Eleven mutant lines exhibiting decreased numbers of chloroplasts per cell were isolated from 8 800 tagged mutant lines of Physcomitrella patens by microscopic observations. Chloronema subapical cells in wild-type plants had a mean of 48 chloroplasts, whereas chloroplast numbers in subapical cells in mutant lines 215 and 222 decreased to 75 % of that in the wild type. Seven mutant lines - 473, 122, 221, 129, 492, 207, and 138 - had about half as many chloroplasts as the wild type. Mutant line 11 had a few remarkably enlarged chloroplasts, and mutant line 347 had chloroplasts of various sizes. Whereas the cell volume was the same as in the wild type in mutant lines 222, 473, 221, 129, 492, and 207, the cell volume of the other mutants increased. The chloroplast number of leaf cells was the same as that of chloronema cells in each mutant line when gametophores could be formed. Treatment with ampicillin decreased the number of chloroplasts in all mutant lines. Southern hybridization using DNA in tags as probes showed that only one insertion occurred in mutant lines 473 and 221. To determine whether the tagged DNA inserted into the known genes for plastid division, we isolated the PpMinD1, PpMinD2, and PpMinE1 genes. Genomic polymerase chain reaction analysis showed that the PpFtsZ and PpMinD/E genes were not disrupted by the insertion of the tags in mutant lines 11 and 347, respectively.

SummaryWe investigated two genes that encode a homolog of the bacterial cell division site determinant MinD, PpMinD1 and PpMinD2, which were previously identified in the moss Physcomitrella patens. Southern analysis suggested that the P. patens genome does not contain minD genes other than PpMinD1 and PpMinD2. Molecular phylogenetic analysis and the results of a transient expression assay using MinD : green fluorescent protein (GFP) fusions implied that PpMinD1 and PpMinD2 are cyanobacterium-derived, closely related genes encoding chloroplast-targeted MinD proteins. Disruption of the genomic PpMinD1 or PpMinD2 locus had no detectable effects on plastid division. This implies that there is functional redundancy in the two minD genes in P. patens.