A CRISPR-associated factor Csa3a regulates DNA damage repair in Crenarchaeon Sulfolobus islandicus

Abstract

CRISPR−Cas system provides acquired immunity against invasive genetic elements in prokaryotes. In both bacteria and archaea, transcriptional factors play important roles in regulation of CRISPR adaptation and interference. In the model Crenarchaeon Sulfolobus islandicus, a CRISPR-associated factor Csa3a triggers CRISPR adaptation and activates CRISPR RNA transcription for the immunity. However, regulation of DNA repair systems for repairing the genomic DNA damages caused by the CRISPR self-immunity is less understood. Here, according to the transcriptome and reporter gene data, we found that deletion of the csa3a gene down-regulated the DNA damage response (DDR) genes, including the ups and ced genes. Furthermore, in vitro analyses demonstrated that Csa3a specifically bound the DDR gene promoters. Microscopic analysis showed that deletion of csa3a significantly inhibited DNA damage-induced cell aggregation. Moreover, the flow cytometry study and survival rate analysis revealed that the csa3a deletion strain was more sensitive to the DNA-damaging reagent. Importantly, CRISPR self-targeting and DNA transfer experiments revealed that Csa3a was involved in regulating Ups- and Ced-mediated repair of CRISPR-damaged host genomic DNA. These results explain the interplay between Csa3a functions in activating CRISPR adaptation and DNA repair systems, and expands our understanding of the lost link between CRISPR self-immunity and genome stability.

Cyclic Tetra-Adenylate (cA4) Recognition by Csa3; Implications for an Integrated Class 1 CRISPR-Cas Immune Response in Saccharolobus solfataricus

“…Regardless of the specific mechanisms of transcriptional activation and repression by Csa3a and other Csa3 family members, confirmation of cA 4 as a Csa3a ligand in S. solfataricus now provides a critical, direct link between the type III surveillance complexes that synthesize cA 4 upon detection of invading nucleic acid [ 53 ], and Csa3 family transcription factors that regulate acquisition gene expression, transcription of CRISPR loci, DNA repair and potentially the transcription and regulation of type I-A surveillance complex activity [ 6 , 23 , 24 , 25 , 54 ]. Thus, the general pathway for Csa3a-mediated spacer acquisition can still be inferred ( Figure 10 ).…”

Cas1 and Fen1 Display Equivalent Functions During Archaeal DNA Repair

“…Another example illustrating the interplay of the CRISPR immune system and DNA repair was revealed in Sulfolobus solfataricus. Here, the CRISPR-associated protein Csa3a controls the expression of major CRISPR adaptation genes (Liu et al, 2015), is a key player in the activation of DNA repair genes (Liu et al, 2017), and regulates the DNA damage response (DDR) (Liu et al, 2020). The reason for the synergistic activation of both CRISPR-Cas and DNA repair pathways by Csa3a is probably due to the frequent acquisition of spacers from its own genome (roughly 7%) (Liu et al, 2017).…”

“…The reason for the synergistic activation of both CRISPR-Cas and DNA repair pathways by Csa3a is probably due to the frequent acquisition of spacers from its own genome (roughly 7%) (Liu et al, 2017). The simultaneous activation of DDR genes thus reduces auto-immunity effects caused by self-targeting spacers (Liu et al, 2020).…”