MULTIFUNCTIONAL ACETYL-COA CARBOXYLASE FROM BRASSICA-NAPUS IS ENCODEDBY A MULTIGENE FAMILY - INDICATION FOR PLASTIDIC LOCALIZATION OF AT LEAST ONE ISOFORM

Three genes coding for different multifunctional acetyl-CoA carboxylas e (ACCase; EC 6.4.1.2) isoenzymes from Brassica napus were isolated an d divided into two major classes according to structural features in t heir 5' regions: class I comprises two genes with an additional coding exon of approximately 300 bp at the 5' end, and class II is represent ed by one gene carrying an intron of 586 bp in its 5' untranslated reg ion. Fusion of the peptide sequence encoded by the additional first ex on of a class I ACCase gene to the jellyfish Aequorea victoria green f luorescent protein (GFP) and transient expression in tobacco protoplas ts targeted GFP to the chloroplasts, In contrast to the deduced primar y structure of the biotin carboxylase domain encoded by the class I ge ne, the corresponding amino acid sequence of the class II ACCase shows higher identity with that of the Arabidopsis ACCase, both lacking a t ransit peptide, The Arabidopsis ACCase has been proposed to be a cytos olic isoenzyme, These observations indicate that the two classes of AC Case genes encode plastidic and cytosolic isoforms of multi-functional , eukaryotic type, respectively, and that B. napus contains at least o ne multi-functional ACCase besides the multi-subunit, prokaryotic type located in plastids, Southern blot analysis of genomic DNA from B. na pus, Brassica mpa, and Brassica oleracea, the ancestors of amphidiploi d rapeseed, using a fragment of a multi-functional ACCase gene as a pr obe revealed that ACCase is encoded by a multi-gene family of at least five members.