Figure 2. Effect of RsmY and RsmZ overexpression on expression of the exoS gene and on biofilm development.

A. Expression of the exoS gene was measured using Miller assays in P. aeruginosa strains carrying an exoS-lacZ reporter fusion and rsmY or rsmZ cloned with their native promoter into a promoterless plasmid pPSV40, or under the control of a constitutively active PlacUV5 promoter in a derivative of the pMMB67 plasmid (See Experimental procedures). As induction of the T3S genes occurs under conditions of low calcium, cultures were grown in Ca²⁺-depleted (grey bars) and Ca²⁺-replete (white bars) LB media, at 37°C for 4 hours.

B. Biofilm phenotypes were measured in P. aeruginosa wild type and gacA mutant strains with rsmY or rsmZ cloned with their native promoter into a promoterless plasmid pPSV40, or under the control of a constitutively active PlacUV5 promoter in a derivative of the pMMB67 plasmid. Biofilms were scored as the attachment of P. aeruginosa to tubes and formation of the biofilm rings and pellicles after 20 h static incubation at 30°C.