The psbB gene cluster of the Chlamydomonas reinhardtii chloroplast: sequence and transcriptional analyses of psbN and psbH

Abstract

We have sequenced and characterized the complete psbB gene cluster of Chlamydomonas reinhardtii chloroplast DNA. Although the petB and petD genes are located elsewhere, the sequential order of psbB, ORF31, psbN and psbH is identical to that of the psbB operon in higher plants. Also, intergenic non-coding regions are much larger in the Chlamydomonas gene cluster. Northern blot analyses indicate the formation of dicistronic transcripts of psbB and ORF31 and monocistronic transcripts of psbN and psbH. It is unclear whether a psbB operon is transcribed to yield a large polycistronic precursor but northern blot analysis with total RNA from cells grown at 15°C does not detect an increased complexity of the transcripts, as has been found in studies of the psbB operon of higher plants. From primer extension and nuclease protection assays, it is apparent that 5′ and 3′ processing of the primary psbH transcript results in the accumulation of a heterogenous population of mRNAs. Northern blot analyses reveal transcription of Chlamydomonas psbN and show that its mRNA is much larger than that identified in liverwort and pea. The sequence identities of the PSII-H and PSII-N polypeptides as compared to their vascular plant counterparts is 50 to 62%. While the amino acid sequences of PSII-H and PSII-N proteins are significantly conserved, the mass of PSII-H from Chlamydomonas is significantly larger.