The nucleus-encoded HCF107 gene of Arabidopsis provides a link between intercistronic RNA processing and the accumulation of translation-competent psbH transcripts in chloroplasts

To understand the functional significance of RNA processing for the express ion of plastome-encoded photosynthesis genes, we Investigated the nuclear m utation hcf107 of Arabidopsis. The mutation is represented by two alleles, both of which lead to a defective photosystem II (PSII). In vivo protein la beling, in vitro phosphorylation, and immunoblot experiments revealed that the psbB gene product (CP47) and an 8-kD phosphoprotein, the psbH gene prod uct (PsbH), are absent in mutant plants. PsbH and PsbB are essential requir ements for PSII assembly in photosynthetic eukaryotes, and their absence in hcf107 is consistent with the PSII-less mutant phenotype. RNA gel blot hyb ridizations showed that the hcf107 mutation specifically impairs the accumu lation of some but not all oligocistronic psbH transcripts that are release d from the pentacistronic psbB-psbT-psbH-petB-petD precursor RNA by interge nic endonucleolytic cleavage. In contrast, neither the levels nor the sizes of psbB-containing RNAs are affected. S1 nuclease protection analyses reve aled that psbH RNAs are lacking only where psbH is the leading cistron and that they are processed at position -45 in the 5' leader segment of psbH. T hese data and additional experiments with the cytochrome b(6)f complex muta nt hcf152, which is defective in 3' psbH processing, suggest that only thos e psbH-containing transcripts that are processed at their -45 5' ends can b e translated. Secondary structure analysis of the 5' psbH leader predicted the formation of stable stem loops in the nonprocessed transcripts, which a re unfolded by processing at the -45 site. We propose that this unfolding o f the psbH leader segment as a result of RNA processing is essential for th e translation of the psbH reading frame. We suggest further that HCF107 has dual functions: it is involved in intercistronic processing of the psbH 5' untranslated region or the stabilization of 5' processed psbH RNAs, and co ncomitantly, it is required for the synthesis of CP47.