PALE CRESS binds to plastid RNAs and facilitates the biogenesis of the 50S ribosomal subunit

The plant-specific PALE CRESS (PAC) protein has previously been shown to be essential for photoautotrophic growth. Here we further investigated the molecular function of the PAC protein. PAC localizes to plastid nucleoids and forms large proteinaceous and RNA-containing megadalton complexes. It co-immunoprecipitates with a specific subset of chloroplast RNAs including psbK-psbI, ndhF, ndhD, and 23S ribosomal RNA (rRNA), as demonstrated by RNA immunoprecipitation in combination with high throughput RNA sequencing (RIP-seq) analyses. Furthermore, it co-migrates with premature 50S ribosomal particles and specifically binds to 23S rRNA invitro. This coincides with severely reduced levels of 23S rRNA in pac leading to translational deficiencies and related alterations of plastid transcript patterns and abundance similar to plants treated with the translation inhibitor lincomycin. Thus, we conclude that deficiency in plastid ribosomes accounts for the pac phenotype. Moreover, the absence or reduction of PAC levels in the corresponding mutants induces structural changes of the 23S rRNA, as demonstrated by invivo RNA structure probing. Our results indicate that PAC binds to the 23S rRNA to promote the biogenesis of the 50S subunit. Significance Statement PAC is an RNA-binding protein localized in plastid nucleoids that is associated invivo with the 23S rRNA as well as with psbK-psbI, ndhF, and ndhD mRNAs, and forms high molecular weight complexes. The pronounced impact of PAC on the biogenesis of plastid ribosomes severely influences translation, leaf coloration, plant growth and leads to seedling lethality.