3.2 Mutagenic potential of AFB1-FapyGua in
relation to mutational signature of AFB1
High levels of base substitutions at the C/G sites, predominantly C/G
> A/T transversions, is a common feature of DNA isolated
from AFB1-exposed cells or animals and human
AFB1-driven cancers (COSMIC signature SBS24)
(Chawanthayatham et al. 2017; Huang et al. 2017; Zhang et al. 2017;
Fedeles and Essigmann 2018; Kucab et al. 2019; Alexandrov et al. 2020;
Volkova et al. 2020). AFB1-FapyGua has been implicated
as a major contributor to AFB1-induced mutagenesis and
carcinogenesis (reviewed in (McCullough and Lloyd 2019)) particularly,
based on observed correlation between the mutational spectra of
AFB1 and the miscoding properties of
AFB1-FapyGua in replication bypass reactions (Lin et al.
2016) and cultured primate cells (Lin et al. 2014). However, a
limitation of these site-specific studies was that the analyses were
restricted to a single, TXA, local sequence context. Here, we
substantiated the hypothesis about a major role of
AFB1-FapyGua in AFB1-induced mutagenesis
and carcinogenesis by demonstrating its high potential for inducing base
substitutions, including high frequency G > T
transversions, in four additional sequence contexts (Figure 2).
The G > T transversions that constitute signature SBS24 are
not distributed randomly, but preferentially arise in a limited subset
of trinucleotide sequences (Chawanthayatham et al. 2017; Huang et al.
2017; Zhang et al. 2017; Fedeles and Essigmann 2018; Kucab et al. 2019;
Alexandrov et al. 2020; Volkova et al. 2020). Several mechanisms have
been considered through which this signature is manifested, including
sequence-dependent DNA adduct formation, stability of the initial
AFB1-N7-dG adduct, rates of repair, and fidelity of
replication bypass. The present data, in combination with previous
analyses (Lin et al. 2014), strongly suggest that differential fidelity
of replication bypass of AFB1-FapyGua does not account
for the occurrence of these mutational hot-spots since the adduct was
highly mutagenic in all five sequence contexts. The difference between
the hot-spot CGC sequence and three other sequences analyzed in the
present study was insignificant (Figure 2). Thus, mechanism(s) that
define sequence-dependent occurrence of AFB1-induced
mutations likely operates prior to replication. Consistent with such
possibility, we recently have demonstrated that sequence context
modulates the catalytic efficiency of human glycosylase NEIL1 acting on
AFB1-FapyGua, with rates of reactions being inversely
correlated with thermal stability of the adduct-containing DNA (Tomar et
al. 2021). We speculate that AFB1-FapyGua in a more
thermally-stable local environment would be more likely to escape repair
not only by NEIL1, but also by nucleotide excision repair and thus,
largely contribute to the final mutational signature of
AFB1. In support for a role of pre-replication events in
shaping the final mutational signature of AFB1 is the
distribution of AFB1-induced mutations throughout
genome: these are enriched in the intergenic and late-replicating
regions and the signature has a transcriptional strand asymmetry
(https://cancer.sanger.ac.uk/signatures/sbs/sbs24/).