The contributions of the Sgs1, Mph1, and Srs2 DNA helicases during mitotic double-strand break (DSB) repair in yeast were investigated using a gap-repair assay. A diverged chromosomal substrate was used as a repair template for the gapped plasmid, allowing mismatch-containing heteroduplex DNA (hDNA) formed during recombination to be monitored. Overall DSB repair efficiencies and the proportions of crossovers (COs) versus noncrossovers (NCOs) were determined in wild-type and helicase-defective strains, allowing the efficiency of CO and NCO production in each background to be calculated. In addition, the products of individual NCO events were sequenced to determine the location of hDNA. Because hDNA position is expected to differ depending on whether a NCO is produced by synthesis-dependent-strand-annealing (SDSA) or through a Holliday junction (HJ)–containing intermediate, its position allows the underlying molecular mechanism to be inferred. Results demonstrate that each helicase reduces the proportion of CO recombinants, but that each does so in a fundamentally different way. Mph1 does not affect the overall efficiency of gap repair, and its loss alters the CO-NCO by promoting SDSA at the expense of HJ–containing intermediates. By contrast, Sgs1 and Srs2 are each required for efficient gap repair, strongly promoting NCO formation and having little effect on CO efficiency. hDNA analyses suggest that all three helicases promote SDSA, and that Sgs1 and Srs2 additionally dismantle HJ–containing intermediates. The hDNA data are consistent with the proposed role of Sgs1 in the dissolution of double HJs, and we propose that Srs2 dismantles nicked HJs.

Natural compounds capable of increasing root depth and branching are desirable tools for enhancing stress tolerance in crops. We devised a sensitized screen to identify natural metabolites capable of regulating root traits in Arabidopsis. β-Cyclocitral, an endogenous root compound, was found to promote cell divisions in root meristems and stimulate lateral root branching. β-Cyclocitral rescued meristematic cell divisions in ccd1ccd4 biosynthesis mutants, and β-cyclocitral–driven root growth was found to be independent of auxin, brassinosteroid, and reactive oxygen species signaling pathways. β-Cyclocitral had a conserved effect on root growth in tomato and rice and generated significantly more compact crown root systems in rice. Moreover, β-cyclocitral treatment enhanced plant vigor in rice plants exposed to salt-contaminated soil. These results indicate that β-cyclocitral is a broadly effective root growth promoter in both monocots and eudicots and could be a valuable tool to enhance crop vigor under environmental stress.

SUMMARY Heteroduplex DNA (hetDNA) is a key molecular intermediate during the repair of mitotic double-strand breaks by homologous recombination, but its relationship to 5′-end resection and/or 3′-end extension is poorly understood. In the current study, we examined how perturbations in these processes affect the hetDNA profile associated with repair of a defined double-strand break (DSB) by the synthesis-dependent strand-annealing (SDSA) pathway. Loss of either the Exo1 or Sgs1 long-range resection pathway significantly shortened hetDNA, suggesting that these pathways normally collaborate during DSB repair. In addition, altering the processivity or proofreading activity of DNA polymerase δ shortened hetDNA length or reduced break-adjacent mismatch removal, respectively, demonstrating that this is the primary polymerase that extends both 3′ ends. Data are most consistent with the extent of DNA synthesis from the invading end being the primary determinant of hetDNA length during SDSA.

During the transition from darkness to light, a suite of light sensors guides gene expression, biochemistry, and morphology to optimize acclimation to the new environment. Ultraviolet, blue, red, and far-red light all have demonstrated roles in modulating light responses, such as changes in gene expression and suppression of stem growth rate. However, green wavebands induce stem growth elongation, a response not likely mediated by known photosensors. In this study, etiolated Arabidopsis (Arabidopsis thaliana) seedlings were treated with a short, dim, single pulse of green light comparable in fluence and duration to that previously shown to excite robust stem elongation. Genome microarrays were then used to monitor coincident changes in gene expression. As anticipated, phytochrome A-regulated, nuclear-encoded transcripts were induced, confirming proper function of the sensitive phytochrome system. In addition, a suite of plastid-encoded transcripts decreased in abundance, including several typically upregulated after phytochrome and/or cryptochrome activation. Further analyses using RNA gel-blot experiments demonstrated that the response is specific to green light, fluence dependent, and detectable within 30 min. The response obeys reciprocity and persists in the absence of known photosensors. Plastid transcript down-regulation was also observed in tobacco (Nicotiana tabacum) with similar temporal and fluence-response kinetics. Together, the down-regulation of plastid transcripts and increase in stem growth rate represent a mechanism that tempers progression of early commitment to the light environment, helping tailor seedling development during the critical process of establishment.