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 degrees 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.

PsbT Polypeptide Is Required for Efficient Repair of Photodamaged Photosystem II Reaction Center

“…However, it remains unclear whether PsbT has a direct interaction with D1. Although the psbT gene is co-expressed with the psbB gene (15,29), no biochemical evidence for functional and/or structural interaction between PsbT and CP47 was obtained in the present study. Improved resolution of the three-dimensional structure of the PSII complex might demonstrate a possible interaction between PsbT and CP47.…”

The petD gene is transcribed by functionally redundant promoters in Chlamydomonas reinhardtii chloroplasts.

“…However, instances of cotranscription in Chlamydomonas chloroplasts have been documented. For, example cotranscripts of psaA exon II and psbD have been observed in psaA processing mutants (8), and a stable psbB-ORF31 cotranscript has been detected in wildtype cells (18). Our observation of apetA-petD cotranscript in the FUD6 strain raises the possibility that other regions of the chloroplast genome are also cotranscribed.…”

“…In the unicellular green alga Chlamydomonas reinhardtii, the genetic content of the chloroplast genome is similar to that of higher plants, but the gene order is different (9,14). In contrast to higher plants, only two instances of cotranscription have been well documented, one in which the psbD gene is cotranscribed with exon II of psaA (8) and the other in which psbB is cotranscribed with a conserved open reading frame, ORF31 (18,28). Several C. reinhardtii chloroplast promoters have been identified by their ability to direct transcription of reporter genes in transformed chloroplasts (1,12,21,33).…”

Redundancy, phylogeny and differential expression of Histoplasma capsulatum catalases The GenBank accession numbers for the cDNA sequences reported in this paper are AF139985 (CATB), AF189368 (CATA) and AF189369 (CATP).

“…RNA samples were electrophoresed in formaldehyde\1% (w\v) agarose (Sambrook et al, 1989), transferred by capillary blotting to Hybond-N membrane (Amersham Pharmacia) and hybridized to radiolabelled probes in the hybridization solution of Church & Gilbert (1984) according to the procedure described by Johnson & Schmidt (1993). Results were obtained both by autoradiography and by phosphorimaging (Molecular Dynamics Storm Phosphorimager, Amersham Pharmacia Biotech).…”