Differential regulation of genes transcribed by nucleus-encoded plastid RNA polymerase, and DNA amplification, within ribosome-deficient plastids in stable phenocopies of cereal albino mutants

Abstract

We isolated stable albino plants of barley and maize by inhibiting plastid protein synthesis with streptomycin and propagating bleached seedlings in the absence of antibiotics in vitro. Albino plants are deficient in plastid translation products and plastid ribosomal RNAs, and are stable phenocopies of the barley albostrians and maize iojap mutants, which contain ribosome-free plastids. Once plastid ribosomes are lost they cannot be re-synthesized because about one-third of plastid ribosomal proteins are themselves plastid encoded. The group II/subgroup IIA intron in plastid rpl2 transcripts was not processed in albinos, providing strong evidence for the absence of plastid translation. Photosynthesis-related plastid mRNAs and plastid tRNAs were down-regulated in albino leaves. A differential influence of plastid ribosome deficiency on mRNA levels allowed us to divide genes transcribed by nucleus-encoded plastid RNA polymerase into two groups. Northern analysis revealed increases in the levels of clpP, rpl2, rpl23, rps15 and rpoB mRNAs in total RNA from albino leaves relative to those in green leaves. In contrast, albinism did not increase the band intensities of rps2 and rps4 messages. Plastid ribosome-associated factor(s) or plastid-encoded product(s) play a role in the initiation, termination, processing or stability of transcripts containing trnG(UCC) and rps4. Excision and 100-fold amplification of a 5.2-kb region of plastid DNA encompassing the trnG(UCC) and trnE(UUC) genes was observed in one of four albino barley plants. Gene amplification was correlated with the accumulation of abundant novel transcripts derived from regions flanking the trnG(UCC) gene.