Photochemical Nickel-Catalyzed Reductive Migratory Cross-Coupling of Alkyl Bromides with Aryl Bromides

A novel method to access 1,1-diarylalkanes from readily available, nonactivated alkyl bromides and aryl bromides via visible-light-driven nickel and iridium dual catalysis, wherein diisopropylamine (^<i>i</i>Pr₂NH) is used as the terminal stoichiometric reductant, is reported. Both primary and secondary alkyl bromides can be successfully transformed into the migratory benzylic arylation products with good selectivity. Additionally, this method showcases tolerance toward a wide array of functional groups and the presence of bases

Densitometry for yeast cycloheximide chase experiments.

(XLSX)</p

Colony count for DRR assays.

(XLS)</p

List of yeast strains used in this study.

(XLSX)</p

Overexpression of <i>RECQL4</i> results in increased RAD51 foci and decreased tail moment.

(A) Overexpression of RECQL4 results in increased RAD51 foci, which is dependent on its helicase activity. U2OS cells were transfected with an empty plasmid or a plasmid expressing RECQL4 or RECQL4-K508A under a CMV promoter. The cells were either mock or cisplatin treated for one hour and after a two-hour recovery, imaged for RAD51 foci or DAPI by immunofluorescence. RAD51 foci was quantified from 200 cells per condition for each experiment. The experiment was performed three to five times and the median was graphed (Unprocessed foci count in S9 Data). Representative images are shown. (B) Overexpression of RECQL4 results decreased tail moment following cisplatin exposure, which is dependent on its helicase activity. U2OS cells were treated similarly to the immunofluorescence experiment, before being harvested for neutral comet assay. At least 40 comets were counted per condition for each experiment. The experiment was performed four times and the mean and standard deviation was graphed (Unprocessed tail moments in S10 Data).</p

Unprocessed data for MTS assay.

(XLSX)</p

Hrq1 function and regulation in ICL repair is distinct from intrastrand crosslink repair.

(A) Hrq1 functions in a different pathway as Rev1 and Ubc13 to repair ICL. The indicated yeast strains were five-fold serial diluted onto YPD medium containing DMSO and/or YPD medium containing the indicated amount of MMC. The plates were photographed after 2 days of incubation at 30°C in the dark. (B) Hrq1 protein levels does not decreased upon 100 μg/ml MMC treatment. Exponentially growing cells with Hrq1-9xMYC were incubated with cycloheximide in the presence or absence of 100 μg/ml MMC and/or 0.1% DMSO. Protein extracts were analyzed by western blot for Hrq1 protein levels (α-MYC) or a loading control, Kar2 (α-Kar2) at the indicated time points. Quantification from three separate experiment is shown, with mean and SEM graphed (Raw densitometry data in Sheets S-U in S1 Data). (C) Hrq1 functions in the same pathway as Rad5 and Ubc13 to repair intrastrand adducts. The indicated yeast strains were five-fold serial diluted onto YPD medium before being treated with the indicated dosages of UV-C. The plates were photographed after 2 days of incubation at 30°C in the dark. (D) Hrq1 functions in different pathway as Rev1 to repair intrastrand adducts. The indicated yeast strains were five-fold serial diluted onto YPD medium before being treated with the indicated dosage of UV-C. The plates were photographed after 2 days of incubation at 30°C in the dark. (E) Hrq1 protein levels decreased following UV-C treatment. Exponentially growing Hrq1-9xMYC cells were incubated with cycloheximide then treated with indicated dosage of UV-C. Protein was extracted similarly to (B). Quantification is from three separate experiments with mean and SEM graphed (Raw densitometry data in Sheets V-X in S1 Data). (TIF)</p

Tail moment data from neutral comet assay.

(XLSX)</p

Hrq1 functions during error-free post-replicative repair.

(A) Hrq1 expression increases during S/G2 and then plateaus. Hrq1-9xMYC expressing cells were either untreated (asynchronous, AS) or cell cycle arrested in G1 with α-factor. The α-factor arrested cells were subsequently released into fresh YPD medium (0 min) and grown for 120 min. Protein samples from the indicated time points were analyzed by western blot for Hrq1 (anti-MYC), the G2/M cyclin, Clb2, (anti-Clb2), or a loading control, Kar2 (anti-Kar2). Quantification from three experiments, the mean with SEM is shown (Raw densitometry data in S2 Data). The cell cycle stage was analyzed FACS. (B) Schematic of post-replicative repair pathways. When the replication fork stalls, Pol30 (yellow trimer) is initially monoubiquitylated (Ub, purple circle) via Rad6-Rad18 (light and dark blue circles) on lysine 164 (K164). Monoubiquitylation of PCNA recruits the error-prone translesion synthesis polymerases, i.e. Rev1 (gray circle) to bypass the lesion. Alternatively, PCNA is further poly-ubiquitylated by Rad5-Ubc13-Mms2 (light, medium, and dark purple complex). Polyubiquitylation of PCNA mediates error-free repair through template switching, which is a homolog directed process. A bypass intermediate is shown with the newly synthesized DNA in red and the lesion as a yellow star. (C, D) Hrq1 functions in the same pathway as Rad6 and Pol30. The indicated yeast strains were five-fold serial diluted onto SC medium containing DMSO and/or SC medium containing the indicated amount of cisplatin. The plates were photographed after 2 days of incubation at 30°C in the dark. (E) Hrq1 functions in the same pathway as Rad5 and Ubc13. Serial dilutions of the indicated yeast strains were performed as described in (C). (F) Hrq1 functions in a different pathway than Rev1. Serial dilutions of the indicated yeast strains were performed as described in (C).</p

9-Myc tagging Hrq1 does not result in cisplatin sensitivity.

(A) Hrq1-9myc cells displayed similar cisplatin sensitivity compared to WT cells. The indicated yeast strains were five-fold serial diluted onto SC medium containing DMSO and/or SC medium containing the indicated amount of cisplatin. The plates were photographed after 2 days of incubation at 30°C in the dark. (B) Hrq1 protein level is unchanged with HU and MMS treatment. Hrq1-9myc cells were grown to log phase and then treated with indicated agents for two hours before being collected for TCA prep. Cisplatin was used at 100 μg/ml (Raw densitometry data in S11 Data). (TIF)</p