Molecular analyses of Physcomitrella patens MIKCc-type MADS-box genes establish an evolutionary context in plants

MADS-box genes encode a family of highly conserved transcription factors involved in numerous developmental processes in higher eukaryotes. While the members of this transcription factor family have a mostly tissue-specific expression pattern in seed plants, MADS-box genes in non-seed plants show broad expression domains often comprising the gametophytic as well as the sporophytic phase of the plant life cycle. Their functions in non-seed plants remain elusive, although a number of genes could be isolated and characterized from lycophytes, pteridophytes, bryophytes and green algae. To investigate MADS-box genes in a representative of early land plants, the moss Physcomitrella patens was selected. PPMC1 and PPMC2 are two out of six members of the small Physcomitrella patens gene family of MIKCc-type MADS-box genes. While their transcripts are detected equally during all stages of the P. patens life cycle, translational reporter gene fusions reveal distinct expression patterns in both generations of the moss life cycle. In addition to a weak ubiquitous signal, PPMC2 shows strong expression in both male and female gametangia of the gametophyte as well as in the haustorium of the diploid sporophyte. As a major suspect of regulation, the unusually long 5� UTR with features known to be involved in translational activation or repression was investigated. However, replacement of the PPMC2 5� UTR by the 5� UTR of the Antirrhinum gene DEFICIENS in a promoter::GUS fusion disproved regulation on the translational level and, instead, confirmed transcriptional regulation. The translational reporter gene fusion of PPMC1 provides evidence that the protein is first expressed in the apices of gametophores carrying mature gametangia. Later, developing sporophytes show broad protein distribution that is gradually restricted to the upper half of the short seta in older stages and finally disappears. Several gene disruption lines have been generated to help reveal the functions of PPMC1 and PPMC2, but so far no obvious phenotype has been detected. Furthermore, a translational reporter gene fusion of PPMC2 in a PPMC1 gene disruption background convincingly supports that the genes are non-redundant. Based on these findings, putative functions for PPMC1 and PPMC2 in the moss are discussed. The genes likely participate in UV-light protection or sink tissue definition in both generations of the moss life cycle, however, gene functions may have diverged between the haploid and the diploid generation. These findings strongly support the hypothesis of MADS-box gene recruitment from the gametophyte into the sporophyte during the evolution of land plants