How Many Genes Are Essential?

KEY CONCEPTS
-Not all genes are essential. In yeast and flies, individual deletions of less than 50% of the genes have detectable effects.
-When two or more genes are redundant, a mutation in any one of them might not have detectable effects.
-We do not fully understand the persistence of genes that are apparently dispensable in the genome.
The force of natural selection ensures that functional genes are retained in the genome. Mutations occur at random, and a common mutational effect in an ORF will be to damage the protein product.
An organism with a damaging mutation will be at a disadvantage in competition and ultimately the mutation might be eliminated from a population. However, the frequency of a disadvantageous allele in the population is balanced between the generation of new copies of the allele by mutation and the elimination of the allele by selection.
Reversing this argument, whenever we see an intact, expressed ORF in the genome, researchers assume that its product plays a useful role in the organism. Natural selection must have prevented mutations from accumulating in the gene. The ultimate fate of a gene that ceases to be functional is to accumulate mutations until it is no longer recognizable.
The maintenance of a gene implies that it does not confer a selective disadvantage to the organism. However, in the course of evolution, even a small relative advantage can be the subject of natural selection, and a phenotypic defect might not necessarily be immediately detectable as the result of a mutation. Also, in diploid organisms, a new recessive mutation can be “hidden” in heterozygous form for many generations. However, researchers would like to know how many genes are actually essential, meaning that their absence is lethal to the organism. In the case of diploid organisms, it means, of course, that the homozygous null mutation is lethal.
We might assume that the proportion of essential genes will decline with an increase in genome size, given that larger genomes can have multiple related copies of particular gene functions. So far this expectation has not been borne out by the data.
One approach to the issue of gene number is to determine the number of essential genes by mutational analysis. If we saturate some specified region of the chromosome with mutations that are lethal, the mutations should map into a number of complementation groups that correspond to the number of lethal loci in that region. By extrapolating to the genome as a whole, we can estimate the total essential gene number.
In the organism with the smallest known genome (M. genitalium), random insertions have detectable effects in only about two-thirds of the genes. Similarly, fewer than half of the genes of E. coli appear to be essential. The proportion is even lower in the yeast S. cerevisiae. When insertions were introduced at random into the genome in one early analysis, only 12% were lethal and another 14% impeded growth. The majority (70%) of the insertions had no effect. A more systematic survey based on completely deleting each of 5,916 genes (more than 96% of the identified genes) shows that only 18.7% are essential for growth on a rich medium (i.e., when nutrients are fully provided). FIGURE 1. shows that these include genes in all categories. The only notable concentration of defects is in genes encoding products involved in protein synthesis, for which about 50% are essential. Of course, this approach underestimates the number of genes that are essential for the yeast to live in the wild when it is not so well provided with nutrients.

FIGURE 1. Essential yeast genes are found in all classes. Blue bars show the total proportion of each class of genes, and pink bars show those that are essential.
FIGURE 2 summarizes the results of a systematic analysis of the effects of loss of gene function in the nematode worm C. elegans. The sequences of individual genes were predicted from the genome sequence, and by targeting an inhibitory RNA against these sequences  a large collection of worms was made in which one predicted gene was prevented from functioning in each worm. Detectable effects on the phenotype were only observed for 10% of these knockdowns, suggesting that most genes do not play essential roles.

FIGURE 2. A systematic analysis of loss of function for 86% of worm genes shows that only 10% have detectable effects on the phenotype.
There is a greater proportion of essential genes (21%) among those worm genes that have counterparts in other eukaryotes, suggesting that highly conserved genes tend to have more basic functions. There is also an increased proportion of essential genes among those that are present in only one copy per haploid genome, compared with those for which there are multiple copies of related or identical genes. This suggests that many of the multiple genes might be relatively recent duplications that can substitute for one another’s functions.
Extensive analyses of essential gene number in a multicellular eukaryote have been made in Drosophila through attempts to correlate visible aspects of chromosome structure with the number of functional genetic units. The notion that this might be possible originated from the presence of bands in the polytene chromosomes of D. melanogaster. (These chromosomes are found at certain developmental stages and represent an unusually extended physical form in which a series of bands [more formally called chromomeres] are evident; see the Chromosomes chapter.) From the time of the early concept that the bands might represent a linear order of genes, there has been an attempt to correlate the organization of genes with the organization of bands. There are about 5,000 bands in the D. melanogaster haploid set; they vary in size over an order of magnitude, but on average there are about 20 kb of DNA per band.
The basic approach is to saturate a chromosomal region with mutations. Usually the mutations are simply collected as lethals without analyzing the cause of the lethality. Any mutation that is
lethal is taken to identify a locus that is essential for the organism. Sometimes mutations cause visible deleterious effects short of lethality, in which case we also define them as essential loci.
When the mutations are placed into complementation groups, the number can be compared with the number of bands in the region, or individual complementation groups might even be assigned to individual bands. The purpose of these experiments has been to determine whether there is a consistent relationship between bands and genes. For example, does every band contain a single gene?
Totaling the analyses that have been carried out since the 1970s, the number of essential complementation groups is about 70% of the number of bands. It is an open question as to whether there is any functional significance to this relationship. Regardless of the cause, the equivalence gives us a reasonable estimate for the essential gene number of around 3,600. By any measure, the number of essential loci in Drosophila is significantly less than the total number of genes.
If the proportion of essential human genes is similar to that of other eukaryotes, we would predict a range of 4,000 to 8,000 genes in which mutations would be lethal or produce evidently damaging effects. As of 2015, nearly 8,000 human genes in which mutations cause evident defects have been identified. This might actually exceed the upper range of the predicted total, especially in view of the fact that many lethal genes are likely to act so early in
development that we never see their effects. This sort of bias might also explain the results in TABLE 1, which show that the majority of known genetic defects are due to point mutations (where there is more likely to be at least some residual function of the gene).
TABLE 1 Most known genetic defects in human genes are due to point mutations. The majority directly affect the protein sequence.The remainder is due to insertions, deletions, or rearrangements of varying sizes.

How do we explain the persistence of genes whose deletion appears to have no effect? The most likely explanation is that the organism has alternative ways of fulfilling the same function. The simplest possibility is that there is redundancy, with some genes present in multiple copies. This is certainly true in some cases, in which multiple related genes must be knocked out in order to produce an effect. In a slightly more complex scenario, an organism might have two separate biochemical pathways capable of providing some activity. Inactivation of either pathway by itself would not be damaging, but the simultaneous occurrence of mutations in genes from both pathways would be deleterious.
Such situations can be tested by combining mutations. In this approach, deletions in two genes, neither of which is lethal by itself, are introduced into the same strain. If the double mutant dies, the strain is called a synthetic lethal. This technique has been used to great effect with yeast, for which the isolation of double mutants can be automated. The procedure is called synthetic genetic array analysis (SGA). FIGURE 3 summarizes the results of an analysis in which an SGA screen was made for each of 132 viable deletions by testing whether it could survive in combination with any one of 4,700 viable deletions. Every one of the tested genes had at least one partner with which the combination was lethal, and most of the tested genes had many such partners; the median is 25 partners and the greatest number is shown by one tested gene that had 146 lethal partners. A small proportion (about 10%) of the interacting mutant pairs encode polypeptides that interact physically.

FIGURE 3. All 132 mutant test genes have some combinations that are lethal when they are combined with each of 4,700 nonlethal mutations. This chart shows how many lethal interacting genes there are for each test gene.
This result goes some way toward explaining the apparent lack of effect of so many deletions. Natural selection will act against these deletions when they are found in lethal pair-wise combinations. To some degree, the organism is protected against the damaging effects of mutations by built-in redundancy. There is, however, a price in the form of accumulating the “genetic load” of mutations that are not deleterious in themselves but that might cause serious problems when combined with other such mutations in future generations. Presumably, the loss of the individual genes in such circumstances produces a sufficient disadvantage to maintain the functional gene during the course of evolution.