DNA fragmentation

DNA fragmentation is the separation or breaking of DNA strands into pieces. It can be done intentionally by laboratory personnel or by cells, or can occur spontaneously. Spontaneous or accidental DNA fragmentation is fragmentation that gradually accumulates in a cell. It can be measured by e.g. the Comet assay.

The degree of DNA fragmentation can predict outcomes for in vitro fertilization[1] (IVF) and its expansion intracytoplasmic sperm injection[2] (ICSI). The sperm chromatin dispersion test (SCD) and TUNEL assay are both effective in detecting sperm DNA damage.[3] Using bright-field microscopy, the SCD test appears to be more sensitive than the TUNEL assay.[3]

Its main units of measurement is the DNA Fragmentation Index (DFI).[2] A DFI of 20% or more significantly reduces the success rates after ICSI.[2]

DNA fragmentation was first documented by Williamson in 1970 when he observed discrete oligomeric fragments occurring during cell death in primary neonatal liver cultures. He described the cytoplasmic DNA isolated from mouse liver cells after culture as characterized by DNA fragments with a molecular weight consisting of multiples of 135 kDa. This finding was consistent with the hypothesis that these DNA fragments were a specific degradation product of nuclear DNA.[4]

Intentional

DNA fragmentation is often necessary prior to library construction or subcloning for DNA sequences. A variety of methods involving the mechanical breakage of DNA have been employed where DNA is fragmented by laboratory personnel. Such methods include sonication, needle shear, nebulisation, point-sink shearing and passage through a pressure cell.[5]

Spontaneous

Apoptotic DNA fragmentation is a natural fragmentation that cells perform in apoptosis (programmed cell death). DNA fragmentation is a biochemical hallmark of apoptosis. In dying cells, DNA is cleaved by an endonuclease that fragments the chromatin into nucleosomal units, which are multiples of about 180-bp oligomers and appear as a DNA ladder when run on an agarose gel. The enzyme responsible for apoptotic DNA fragmentation is the Caspase-activated DNase. CAD is normally inhibited by another protein, the Inhibitor of Caspase Activated DNase (ICAD). During apoptosis, the apoptotic effector caspase, caspase 3, cleaves ICAD and thus causes CAD to become activated.[9]

A DNA double strand wrapped around a core of histone proteins
A nucleosome, consisting of DNA (grey) wrapped around a histone tetramer (coloured). In apoptotic DNA fragmentation, the DNA is cleaved in the internucleosomal linker region, which is the part of the DNA not wrapped around the histones.

CAD cleaves the DNA at the internucleosomal linker sites between the nucleosomes, protein-containing structures that occur in chromatin at ~180-bp intervals. This is because the DNA is normally tightly wrapped around histones, the core proteins of the nucleosomes. The linker sites are the only parts of the DNA strand that are exposed and thus accessible to CAD.

Uses

DNA Fragmentation plays an important part in forensics, especially that of DNA profiling.

References

  1. Simon L, Brunborg G, Stevenson M, Lutton D, McManus J, Lewis SE (May 2010). "Clinical significance of sperm DNA damage in assisted reproduction outcome". Hum Reprod 25 (7): 1594–1608. doi:10.1093/humrep/deq103. PMID 20447937.
  2. 2.0 2.1 2.2 Speyer BE, Pizzey AR, Ranieri M, Joshi R, Delhanty JD, Serhal P (May 2010). "Fall in implantation rates following ICSI with sperm with high DNA fragmentation". Hum Reprod 25 (7): 1609–1618. doi:10.1093/humrep/deq116. PMID 20495207.
  3. 3.0 3.1 Zhang LH, Qiu Y, Wang KH, Wang Q, Tao G, Wang LG (June 2009). "Measurement of sperm DNA fragmentation using bright-field microscopy: comparison between sperm chromatin dispersion test and terminal uridine nick-end labeling assay". Fertil. Steril. 94 (3): 1027–1032. doi:10.1016/j.fertnstert.2009.04.034. PMID 19505686.
  4. Williamson, Robert. "Properties of rapidly labelled deoxyribonucleic acid fragments isolated from the cytoplasm of primary cultures of embryonic mouse liver cells". Article. Elsevier Ltd. Retrieved 8 April 2013.
  5. Quail, Michael Andrew. "DNA: Mechanical Breakage". Online Library. John Wiley & Sons. Retrieved 1 April 2013.
  6. Phillips, Thearesa. "Restriction Enzymes Explained". Biotech / Biomedical. About.com. Retrieved 2 April 2013.
  7. 7.0 7.1 7.2 7.3 7.4 7.5 "DNA Fragmentation". New England Biolabs. Retrieved 2 April 2013.
  8. Sambrook, Joseph, Russell, David W. "Fragmentation of DNA by Nebulization". Article. Cold Spring Harbor Laboratory Press. Retrieved 3 April 2013.
  9. Enari, Masato; Sakahira, Hideki; Yokoyama, Hideki; Okawa, Katsuya; Iwamatsu, Akihiro; Nagata Shigekazu. "A caspase-activated DNase that degrades DNA during apoptosis, and its inhibitor ICAD". Article. Nature Publishing Group. Retrieved 8 April 2013.
  10. 10.0 10.1 "DNA Forensics". U.S. Department of Energy Genome Programs. Retrieved 8 April 2013.