A PUTATIVE MG CHELATASE SUBUNIT FROM ARABIDOPSIS-THALIANA CV C24 - SEQUENCE AND TRANSCRIPT ANALYSIS OF THE GENE, IMPORT OF THE PROTEIN INTOCHLOROPLASTS, AND IN-SITU LOCALIZATION OF THE TRANSCRIPT AND PROTEIN

We have isolated and sequenced a cDNA from Arabidopsis thaliana cv C24 that encodes a putative Mg chelatase subunit. The deduced amino acid sequence shows a very high level of identity to a gene previously char acterized from Antirrhinum majus (olive) and also high similarity to b chH, a bacterial gene involved in the Mg chelatase reaction of bacteri ochlorophyll biosynthesis. We suggest that this gene be called CHL H. Northern blot analyses were used to investigate the expression of CHL H, another putative Mg chelatase gene, ch-42, and ferrochelatase. The CHL H transcript was observed to undergo a dramatic diurnal variation, rising almost to its maximum level by the end of the dark period, the n increasing slightly at the onset of the light and declining steadily to a minimum by the end of the light period; in contrast, transcripts for ch-42 and ferrochelatase remained constant. A model is proposed i n which the CHL H protein plays a role in regulating the levels of chl orophyll during this cycle. In situ hybridization revealed that the tr anscripts are located over the surface of the chloroplasts, a feature in common with transcripts for the ch-42 gene. The CHL H protein was i mported into the stromal compartment of the chloroplast and processed in an in vitro assay. Immunoblotting showed that the distribution of C HL H protein between the stroma and chloroplast membranes varies depen ding on the concentration of Mg2+. In situ immunofluorescence was used to establish that the CHL H and CH-42 proteins are localized within t he chloroplast in vivo.