DOWN-REGULATION OF EXPRESSION OF THE RHIZOBIUM-LEGUMINOSARUM OUTER-MEMBRANE PROTEIN GENE ROPA OCCURS ABRUPTLY IN INTERZONE II-III OF PEA NODULES AND CAN BE UNCOUPLED FROM NIF GENE ACTIVATION

The expression of the Rhizobium leguminosarum bv. viciae outer membrane protein gene ropA during nodule development was studied using immune-electron microscopy and in situ hybridization. Using immunochemical detection in isolated cell envelopes it had been shown earlier that the RopA outer membrane antigen disappears during bacteroid development (R. A. de Maagd, R. de Rijk, I H. M. Mulders, and B. J. J. Lugtenberg, J. Bacteriol. 171:1136-1142, 1989). In the present study,ve used immune-electron microscopy on vetch nodule sections to show that the decrease in RopA protein expression occurs in the nodule after release of the bacteria from the infection thread, during the transition in morphology from that of the free-living bacterium into that of the bacteroid. Detection of ropA mRNA in sections of pea nodules by in situ hybridization revealed a sudden decrease in messenger level at the transition from prefixation zone TI to interzone II-III. This decrease coincided with a sudden increase in nifH mRNA levels. Although the decrease in ropA messenger and appearance of nif messenger are spatially correlated we could show that ropA down-regulation can be uncoupled from nif gene activation by using a strain that induces non-nitrogen fixing nodules on pea but does develop into bacteroids. The importance of the transition of prefixation zone II to interzone II-LII as a developmental snitch for bacteroid development is discussed.