Posttranscriptional control of puc operon expression of B800-850 light-harvesting complex formation in Rhodobacter sphaeroides

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Abstract
The puc operon of Rhodobacter sphaeroides comprises the pucBA structural genes which encode B800-850 light-harvesting beta and alpha polypeptides, respectively. Northern (RNA) blot hybridization analysis of puc operon expression has identified two pucBA-specific transcripts. The small (0.5-kilobase [kb]) transcript encodes the beta and alpha polypeptides and, under photoheterotrophic growth conditions, was approximately 200-fold more abundant than the large (2.3-kb) transcript. The 5' end of the 0.5-kb transcript was mapped at 117 nucleotides upstream from the start of pucB. The 3' ends of the 0.5-kb transcript were mapped to two adjacent nucleotides, which follow a stem-loop structure immediately 3' to the pucA stop codon. Two mutant strains, PUC705-BA and PUC-Pv, were constructed by replacement of the pucBA genes and adjacent DNA in the former case or by insertional interruption of the DNA downstream of the pucBA genes in the latter case. The two mutant strains were devoid of B800-850 complexes during photosynthetic growth but were otherwise apparently normal. The B800-850 phenotype of both PUC705-BA and PUC-Pv was not complemented in trans with a 2.5-kb PstI restriction endonuclease fragment extending from 0.75 kb upstream of pucBA to 1.3 kb downstream of pucBA, despite the presence of the 0.5-kb pucBA-specific transcript. Both of the mutant strains, however, showed restoration of B800-850 expression with a 10.5-kb EcoRI restriction endonuclease fragment in trans encompassing the 2.5-kb PstI fragment. Western immunoblot analysis revealed no B800-850-beta polypeptide as well as no polypeptide designated 15A in either mutant. Nonetheless, under photoheterotrophic growth conditions, the 0.5-kb pucBA-specific transcript was present in PUC-Pv, although no 2.3-kb transcript was detectable. We suggest that the DNA region immediately downstream of pucBA encodes a gene product(s) essential for translational or posttranslational expression of the B800-850 beta and alpha polypeptides.