The requirement for recombination factors differs considerably between different pathways of homologous double‐strand break repair in somatic plant cells

Abstract

SUMMARYIn recent years, multiple factors involved in DNA double-strand break (DSB) repair have been characterised in Arabidopsis thaliana. Using homologous sequences in somatic cells, DSBs are mainly repaired by two different pathways: synthesis-dependent strand annealing (SDSA) and single-strand annealing (SSA). By applying recombination substrates in which recombination is initiated by the induction of a site-specific DSB by the homing endonuclease I-SceI, we were able to characterise the involvement of different factors in both pathways. The nucleases MRE11 and COM1, both involved in DSB end processing, were not required for either SDSA or SSA in our assay system. Both SDSA and SSA were even more efficient without MRE11, in accordance with the fact that a loss of MRE11 might negatively affect the efficiency of non-homologous end joining. Loss of the classical recombinase RAD51 or its two paralogues RAD51C and XRCC3, as well as the SWI2/SNF2 remodelling factor RAD54, resulted in a drastic deficiency in SDSA but had hardly any influence on SSA, confirming that a strand exchange reaction is only required for SDSA. The helicase FANCM, which is postulated to be involved in the stabilisation of recombination intermediates, is surprisingly not only needed for SDSA but to a lesser extent also for SSA. Both SSA and SDSA were affected only weakly when the SMC6B protein, implicated in sister chromatid recombination, was absent, indicating that SSA and SDSA are in most cases intrachromatid recombination reactions.

A DNA2 Homolog Is Required for DNA Damage Repair, Cell Cycle Regulation, and Meristem Maintenance in Plants

“…The expression of BRCA1, which functions in genome surveillance and DNA damage repair (Wang et al, 2000;Lafarge and Montané, 2003), was increased more than 4-fold in jhs1, suggesting that much DNA damage occurs in this mutant. The GR1 and MRE11 nucleases, which are involved in DSB end processing (Bundock and Hooykaas, 2002;Uanschou et al, 2007;Roth et al, 2012), were also induced in the mutant. The expression level of RAD51, which mediates DNA repair directly by homologous recombination (Li et al, 2004), was increased more than 4-fold in jhs1.…”

“…BREAST CANCER SUSCEPTIBILITY1 (ATBRCA1) is a homolog of the human breast cancer susceptibility gene and has an important role in DNA damage repair as a key component of the BRCA1-associated genome surveillance complex (Wang et al, 2000;Lafarge and Montané, 2003). GAMMA RESPONSE1 and MRE11 are nucleases involved in DSB end-processing in the repair of crosslinked DNA (Bundock and Hooykaas, 2002;Uanschou et al, 2007;Roth et al, 2012). KU70 plays an important role in the maintenance of the telomeric region and DSB repair by modulating the nonhomologous end-joining pathway (Tamura et al, 2002).…”

DNA Damage Repair in the Context of Plant Chromatin

“…More recently, RAD54 was specified to participate mainly in the synthesis-dependent strandannealing mechanism of DSB repair by HR (Roth et al, 2012).…”

“…Chromatin is expected to especially affect the access, resection, and restoration of the original arrangement, and several factors that can dissolve higher order structures, slide, evict, or exchange nucleosomes, or restore chromatin have been implicated in DDR (for review, see Lukas et al, 2011;Czaja et al, 2012;Dinant et al, 2012;Euskirchen et al, 2012;Lans et al, 2012;Soria et al, 2012;Altmeyer and Lukas, 2013;Dion and Gasser, 2013;Herceg, 2013a, 2013b;Ohsawa et al, 2013;PapamichosChronakis and Peterson, 2013;Peterson and Almouzni, 2013;Price and D'Andrea, 2013;Stanley et al, 2013;House et al, 2014;Jeggo and Downs, 2014;Swygert and Peterson, 2014;Polo, 2015). The repair process in plants has been the focus of several reviews (Roth et al, 2012;Donà et al, 2013;Knoll et al, 2014a;Missirian et al, 2014). In this Update, we summarize recent literature describing the connection of repair and chromatin in plants, adding to earlier reviews (Balestrazzi et al, 2011;Zhu et al, 2011;Roy, 2014).…”

The moss Physcomitrella patens: methods and tools from cultivation to targeted analysis of gene function

“…These specific features make P. patens highly attractive, not only as model organism for the scientific community, but also for the pharmaceutical industry as a producer of biopharmaceutical agents. homologous end joining (NHEJ) and the HR, are highly conserved DNA DSB repair mechanisms (Pâques and Haber, 1999) that can be found in all three domains of life and allow a cell to keep its genome fully functional (Sung and Klein, 2006;Roth et al, 2012). The NHEJ operates via illegitimate recombination (IR) without sequence-specific requirements, whereas HR depends on sequence similarity for the DSB repair (Fig.…”