure 3.12: Exons are typically 100-200 bp. 3.5 Genes Show a Wide Distribution of Sizes Primarily Due to Intron Size and Number Variation Exons are usually short, typically coding for ~100 amino acids. Figure 3.12: Exons are typically 100-200 bp.

Figure 3.13: Introns have wide length variation. 3.5 Genes Show a Wide Distribution of Sizes Primarily Due to Intron Size and Number Variation Figure 3.13: Introns have wide length variation.

3.6 Some DNA Sequences Code for More Than One Polypeptide The use of alternative initiation or termination codons allows two proteins to be generated where one is equivalent to a fragment of the other. Figure 3.14: Alternative starts (or stops) generate related proteins.

3.6 Some DNA Sequences Code for More Than One Polypeptide Nonhomologous protein sequences can be produced from the same sequence of DNA when it is read in different reading frames by two (overlapping) genes. Figure 3.15: Overlapping triplets may be used in different reading frames.

3.6 Some DNA Sequences Code for More Than One Polypeptide Homologous proteins that differ by the presence or absence of certain regions can be generated by differential (alternative) splicing when certain exons are included or excluded. This may take the form of including or excluding individual exons or of choosing between alternative exons.

Figure 3.16: Alternative splicing can subsititute exons.

3.6 Some DNA Sequences Code for More Than One Polypeptide Figure 3.17: Different combinations of exons are used in alternative splicing.

3.7 How Did Interrupted Genes Evolve? A major evolutionary question is whether genes originated as sequences interrupted by introns or whether they were originally uninterrupted. More evidence supports the first (“introns early”) hypothesis, though it appears that introns can be inserted into genes. Most protein-coding genes probably originated in an interrupted form, but interrupted genes that code for RNA may have originally been uninterrupted. A special class of introns are mobile and can insert themselves into genes.

Figure 3.18: Random translocations may produce functional genes.

3.8 Some Exons Can Be Equated with Protein Functions Many exons can be equated with coding for polypeptide sequences that have particular functions. Related exons are found in different genes.

3.8 Some Exons Can Be Equated with Protein Functions Figure 3.19: Immunoglobulin exons code for protein domains

Figure 3.20: Exons in two proteins can be related.

3.9 The Members of a Gene Family Have a Common Organization A common feature in a set of genes is assumed to identify a property that preceded their separation in evolution. All globin genes have a common form of organization with three exons and two introns. This suggests that they are descended from a single ancestral gene.

3.9 The Members of a Gene Family Have a Common Organization Figure 3.21: Leghemoglobin has an extra intron.

Figure 3.22: One rat insulin gene has lost an intron.

Figure 3.23: Many changes in introns have occured in actin gene evolution.