Evolutionary conservation and modulation of a juvenile growth-regulating genetic program

Abstract

Body size varies enormously among mammalian species. In small mammals, body growth is typically suppressed rapidly, within weeks, whereas in large mammals, growth is suppressed slowly, over years, allowing for a greater adult size. We recently reported evidence that body growth suppression in rodents is caused in part by a juvenile genetic program that occurs in multiple tissues simultaneously and involves the downregulation of a large set of growth-promoting genes. We hypothesized that this genetic program is conserved in large mammals but that its time course is evolutionarily modulated such that it plays out more slowly, allowing for more prolonged growth. Consistent with this hypothesis, using expression microarray analysis, we identified a set of genes that are downregulated with age in both juvenile sheep kidney and lung. This overlapping gene set was enriched for genes involved in cell proliferation and growth and showed striking similarity to a set of genes downregulated with age in multiple organs of the juvenile mouse and rat, indicating that the multiorgan juvenile genetic program previously described in rodents has been conserved in the 80 million years since sheep and rodents diverged in evolution. Using microarray and real-time PCR, we found that the pace of this program was most rapid in mice, more gradual in rats, and most gradual in sheep. These findings support the hypothesis that a growth-regulating genetic program is conserved among mammalian species but that its pace is modulated to allow more prolonged growth and therefore greater adult body size in larger mammals.

Keywords: evolutionary conservation; gene expression; growth limitation; phylogeny.

Fig. 1

A multiorgan growth-regulating genetic program common to mice, rats, and sheep. A) Venn diagrams showing the number of genes significantly downregulated and upregulated with age by microarray in sheep vs. mouse and rat. Genes that showed significant age regulation (FDR<0.1) in both kidney and lung in sheep were compared with the 174 genes downregulated or the 58 genes upregulated in all organs studied in mouse and rat. * P<0.001. B) Gene ontology analysis of the genes that were age-regulated in both organs in sheep (FDR<0.1, fold change>1.5). Downregulated (401) and upregulated (334) genes were analyzed separately using Ingenuity Pathway Analysis. The top 6 most strongly overrepresented molecular, cellular, or physiological functions are shown. Grey bars, -log (P value); black, number of molecules. C) Heat map based on microarray analysis of the 232 genes that were commonly down- or upregulated with age in the mouse (columns 1–4) and rat (columns 5–6) and present in the sheep microarray (columns 7–10). Each row corresponds to a gene, ranked by average fold-change in mouse and rat. D) Heat map similar to (C) but excluding 78 genes that have a known function specific to the cell-cycle. Green, downregulation with age; red, upregulation. Scale values are log₂ (fold difference). P, postnatal age (weeks); F, fetal age (days); AD, adult (21 months).

Fig. 2

Rate of decline in growth velocity varies among mouse, rat, and sheep. Percent change in body weight per change in time was calculated for mouse (dotted line) based on weights at 4, 7, 9, 12, 15, and 19 wk post-fertilization; rat (dashed line) at 6, 7, 9, 12, and 15 wk post-fertilization; and sheep (solid line) at 9, 13, 24, 31, 42, and 111 wk post-fertilization. Age is expressed as wk post-fertilization to account for interspecies differences in gestational age.

Fig. 3

The pace of the multi-organ juvenile genetic program differs among species of different body size. A) Mice versus rats. For the 235 genes that were downregulated with age in all organs studied in mouse and rat, we compared the fold change in expression from 1 to 4 wks of age in the mouse versus 1 to 5 wks in the rat by microarray. Negative values indicate decreasing expression. Boundaries of the box, 25^th and 75^th percentile; bars, 10^th and 90^th percentiles; central line segment, median. B) Mice versus sheep. For genes that were commonly downregulated in all organs studied of all 3 species, we compared the expression patterns in mice and sheep by microarray. Age is expressed as weeks post-fertilization to account for differences in length of gestation. Relative expression was normalized to the young adult time point in each species (mouse, 11wk; sheep, 111wk). Dotted line separates data from different microarray experiments.

Fig. 4

Downregulation of specific growth-regulating genes occurs more slowly in sheep (open circles) than in mice (closed circles). mRNA relative expression (mean ± SEM) was assessed by quantitative RT-PCR. Top to bottom row: kidney, lung, heart, liver. Relative expression was normalized to the young adult time point in each species (mouse, 11wk; sheep, 111wk).