DOI: 10.5281/zenodo.16362591
title: van Der Sluijs et al. "DNA polymerase kappa is the primary translesion synthesis polymerase for aldehyde ICLs"

description: Paired-end raw 150 base-pair Illumina short-read sequencing data corresponding to four experimental replicate libraries: MutSeq 8, MutSeq 9, Mutseq 9 - depletion, and Mutseq 10 - depletion.

Abstract: 

DNA interstrand crosslinks (ICLs) are highly cytotoxic lesions that block essential cellular processes like replication and transcription. Endogenous ICLs can be induced by reactive aldehydes produced during normal cellular metabolism. Defective repair of these aldehyde-induced ICLs is associated with Fanconi anemia (FA), a cancer predisposition syndrome. We previously showed that acetaldehyde-induced ICLs are repaired by the Fanconi anemia (FA) pathway and a novel excision-independent pathway. Here, we demonstrate that ICLs induced by acrolein, another cellular aldehyde, are also repaired by both pathways, establishing the generality of aldehyde ICL repair. Focusing on the FA pathway, we identify DNA polymerase kappa (Polκ) as the primary translesion synthesis (TLS) polymerase responsible for the insertion step during lesion bypass of unhooked aldehyde ICLs. This function requires Polκ's catalytic activity and PCNA interaction domains but is independent of Rev1 interaction. In contrast, Polκ has a minor non-catalytic role in the extension step of cisplatin ICL repair that is dependent on Rev1 interaction. Our work reveals a key role for Polκ in aldehyde ICL repair and provides mechanistic insights into how different ICL structures determine the choice of TLS polymerases during repair.

 

 
description: The following Tables 1-3 below provide information on the barcodes and sample conditions for all samples used to generate the pooled libraries in this study. Some barcodes were used for different sample conditions and contexts, hence the necessity to refer to these tables. Left and right flanking sequences and corresponding reference plasmid are provided for generation of mutation profiles with the SIQ program (van Schendel et al. 2022). Processed reads from corresponding barcodes/samples were mapped to the matching plasmid reference with bwamem2 (Vasimuddin et al. 2019).

 

Table 1: Mutseq 8 and Mutseq 9 Library Barcodes and Sample Conditions. 






Condition



Left Flank



Right Flank



ID



Barcode



Reference





Pt



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B1



CAGTTAGT



CTRL.fa





AA-red



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B2



AATAGGAT



CTRL.fa





Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



B3



AACACGTG



Ctrl-Acr-red.fa





AA-nat



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B4



TGTAAAGG



CTRL.fa





Acr-nat



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B5



GAATCTAG



CTRL.fa





mono-AA



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B6



GAGTTCTC



CTRL.fa





mono-Acr



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B7



AACTGAGC



CTRL.fa





Ctrl



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B8



GCAAGTAC



CTRL.fa





Ctrl-Acr



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



B9



GATTCGTA



Ctrl-Acr-red.fa





AA-nat DMSO



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



A1



TAGGTGTT



CTRL.fa





Acr-nat DMSO



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



A2



AGCTACGT



CTRL.fa





AA-nat p97i



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



A3



TCCTATAT



CTRL.fa





Acr-nat p97i



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



A4



CAACTGTG



CTRL.fa





 

Table 2: Mutseq 9 - depletion Library Barcodes and Sample Conditions. 






Condition



Left Flank



Right Flank



ID



Barcode



Reference





Mock Pt



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B1



CAGTTAGT



CTRL.fa





Mock AA-red



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B2



AATAGGAT



CTRL.fa





Mock Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



B3



AACACGTG



Ctrl-Acr-red.fa





Mock Ctrl



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B4



TGTAAAGG



CTRL.fa





Mock Ctrl-Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



B5



GAATCTAG



Ctrl-Acr-red.fa





∆Rev1 Pt



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B6



GAGTTCTC



CTRL.fa





∆Rev1 AA-red



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B7



AACTGAGC



CTRL.fa





∆Rev1 Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



B8



GCAAGTAC



Ctrl-Acr-red.fa





∆Rev1 Ctrl



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B9



GATTCGTA



CTRL.fa





∆Rev1 Ctrl-Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



B10



TAGTAGGA



Ctrl-Acr-red.fa





∆PolK Pt



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



A1



TAGGTGTT



CTRL.fa





∆PolK AA-red



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



A2



AGCTACGT



CTRL.fa





∆PolK Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



A3



TCCTATAT



Ctrl-Acr-red.fa





∆PolK Ctrl



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



A4



CAACTGTG



CTRL.fa





∆PolK Ctrl-Acr



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



A5



GTTGTAGG



Ctrl-Acr-red.fa





 

Table 3: Mutseq 10 - depletion Barcodes and Sample Conditions. 






Condition



Left Flank



Right Flank



ID



Barcode



Reference





Mock Pt



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B1



CAGTTAGT



CTRL.fa





Mock AA-red



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B5



GAATCTAG



CTRL.fa





Mock Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



A1



TAGGTGTT



Ctrl-Acr-red.fa





Mock Ctrl



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B8



GCAAGTAC



CTRL.fa





Mock Ctrl-Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



A6



AGCCGACG



Ctrl-Acr-red.fa





∆Rev1 Pt



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B2



AATAGGAT



CTRL.fa





∆Rev1 AA-red



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B6



GAGTTCTC



CTRL.fa





∆Rev1 Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



A2



AGCTACGT



Ctrl-Acr-red.fa





∆Rev1 Ctrl



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B9



GATTCGTA



CTRL.fa





∆Rev1 Ctrl-Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



A8



CGAAACGC



Ctrl-Acr-red.fa





∆PolK Pt



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B4



TGTAAAGG



CTRL.fa





∆PolK AA-red



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B7



AACTGAGC



CTRL.fa





∆PolK Acr-red



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



A5



GTTGTAGG



Ctrl-Acr-red.fa





∆PolK Ctrl



AGACAGCCCTCTTCC



TCTTCTTTCGTGCGC



B10



TAGTAGGA



CTRL.fa





∆PolK Ctrl-Acr



AGACAGCCCTCTCAC



GCTTCTTTCGTGCGC



A9



GTCAGACC



Ctrl-Acr-red.fa