Endoreduplication Mediated by the Anaphase-Promoting Complex Activator CCS52A Is Required for Symbiotic Cell Differentiation in Medicago truncatula Nodules

Abstract

In Medicago nodules, endoreduplication cycles and ploidy-dependent cell enlargement occur during the differentiation of bacteroid-containing nitrogen-fixing symbiotic cells. These events are accompanied by the expression of ccs52A , a plant ortholog of the yeast and animal cdh1/srw1/fzr genes, acting as a substrate-specific activator of the anaphase-promoting complex (APC) ubiquitin ligase. Because CCS52A is involved in the transition of mitotic cycles to endoreduplication cycles, we investigated the importance of somatic endoploidy and the role of the M. truncatula ccs52A gene in symbiotic cell differentiation. Transcription analysis and ccs52A promoter-driven ␤ -glucuronidase activity in transgenic plants showed that ccs52A was dispensable for the mitotic cycles and nodule primordium formation, whereas it was induced before nodule differentiation. The CCS52A protein was present in the nucleus of endoreduplication-competent cells, indicating that it may activate APC constitutively during the endoreduplication cycles. Downregulation of ccs52A in transgenic M. truncatula plants drastically affected nodule development, resulting in lower ploidy, reduced cell size, inefficient invasion, and the maturation of symbiotic cells, accompanied by early senescence and finally the death of both the bacterium and plant cells. Thus, ccs52A expression is essential for the formation of large highly polyploid symbiotic cells, and endoreduplication is an integral part of normal nodule development.

Eukaryotic control on bacterial cell cycle and differentiation in the Rhizobium –legume symbiosis

“…Few other examples in the bacterial world describe polyploid cells, but they all point to a similar correlation (18)(19)(20). Furthermore, it is remarkable that prokaryotic bacteroids and their eukaryotic host cells in the infection zone of indeterminate nodules follow similar differentiation paths proceeding synchronously and involving genome multiplication and cell enlargement (7,10).…”

“…This cell enlargement is predominantly responsible for the growth of the nodule organ and is mediated by repeated endoreduplication cycles resulting in 64C or 128C polyploid nodule cells (C being the haploid DNA content) (3)(4)(5)(6)(7)(8). The endoreduplication cycle is a modified cell cycle with replication of the genome (S phase) but without mitosis and cytokinesis (M phase).…”

“…Moreover, in many cell types endoreduplication is tightly linked to cell differentiation, and inhibition of endoreduplication results in developmental abnormalities. Reducing endoreduplication in nodule cells of transgenic Medicago truncatula plants by downregulation of the ccs52A gene, a key regulator of the endoreduplication cycles, aborted nodule differentiation (7). The absence of nitrogen-fixing symbiotic cells in these nodules proved that endoreduplication is an integral part of the symbiotic cell differentiation.…”

Seven in Absentia Proteins Affect Plant Growth and Nodulation inMedicago truncatula

“…Until now, only two E3 ligases have been shown to play a role in nodule development (Vinardell et al, 2003;Shimomura et al, 2006). The anaphase-promoting complex, APC CCS52A , degrades mitotic cyclins in the infected cells to induce endoreduplication, resulting in polyploidy needed to assure cell enlargement to host the huge numbers of nitrogenfixing bacteroids (Vinardell et al, 2003;Kondorosi et al, 2005).…”

How rhizobial symbionts invade plants: the Sinorhizobium–Medicago model

“…These layers of invasion-competent cells have exited mitosis and become polyploid owing to cycles of genomic endoreduplication without cytokinesis 14,66 . Plant cell endoreduplication is required for the formation of functional nodules, and is dependent on the degradation of mitotic cyclins by anaphase-promoting complex (APC), an E3 ubiquitin ligase 66,67 . The advantages of cellular polyploidy are a higher transcription rate and a higher metabolic rate than cells with the normal 2n DNA content 68 -characteristics that might be important for the invaded plant cells to support bacterial nitrogen fixation (discussed later).…”