Cloning and in vivo analysis of a nuclear-encoded chloroplast-localized 28 kDa RNA-binding protein

A cDNA encoding a chloroplast-localized 28 kDa RNP, At28RNP, was isolated from Arabidopsis. Chloroplast import experiments and RNA UV-crosslinking have confirmed the localization and RNA-binding ability of this protein suggested by its deduced sequence. Expression analysis of At28RNP revealed that the mRNA and the encoded plastid-localized protein accumulate in the active tissues. The At28RNP protein has high sequence identity (60.6%) to a spinach chloroplast protein, So28RNP, which binds to the 3′ ends of plastid RNAs that contain transcript stabilizing inverted repeat sequences (IRs) in their untranslated regions. In vitro RNA-processing assays have shown that the So28RNP is necessary for the proper processing of a precursor RNA to a mature form. The identification and isolation of a homologous protein n Arabidopsis has allowed for the in vivo analysis of plastid localized RNP function using a reverse genetic approach. Transgenic Arabidopsis plants were generated with both increased and decreased levels of At28RNP and while protein levels were dramatically effected in both types of transgenic plants, no significant differences were found between transgenic plants and controls with respect to affects on plastid RNA maturation or accumulation. However, the stable inheritance of altered At28RNP protein expression within the transgenic lines may prove useful in further studies of RNPs in chloroplast gene expression.