Two functional psbD genes in the cyanobacterium Synechococcus sp. strain PCC 7942

The cyanobacterium Synechococcus sp. strain PCC 7942 has two copies of the psbD gene which encodes the D2 polypeptide of the photosystem II (PSII) reaction center. One of the genes, psbDI, overlaps the open reading frame of another photosystem II gene, psbC; the psbDII gene is monocistronic. Gene inactivation experiments had previously shown that psbDII is dispensable under normal laboratory growth conditions. However, similar experiments with psbDI never produced viable psbDI-inactivated mutants, presumably because psbC expression depends on transcription through psbDI. The experiments described here were designed to assess the need for psbDI independent of the need for expression of psbC. A strain, AMC027, was engineered in which a second copy of psbC was expressed from the psbDII locus. Northern (RNA) blot analysis confirmed that both psbDI and psbDII gave rise to dicistronic messages containing psbC sequences in AMC027. In this genetic background, it was possible to inactivate psbDI, creating strain AMC050 and indicating that the psbDII gene is functional. Western immunoblot analysis showed that the products of psbD and psbC, the PSII proteins D2 and CP43, respectively, were present in thylakoids of AMC050, but at reduced levels relative to the wild type, the mutant AMC027, and two psbDII-inactivated mutants. AMC050 consistently formed small colonies on plates and competed poorly in mixed-culture experiments. This suggested that, although not essential for viability, expression from the psbDI locus is required to produce sufficient D2 and CP43 for optimal growth.