Human X ChromosomeMouse X Chromosome
6.11 Gene Prediction
nucleotides results in minor changes in the protein. For example, the phrase THE SLY FOX AND THE SHY DOG(written in triplets) turns into gibber- ish after deleting one letter (THE SYF OXA NDT HES HYD OG) or two let- ters (THE SFO XAN DTH ESH YDO G), but makes some sense after delet- ing three nucleotides THE SOX AND THE SHY DOG. Inspired by this ex- periment Charles Yanofsky proved that a gene and its protein product are collinear, that is, the first codon in the gene codes for the first amino acid in the protein, the second codon codes for the second amino acid (rather than, say, the seventeenth), and so on. Yanofsky’s ingenious experiment was so influential that nobody even questioned whether codons are represented by continuous stretches in DNA, and for the subsequent fifteen years biologists believed that a protein was encoded by a long string of contiguous triplets.
However, the discovery of split human genes in 1977 proved that genes are often represented by acollectionof substrings, and raised the computational problem of predicting the locations of genes in a genome given only the ge- nomic DNA sequence.
The human genome is larger and more complex than bacterial genomes.
This is not particularly surprising since one would expect to find more genes in humans than in bacteria. However, the genome size of many eukaryotes does not appear to be related to an organism’s genetic complexity; for exam- ple, the salamander genome is ten times larger than the human genome. This apparent paradox was resolved by the discovery that many organisms con- tain not only genes but also large amounts of so-calledjunk DNAthat does not code for proteins at all. In particular, most human genes are broken into pieces calledexonsthat are separated by this junk DNA. The difference in the sizes of the salamander and human genomes thus presumably reflects larger amounts of junk DNA and repeats in the salamander genome.
Split genes are analogous to a magazine article that begins on page 1, con- tinues on page 13, then takes up again on pages 43, 51, 74, 80, and 91, with pages of advertising appearing in between. We do not understand why these jumps occur. and a significant portion of the human genome is this junk “ad- vertising” that separates exons.
More confusing is that the jumps between different parts of split genes are inconsistent from species to species. A gene in an insect edition of the genome will be organized differently than the related gene in a worm genome.
The number of parts (exons) may be different: the information that appears in one part in the human edition may be broken up into two in the mouse version, or vice versa. While the genes themselves are related, they may be quite different in terms of the parts’ structure.
6.11 Gene Prediction 195 Split genes were first discovered in 1977 in the laboratories of Phillip Sharp and Richard Roberts during studies of the adenovirus. The discovery was such a surprise that the paper by Roberts’s group had an unusually catchy ti- tle for the journalCell: “An Amazing Sequence Arrangement at the 5’ End of Adenovirus 2 Messenger RNA.” Sharp’s group focused their experiments on an mRNA17 that encodes a viral protein known ashexon. To map the hexon mRNA in the viral genome, mRNA was hybridized to adenovirus DNA and the hybrid molecules were analyzed by electron microscopy. Strikingly, the mRNA-DNA hybrids formed in this experiment displayed three loop struc- tures, rather than the continuous duplex segment suggested by the classic continuous gene model (figure 6.23). Further hybridization experiments re- vealed that the hexon mRNA is built from four separate fragments of the adenovirus genome. These four continuous segments (calledexons) in the adenovirus genome are separated by three “junk” fragments calledintrons.
Gene prediction is the problem of locating genes in a genomic sequence.
Human genes constitute only3%of the human genome, and no existing in silico gene recognition algorithm provides completely reliable gene recogni- tion. The intron-exon model of a gene seems to prevail in eukaryotic organ- isms; prokaryotic organisms (like bacteria) do not have broken genes. As a result, gene prediction algorithms for prokaryotes tend to be somewhat simpler than those for eukaryotes.18
There are roughly two categories of approaches that researchers have used for predicting gene location. The statistical approach to gene prediction is to look for features that appear frequently in genes and infrequently elsewhere.
Many researchers have attempted to recognize the locations ofsplicing signals at exon-intron junctions.19 For example, the dinucleotidesAGandGTon the left- and right-hand sides of an exon are highly conserved (figure 6.24). In addition, there are other less conserved positions on both sides of the exons.
The simplest way to represent such binding sites is by a profile describing the propensities of different nucleotides to occur at different positions. Unfortu-
17. At that time, messenger RNA (mRNA) was viewed as a copy of a gene translated into the RNA alphabet. It is used to transfer information from the nuclear genome to the ribosomes to direct protein synthesis.
18. This is not to say that bacterial gene prediction is a trivial task but rather to indicate that eukaryotic gene finding is very difficult.
19. If genes are separated into exons interspersed with introns, then the RNA that is transcribed from DNA (i.e., the complementary copy of a gene) should be longer than the mRNA that is used as a template for protein synthesis. Therefore, some biological process needs to remove the introns in the pre-mRNA and concatenate the exons into a single mRNA string. This process is known assplicing, and the resulting mRNA is used as a template for protein synthesis in cytoplasm.
Figure 6.23 An electron microscopy experiment led to the discovery of split genes.
When mRNA (below) is hybridized against the DNA that generated it, three dis- tinct loops can be seen (above). Because the loops are present in the DNA and are not present in mRNA, certain parts (introns) must be removed during the process of mRNA formation called splicing.
Exon 1
Intron 1
Exon 2
Intron 2
Exon 3
GT AG GT AG
Figure 6.24 Exons typically are flanked by the dinucleotidesAGandGT.
nately, using profiles to detect splice sites has met with limited success since these profiles are quite weak and tend to match frequently in the genome at nonsplice sites. Attempts to improve the accuracy of gene prediction led to the second category of approaches for gene finding: those based on similar- ity.
The similarity-based approach to gene prediction relies on the observation that a newly sequenced gene has a good chance of being related to one that