In the study, researchers found that these "silent" differences in the nucleotide sequence seem to influence the density of ribosomes, the molecular machines that translate RNA into proteins. Such differences may enable each individual actin form to take on a different role in the cell. This study has been published in the journal eLife.
New findings by Kashina and colleagues have pointed to a surprising answer. The differing functions of these proteins are determined not by their amino acid sequences but by their genetic code. "We like to call it the 'silent code,'" Kashina said. "Our findings show that the parts of genes that we think of as being silent actually encode very key functional information."
When β-actin is lacking, for example, mice die at an early stage of embryonic development. But mice lacking γ-actin, though typically smaller than normal and deaf, can survive to adulthood. The researchers took advantage of the precision gene editing made possible by the CRISPR/Cas-9 system.
While the two actin isoforms differ by only four amino acids, their mRNA coding sequences differ by almost 13 percent because of "silent" nucleotide differences that nevertheless encode the same amino acids. Making changes to only five nucleotides in the β-actin gene, they could transform it so that its amino acid output would be the same as the γ-actin protein. All that would distinguish it would be the silent nucleotide substitutions.
The gene editing worked. Mice with these edits had no β-actin protein. But unlike true β-actin knockouts they were completely healthy and viable, just as if they possessed the proper proporitions of β-actin and γ-actin proteins. They survived to reproduce and averaged the same litter sizes as normal animals.
The researchers performed the same experiment, editing the γ-actin gene to encode the β-actin protein but were only able to change the coding sequence for three of the four amino acids. Still, mice subject to this partial replacement also appeared normal and healthy, despite lacking γ-actin protein.
Kashina's team found that the γ-actin proteins made from the edited β-actin gene formed a normal cytoskeleton and enabled cell to migrate in a normal fashion. Getting at a mechanism, the DNA sequence could influence protein function, the researchers found that ribosomes density on β-actin RNA is more than a thousand times higher than on γ-actin RNA, and indeed all six actin genes had differences in ribosome density.
And curious as to how widespread this phenomenon might be, the researchers looked for protein families with nearly identical members that are encoded by different genes and had significant variations in ribosome density across the family. They found many groups that were shared across mice, zebrafish and human genomes.