Four distinct DNA ligase activities (I-IV) have been identified within mammalian cells. Evidence has indicated DNA ligase I to be central to lagging strand DNA replication, as well as being involved in DNA repair processes. It has also been reported that DNA ligase I is essential for cell viability. A patient with altered DNA ligase I displayed a phenotype similar to Bloom's Syndrome, being immunodeficient, growth retarded, and predisposed to cancer. Fibroblasts isolated from this patient (46BR) exhibited abnormal lagging strand DNA synthesis, and repair deficiency. Gene inactivation was the first step of a 'double replacement' strategy to introduce analogous mutations into the mouse DNA ligase I gene (<I>LigI</I>). Using gene targeting in HPRT-deficient embryonic stem (ES) cells, the last 5 exons of the endogenous <I>Lig1</I> gene were replaced by an <I>HPRT</I> minigene, and subsequently DNA ligase I-deficient mice were produced. Embryos lacking DNA ligase I developed normally to mid-term, when haematopoiesis usually switches to the foetal liver. Thereupon severe disruption to foetal liver erythropoiesis occurred, leading to acute anaemia and prenatal death. <I>In vitro</I> assays indicated that erythroid-committed progenitor cells were present in the liver, but in reduced numbers. Apart from the developing liver, organogenesis was not perceptibly affected and non-erythroid haematopoietic lineages were not reduced. Injection of cell suspensions from single foetal livers were unable to rescue lethally-irradiated mice. However, injection of multiple livers into a single recipient was able to effect long term repopulation of the haematopoietic system. Animals rescued with cells lacking DNA ligase I displayed splenomegaly, anaemia coupled with reticulocytosis, and aplasia in all haematopoietic tissues.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:641545 |
| Date | January 1999 |
| Creators | Bentley, Darren James |
| Publisher | University of Edinburgh |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/1842/11937 |
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