Figure 8–45 A behavioral phenotype detected in a genetic screen. (A) Wild-type C. elegans engage in social feeding. The worms migrate around until they encounter their neighbors and commence feeding on bacteria. (B) Mutant animals feed by themselves. (Courtesy of Cornelia Bargmann, Cell 94: cover, 1998. With permission from Elsevier.) mutant cells proliferate and form a colony at the permissive temperature mutant cells fail to proliferate and form a colony at the nonpermissive conditional mutations. The mutant individuals function normally as long as “permissive” conditions prevail, but demonstrate abnormal gene function when subjected to “nonpermissive” (restrictive) conditions. In organisms with temperature-sensitive mutations, for example, the abnormality can be switched on and off experimentally simply by changing the ambient temperature; thus, a cell containing a temperature-sensitive mutation in a gene essential for survival will die at a nonpermissive temperature but proliferate normally at the permissive temperature (Figure 8–46). The temperature-sensitive gene in such a mutant usually contains a point mutation that causes a subtle change in its protein product; for example, the mutant protein may function normally at low temperatures but unfold at higher temperatures.
Temperature-sensitive mutations were crucial to find the bacterial genes that encode the proteins required for DNA replication. The mutants were identified by screening populations of mutagen-treated bacteria for cells that stop making DNA when they are warmed from 30°C to 42°C. These mutants were later used to identify and characterize the corresponding DNA replication proteins (discussed in Chapter 5). Similarly, screens for temperature-sensitive mutations led to the identification of many proteins involved in regulating the cell cycle, as well as many proteins involved in moving proteins through the secretory pathway in yeast. Related screening approaches demonstrated the function of enzymes involved in the principal metabolic pathways of bacteria and yeast (discussed in Chapter 2) and identified many of the gene products responsible for the orderly development of the Drosophila embryo (discussed in Chapter 21).
Mutations Can Cause Loss or Gain of Protein Function
Gene mutations are generally classed as “loss of function” or “gain of function.” A loss-of-function mutation results in a gene product that either does not work or works too little; thus, it can reveal the normal function of the gene. A gain-of-function mutation results in a gene product that works too much, works at the wrong time or place, or works in a new way (Figure 8–47).
An important early step in the genetic analysis of any mutant cell or organism is to determine whether the mutation causes a loss or a gain of function. A standard test is to determine whether the mutation is dominant or recessive. A dominant mutation is one that still causes the mutant phenotype in the presence of a single copy of the wild-type gene. A recessive mutation is one that is no longer able to cause the mutant phenotype in the presence of a single wild-type copy of the gene. Although cases have been described in which a loss-of-function mutation is dominant or a gain-of-function mutation is recessive, in the vast majority of cases, recessive mutations are loss of function and dominant mutations are gain
Figure 8–46 Screening for temperature-sensitive bacterial or yeast mutants.
Mutagenized cells are plated out at the permissive temperature. They divide and form colonies, which are transferred to two identical Petri dishes by replica plating. One of these plates is incubated at the permissive temperature, the other at the nonpermissive temperature. Cells containing a temperature-sensitive mutation in a gene essential for proliferation can divide at the normal, permissive temperature but fail to divide at the elevated, nonpermissive temperature. Temperature-sensitive mutations of this type were especially useful for identifying genes needed for DNA replication, an essential process.
Figure 8–47 Gene mutations that affect their protein product in different ways.