Regulation of CRISPR-Based Immune Responses

Nucleic acid cleaving CRISPR effector complexes, consisting of Cas protein(s) and crRNAs, provide protection against invading genetic elements, such as phage and (conjugative) plasmids. However, under some conditions, cells may experience a selective advantage if they avoid energy investment in CRISPR defense, for example, if they contain additional defense systems (e.g., R-M systems, phage exclusion systems) that provide sufficient protection. The formation of CRISPR effector complexes is a multistep process that requires (1) expression of the cas genes, (2) assembly of the Cas proteins into a multiprotein complex, (3) transcription of a CRISPR array into a pre-crRNA molecule, and (4) the subsequent sequence-specific processing of the pre-crRNA by a dedicated endoribonuclease, yielding crRNAs that are then loaded on the Cas protein complex. The resulting ribonucleoprotein complex may have intrinsic cleavage activity on complementary nucleic acids (e.g., the RAMP module complex of Pyrococcus furiosus) or may to this end require recruitment of an additional component upon target binding (e.g., Cas3 recruitment by Cascade in Escherichia coli). The different steps toward the formation of the final effector complexes offer several potential targets for regulation of the CRISPR system. Although studies dealing with this regulation are limited and thus far restricted to a few organisms, the number of host factors involved in CRISPR regulation increases rapidly. CRISPR defense can be regulated at the level of (cas gene and/or CRISPR) transcription by DNA-binding global regulators such as H-NS, LeuO, cAMP-CRP, or at the post-transcriptional level by the chaperon HtpG, which has been shown to be essential for Cas3 activity in E. coli. The presence of σ32-dependent promoters within the cas operon and the involvement of the BaeSR two-component system suggest a coupling of CRISPR activity to membrane or heat stress in E. coli. In this chapter, we will summarize the recent findings on the regulation of the CRISPR system, mainly in E. coli, for which several regulatory components have been identified. We will also discuss the role of other potential regulatory mechanisms, such as translational regulation of cas gene expression through overlapping open reading frames on a polycistronic mRNA and the regulation of pre-crRNA stability or processing (Fig. 4.1).