Histone variant HTZ1 shows extensive epistasis with, but does not increase robustness to, new mutations

Abstract

Biological systems produce phenotypes that appear to be robust to perturbation by mutations and environmental variation. Prior studies identified genes that, when impaired, reveal previously cryptic genetic variation. This result is typically interpreted as evidence that the disrupted gene normally increases robustness to mutations, as such robustness would allow cryptic variants to accumulate. However, revelation of cryptic genetic variation is not necessarily evidence that a mutationally robust state has been made less robust. Demonstrating a difference in robustness requires comparing the ability of each state (with the gene perturbed or intact) to suppress the effects of new mutations. Previous studies used strains in which the existing genetic variation had been filtered by selection. Here, we use mutation accumulation (MA) lines that have experienced minimal selection, to test the ability of histone H2A.Z (HTZ1) to increase robustness to mutations in the yeast Saccharomyces cerevisiae. HTZ1, a regulator of chromatin structure and gene expression, represents a class of genes implicated in mutational robustness. It had previously been shown to increase robustness of yeast cell morphology to fluctuations in the external or internal microenvironment. We measured morphological variation within and among 79 MA lines with and without HTZ1. Analysis of within-line variation confirms that HTZ1 increases microenvironmental robustness. Analysis of between-line variation shows the morphological effects of eliminating HTZ1 to be highly dependent on the line, which implies that HTZ1 interacts with mutations that have accumulated in the lines. However, lines without HTZ1 are, as a group, not more phenotypically diverse than lines with HTZ1 present. The presence of HTZ1, therefore, does not confer greater robustness to mutations than its absence. Our results provide experimental evidence that revelation of cryptic genetic variation cannot be assumed to be caused by loss of robustness, and therefore force reevaluation of prior claims based on that assumption.

Figure 1. Revelation of cryptic genetic variation without a change in robustness.

Top: In an abstract space of possible mutations (points), some (surrounded by solid ellipse) are neutral in the context of an allele X1 and some (surrounded by dashed ellipse) are neutral in the context of an allele X2. Alleles X1 and X2 confer equal robustness to mutations because an equal number of mutations are neutral in the context of each. Middle: Under selection, only neutral mutations accumulate. In the X1 genetic background, these are the mutations within the solid ellipse (left), whereas in the X2 genetic background these are the mutations within the dashed ellipse (right). Bottom: Perturbing the system by replacing one X allele with the other reveals cryptic genetic variation (open circles).

Figure 2. Mean principal component values of HTZ1+ and HTZ1− lines.

Each line connects an HTZ1+ MA line with its HTZ1− derivative. The means and standard deviations of line means are indicated by the black circles and bars. The mean of the ancestral strain is shown to the left in each plot.

Figure 3. Differences in principal component values reflect underlying differences in morphological phenotypes.

Individual cells from line pairs with similar (purple) and dissimilar (green) mean principal component values are shown for two different principal components. The line plots to the left are identical to those in Figure 2 or Figure S3, but with all lines grayed out except for those corresponding to the MA lines depicted on the right.

Figure 4. Mutational and environmental variances in HTZ1+ and HTZ1− lines.

(A) Mutational variances estimated for the HTZ1+ and HTZ1− lines for the indicated principal component. Orange = principal components for no-bud phenotypes; black = principal components for small-bud phenotypes; green = principal components for large-bud phenotypes. (B) Environmental variances estimated for the HTZ1+ and HTZ1− lines for the indicated principal component. Color scheme is the same as for A.