CRISPR3 Regulates Exopolysaccharide Production in Myxococcus xanthus

Myxococcus xanthus, a model organism for studying development and Type IV pili (T4P), harbors three Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) on its chromosome. CRISPR systems, which function as an adaptive immune system in prokaryotes, are classified into three types based on CRISPR-associated genes. Evidence suggests that these three types mediate immunity slightly differently. M. xanthus CRISPR1 and CRISPR2 are Type I systems while CRISPR3 is a Type III-B system. In a genetic screen, a mariner transposon insertion in the 13th spacer of CRISPR3 (3SP13) was found to restore exopolysaccharide (EPS) production to a pilA mutant. Since the deletion of CRISPR3 failed to suppress a pilA mutation and expression of CRISPR3 from a heterologous promoter led to pilA suppression, it was concluded that the 3SP13 transposon insertion is a gain-of-function mutation. Deletion of the adjacent Repeat Associated Mysterious Proteins (RAMP) genes indicated that they are essential for the 3SP13 transposon insertion to suppress pilA, providing evidence that Type III-B systems may be involved in the regulation of chromosomal genes. We suggest that one of the spacers, once expressed and processed, may inhibit the expression of a negative regulator of EPS production in M. xanthus.