Cloning And Functional Analysis Of FtsZ Genes From Nicotiana Tobacum.L And Chlamydomonas Reinhardtii

FtsZ is a conservative and primitive cytoskeleton protein in the cell division of almost all known prokaryotoes. The discovery of FtsZ existed in plants proved us a chance to study the molecular mechanism controling the plastids division. The role of plant FtsZs played in the plastids division had been established preliminarily. However, the exact functional pattern of FtsZs involved in the plastids division and/or plastids shape maintaining is still unknown. In addition, the existence of at least two distincty ftsZ gene families, FtsZ1 and FtsZ2, which encodes different FtsZ proteins, respectively, in the nuclear genomes of higher plants makes the understanding of plant FtsZs function(s) in the plastids division and/or plastids shape maintaining more confusedly.Two cDNAs (NtFtsZ1 and NtFtsZ2), encoding the prokaryotic cell division protein FtsZ homologs, have been isolated from Nicotiana tobaccum by RT-PCR and then RACE method. Protein sequence alignments indicate that the two NtFtsZs are similar to other FtsZ proteins and contain the conserved GTP binding motif. Sequences and phylogenetic analysis with the FtsZ proteins from higher plants suggests that the two NtFtsZs are the members of FtsZ1 family. The results of Southern blotting show that ftsZ have multi-copies in the genomes of tobacco and similar expression patterns of the two genes are also confirmed by Northern blotting. Expression of gfp-tagged NtFtsZs in E.coli indicted that NtFtsZs have similar function with their prokaryotic homologs. Antisense and sense expression constuctions were employed to investigate the functions of the two FtsZ proteins. Although antisense expression of NtFtsZs reduced the native proteins level obviously, however, the size and number of chloroplasts in trangenic tobacco plants had not been affected. In contrast, overexpression of NtFtsZs in transgenic plants strikingly changed the number and morphology of chloroplasts suggested that NtFtsZs might have different functions in plastids division and maintaining of plastid shape. All of these results remind us the possible multi-functions of FtsZ proteins, for example, plastokeleton, in higher plant plastids.We used the degenerated primers PCR to produce a cDNA fragment from total RNA of Chlamydomonas reinhardtii. Based on the fragment sequence, a sets of primers are designed toamplify the 5' and 3' region of cDNA by RACE method. The partial cDNA sequence was designated as CrFtsZ (Chlmyidomonas ceinhardtii FtsZ). Sequence analysis of CrFtsZ showed many conservative features that existed in all FtsZ proteins, but there is a distinct difference existed in the C-terminal extension of CrFtsZ compared with other FtsZ proteins. Molecular phylogenetic analysis suggests that CrFtsZ share the common ancestor with FtsZs from a -proteobacteria, a group eubacteria from which eukaryotic mitochondria evolved. Furthermore, CrFtsZ displayed some limited but typical sequence similarities with dynamin, a protein is thought responsible for the division of eukaryotic mitochondria, in its novel C-terminal extension. Based on these results, we speculate that CrFtsZ may play an intermediate during the evolution from FtsZ to dynamin.