Roadblocks & bypasses : protection of genome stability by translesion DNA synthesis in C. elegans

DNA encodes the genetic instructions for living organisms. However, damage to the DNA is inevitable, because DNA itself is an unstable molecule and environmental factors such as UV-radiation or X-rays cause damage to the DNA. A certain type of DNA damages can block DNA replication, an essential step before cell can divide. The polymerases that normally replicate DNA are incredibly efficient and virtually flawless on undamaged DNA, but they cannot replicate damaged DNA. In multi-celled organisms, the most important defense mechanism against this is Translesion DNA synthesis (TLS). TLS protects against various negative consequences of damage to the DNA. For this, TLS utilizes specialized TLS polymerases that can replicate damaged DNA.My experiments show that the strong evolutionary conservation of TLS is explained by the dual functions of TLS: guarding replication potential and genome stability. TLS suppresses genomic instability, by preventing conversion of replication blocks to double-stranded DNA breaks (DSBs). Without functional TLS, DSBs arise and result in larger and more harmful mutations. TLS is beneficial for organisms because it supports continuous reproduction and growth. Although DNA damage is always present and unavoidable, TLS guards against the formation of mutations that would otherwise lead to cancer, aging and congenital disease.Financial support for the printing of this thesis was provided by MRC Holland BV.LUMC / Geneeskund

Translesion synthesis polymerases are dispensable for C. elegans reproduction but suppress genome scarring by polymerase theta-mediated end joining

Author summaryResearch in the fields of DNA repair and mutagenesis has led to enormous insight into the mechanisms responsible for maintaining genetic integrity. However, which processes drive de novo mutations and will thus contribute to inherited diseases are still unclear. One process thought to underlie spontaneous mutagenesis is replication of damaged DNA by specialised so-called "Translesion synthesis" polymerases, which have the ability to replicate across damaged bases, but are not very accurate. To address the impact of TLS or the lack thereof on genome integrity, we have knocked out all TLS enzymes that are encoded by the C. elegans genome, individually and in combination, and monitored mutation accumulation during prolonged culturing of these animals without external sources of DNA damage. We found that TLS is not the major driver of spontaneous mutagenesis in this organism, however, it protects the genome from harmful small deletions that result from mutagenic repair of DNA breaks. We also found that, contrary to what was expected, TLS activity is not essential for reproduction in a multicellular organism with the tissue complexity and genome size of C. elegans.Bases within DNA are frequently damaged, producing obstacles to efficient and accurate DNA replication by replicative polymerases. Translesion synthesis (TLS) polymerases, via their ability to catalyze nucleotide additions to growing DNA chains across DNA lesions, promote replication of damaged DNA, thus preventing checkpoint activation, genome instability and cell death. In this study, we used C. elegans to determine the contribution of TLS activity on long-term stability of an animal genome. We monitored and compared the types of mutations that accumulate in REV1, REV3, POLH1 and POLK deficient animals that were grown under unchallenged conditions. We also addressed redundancies in TLS activity by combining all deficiencies. Remarkably, animals that are deficient for all Y-family polymerases as well as animals that have lost all TLS activity are viable and produce progeny, demonstrating that TLS is not essential for animal life. Whole genome sequencing analyses, however, reveal that TLS is needed to prevent genomic scars from accumulating. These scars, which are the product of polymerase theta-mediated end joining (TMEJ), are found overrepresented at guanine bases, consistent with TLS suppressing DNA double-strand breaks (DSBs) from occurring at replication-blocking guanine adducts. We found that in C. elegans, TLS across spontaneous damage is predominantly error free and anti-clastogenic, and thus ensures preservation of genetic information.Genome Instability and Cance

Roadblocks & bypasses : protection of genome stability by translesion DNA synthesis in C. elegans

DNA encodes the genetic instructions for living organisms. However, damage to the DNA is inevitable, because DNA itself is an unstable molecule and environmental factors such as UV-radiation or X-rays cause damage to the DNA. A certain type of DNA damages can block DNA replication, an essential step before cell can divide. The polymerases that normally replicate DNA are incredibly efficient and virtually flawless on undamaged DNA, but they cannot replicate damaged DNA. In multi-celled organisms, the most important defense mechanism against this is Translesion DNA synthesis (TLS). TLS protects against various negative consequences of damage to the DNA. For this, TLS utilizes specialized TLS polymerases that can replicate damaged DNA.My experiments show that the strong evolutionary conservation of TLS is explained by the dual functions of TLS: guarding replication potential and genome stability. TLS suppresses genomic instability, by preventing conversion of replication blocks to double-stranded DNA breaks (DSBs). Without functional TLS, DSBs arise and result in larger and more harmful mutations. TLS is beneficial for organisms because it supports continuous reproduction and growth. Although DNA damage is always present and unavoidable, TLS guards against the formation of mutations that would otherwise lead to cancer, aging and congenital disease.</table

Combined loss of three DNA damage response pathways renders C-elegans intolerant to light

Genome Instability and Cance

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