The cellular response to genotoxic stress is mediated by a well-characterized 
network of DNA surveillance pathways. The contribution of post-transcriptional 
gene regulatory networks to the DNA damage response (DDR) has not been 
extensively studied. Here, we systematically identified RNA-binding proteins 
differentially interacting with polyadenylated transcripts upon exposure of 
human breast carcinoma cells to ionizing radiation (IR). Interestingly, more 
than 260 proteins, including many nucleolar proteins, showed increased binding 
to poly(A)+ RNA in IR-exposed cells. The functional analysis of DDX54, a 
candidate genotoxic stress responsive RNA helicase, revealed that this protein 
is an immediate-to-early DDR regulator required for the splicing efficacy of its 
target IR-induced pre-mRNAs. Upon IR exposure, DDX54 acts by increased 
interaction with a well-defined class of pre-mRNAs that harbor introns with weak 
acceptor splice sites, as well as by protein-protein contacts within components 
of U2 snRNP and spliceosomal B complex, resulting in lower intron retention and 
higher processing rates of its target transcripts. Because DDX54 promotes 
survival after exposure to IR, its expression and/or mutation rate may impact 
DDR-related pathologies. Our work indicates the relevance of many 
uncharacterized RBPs potentially involved in the DDR.