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Cellular Life

Haploid yeast cells, with only one copy of each vital gene, are always threatened by mutations. Any serious damage in a gene coding for an enzyme that is in an important metabolic pathway, such as the pathway that makes the amino acid tryptophan, can kill the whole cell.

Cells damaged this way cannot complete the synthesis and production of the amino acid, and thus are said to be try-.

Probably long before there were any yeast cells, early proto-eukaryotic cells faced the same problem, and were able to solve this potentially lethal situation by joining forces. Instead of relying on a single set of chromosomes, DNA and genes, cells discovered that if they fused with one another, pooling their biological information, their chances of survival went up enormously.

Cells with two copies of their chromosomes, DNA and genes are termed diploid.

haploids to diploids

Haploid yeast cells can be mutated and damaged in different genes in the same metabolic pathway, with the same result;
both are try-.

Here are two possible examples:

haploid mutant #1

haploid mutant #2

Neither of these mutants can survive on their own unless the experimenter adds the missing nutrient, tryptophan, to their growth medium. But, if these two mutants fuse together, and pool their biological information (genes), then the cytoplasm of such a fused cell would contain all the enzymes necessary to make tryptophan. The diploid cell would not be dependent on an external source of this amino acid.

Diploid Cell

In this example, two haploid cells, mutated in two different genes in the same metabolic pathway, have been fused together to make a new type of cell, a diploid, that showed no deficiencies in its phenotype.

The genotype of the diploid cell still contained the mutant genes inherited from its haploid precursors, but it also inherited the normal, working copies of these genes as well. So the deficiencies were compensated for by the presence of the normal genes.

In genetics, this phenomenon is known as complementation.

If the experimenter fused together two different haploid yeast mutants, and the diploid still needed to be fed extra tryptophan, then these cells were mutated in the same gene!

But it the experimenter fused together two different haploid yeast mutants, and the resulting diploid was now free of its dependence on external tryptophan, then the original haploid cells were mutated in different genes!

Complementation analysis

In the table below the scientist has created five different haploid yeast mutants, none of which can grow unless extra tryptophan is added to their growth medium. They are all try-.

Each mutant is fused in a pair-wise fashion with each other mutant, and the resulting diploids placed in medium that does not contain tryptophan. Only diploids that now have at least one gene for each enzyme in the pathway (i.e. that "complement" each other) will now grow.

From this table it is possible to sort out which mutants are mutated in the same gene and which mutants are mutated in different genes.

haploid mutants
#1 #2 #3 #4 #5
#1 - +++ - +++ -
#2 +++ - - +++ -
#3 - - - - +++
#4 +++ +++ - - -
#5 - - +++ - -

[The number of each haploid mutant yeast cell is written along the top of this table and down the left side. Each mutant cell is fused with each other mutant cell to produce 25 different diploid cells. These diploids are represented by the intersections of the rows and columns (inside the larger table). A "+++" means that this diploid will grow well on medium that does NOT contain any extra amino acid, so the diploid has all the genes necessary to make this ingredient.]

From this table it can be seen that Mutant #1 when fused with Mutant #4 produces a diploid that is no longer dependent on externally added tryptophan, so the mutations must be in different genes.

However, when Mutant #1 is fused with Mutant #3 the diploid that results still cannot grow on its own, so these two mutations must be in the same gene!

See if you can work out how many different (and same) mutants there are in these five examples.

© 2002, Professor John Blamire