Nuclear DNA endoreduplication and expression of the mitotic inhibitor Ccs52 associated to determinate and lupinoid nodule organogenesis.

Lotus japonicus determinate nodules differ greatly from indeterminate nodules in their organogenesis and morphological characteristics, whereas Lupinus albus lupinoid nodules share features of determinate and indeterminate nodules. The mitotic inhibitor Ccs52A is essential for endoreduplication and ploidy-dependent cell enlargement during symbiotic cell differentiation in Medicago truncatula indeterminate nodules. ccs52A homolog genes were isolated from lupin and lotus nodules; the deduced Ccs52A proteins showed high sequence similarity with other Cdh-1-type activators of the anaphase-promoting complex and were grouped with A-type Ccs52 proteins from different plants. In lupin, ccs52A expression was restricted to the earlier stages of nodule development, whereas ccs52A transcripts accumulated in lotus nodule primordia and, to a lesser extent, in mature nodules. Nodule development in Lupinus albus involved a progressive increase in nuclear and cellular size and ploidy level; similarly, Lotus japonicus nodules contained polyploid nuclei and enlarged cells in the infected zone. Nevertheless, in situ hybridization experiments showed the highest ccs52A expression in the inner cortex cells of the lupin nodule primordium, probably associated to the increased size of these cells in mature nodules. In view of our results, Ccs52A-mediated endoreduplication appears to be a universal mechanism required for nodule cell differentiation during the establishment of nitrogen-fixing symbioses.