Editing POLG, the gene that proofreads mitochondrial DNA copies, produced a pig model of accelerated aging
Editing POLG, the gene that proofreads mitochondrial DNA copies, produced a pig model of accelerated aging
On 28 August, a Science Advances paper described 12 pigs in which both copies of POLG carried the D260A substitution. Mitochondrial DNA mutations accumulated in these animals, cells taken from their ears consumed less oxygen, and none of the animals lived longer than seven months.
Mitochondria supply cells with energy and contain their own DNA, known as mtDNA. The POLG protein copies mtDNA, while its exonuclease domain acts as a built-in proofreader that checks the new strand for errors. The authors changed one amino acid in this domain by replacing aspartic acid at position 260 with alanine. This variant is designated D260A. Animals with two edited copies of POLG had a higher mtDNA mutation burden.
The choice of D260A follows directly from a specific experiment conducted in 2004. In a mouse study, the D257A substitution in the corresponding region of Polg increased the number of mtDNA point mutations three to five times and was associated with early age-related features. The same region is located at position 260 in pigs, so the new study tests the same causal hypothesis in a large animal.
The researchers first introduced the required substitution into fibroblasts, which are connective tissue cells, from a pig fetus using prime editing, a method for making precise DNA substitutions. They then transferred nuclei from selected cells into enucleated oocytes and produced 15 males. Both copies of POLG were edited in 12 animals, while one copy was edited in three. Animals with one edited copy grew normally and reached sexual maturity. Whole nuclear genome sequencing of two-week-old piglets showed that the numbers of point mutations and short DNA insertions or deletions in the cloned animals were comparable to those in control pigs.
Two methods for detailed mtDNA sequencing found an excess of mutations throughout mtDNA in animals with two edited copies. Connective tissue cells from their ears consumed less oxygen under both basal and maximal conditions, a measure of mitochondrial energy-producing capacity. These animals lost weight, developed anemia, had thinner skin and subcutaneous fat, and showed more dying cells in the intestines and testes. Six of the 12 animals with two edited copies died at approximately two months of age. The others also died before seven months.
This single pig line connects defective mtDNA proofreading with error accumulation, impaired cellular energy metabolism, and changes across several tissues. It can be used to test how interventions affect individual steps in this sequence, from mutation accumulation to the response of a specific organ.