CHARACTERIZATION OF A NEW RHO-MUTATION THAT RELIEVES POLARITY OF MU-INSERTIONS

We report the identification and characterization of a new rho mutatio n, rho614 that relieves polarity of Mu insertions in Escherichia coli. The mutation was identified by its ability to suppress the polarity o f the Mu-mediated Phi(lamB'-'lacZ)(hyb)61-4 fusion that is located at codon four of the lame signal sequence. The rho614 mutation alters res idue 80 in the proposed RNA-binding domain of Rho and is recessive to wildtype rho. We suggest that in the presence of the rho614 allele tra nscripts initiated at the Mu promoter P-cM fail to terminate at presum ptive Rho-dependent termination sites, namely rut1 and rut2, and conti nue into the 3' 'lamB gene allowing a LamB(+) phenotype. This contenti on is supported by deletion analysis of the region and the observation that insertional inactivation of genes that reduce transcription from P-cM, clpP (ATP protease), himA (IHF-alpha), and himD (IHF-beta), blo ck the LamB(+) phenotype. rho614, rho4 and rho201 alleles suppress the polarity of a malK::Mu insertion on the downstream lamB gene. However , the polarity of the Phi,(lamB'-'lacZ)(hyb)61-4 insertion is only sup pressed by the rho614 mutation. We propose that the rho614 mutation al lows suppression of transcriptional polarity without interfering with translation initiation signals of the truncated 'lamB gene. In additio n to identifying a new rho mutation and Rho-dependent terminator seque nce, this system provides a means of studying Rho protein/terminator r elationships through the identification of new classes of rho mutation s.