Scientists of the Tokyo University of Sciences (Japan) have discovered exactly how circulating free DNA (cfDNA,) is generated, the DNA that is found in small amounts in the blood and that has escaped degradation by enzymes. In addition, they have evidenced possible applications of ‘DNase1L3’, the enzyme primarily responsible for generating cfDNA, as a novel molecule to prevent the spread of tumors.
Before new work, published in the magazine Biochemical and Biophysical Research Communications, these same scientists had already discovered an endonuclease, the 'DNase1L3', and found that it causes fragmentation of cellular DNA during necrosis: when a cell membrane breaks abruptly, the 'DNase1L3' in the bloodstream quickly degrades cellular DNA into individual nucleosomes .
They have also found that ‘DNase1L3’ plays a secondary role to activated DNase (CAD) during apoptosis: CAD degrades the initial DNA called chromatin and apoptotic cells are eliminated by macrophages. However, when some cells escape this process, they flow into the bloodstream and suffer secondary necrosis, after which DNase1L3 breaks down the DNA into nucleosomes.
Now, in this study, researchers using an experimental model genetically engineered to identify the enzymes responsible for generating cfDNA. They induced apoptosis and necrosis in normal individuals, in individuals deficient in CAD, deficient in ‘DNase1L3’ and in subjects with double deficiency in CAD plus ‘DNase1L3’.
Through electrophoresis, they observed that the blood of subjects with ‘DNase1L3’ deficiency had much lower concentrations of cfDNA than blood of CAD and normal deficiency, both in groups induced by apoptosis and by necrosis. Interestingly, the blood of the double-deficient CAD + ‘DNase1L3’ showed no trace of cfDNA at all. The scientists concluded that during apoptosis, ‘DNase1L3’ is crucial as a ‘reserve’ enzyme for the degradation of fragmented condensed chromatin (single nucleosomes), thus giving rise to cfDNA. And in necrosis, ‘DNaseIL3’ is absolutely essential to generate cfDNA.
The researchers also verified the activity of 'DNase1L3' and 'DNase1' (another enzyme that degrades DNA) in the blood and found that apoptosis and necrosis increased the activity of both 'DNase1L3' and 'DNase1' . However, even when cfDNA was not observed in subjects with double deficiency of CAD and ‘DNase1L3’, activity of ‘DNase1’ was observed. This showed that ‘DNase1’ is not essential for the generation of cfDNA.