Expression patterns of MADS-box genes as studied by in situ hybridization
--Jorge Cacharrón, Achim Fischer, Heinz Saedler and Günter Theissen

During recent years, MADS-box genes have been shown to control inflorescence and flower development in dicotyledonous plant species. This let us assume that they may play a similar role in monocots like maize. Therefore, we have taken advantage of cDNA clones to study the expression patterns of several MADS-box genes in maize (Zea mays ssp. mays, T232) and teosinte (Zea mays ssp. parviglumis, Balsas) by in situhybridization experiments. Here the results for three of the MADS-box genes, termed zmm6, zmm7 and zmm8, are reported.

Interestingly, in the case of zmm6, differential expression between maize and teosinte was found. The gene shows the earliest expression, out of the analyzed ones, starting at the stage when spikelet-pair-primordia bifurcate into developing pedicellate and sessile spikelets. At this stage, expression of zmm6 is restricted to one of each spikelet-primordium of a pair. This pattern is visible in male and female inflorescences of maize, while expression of zmm6 in teosinte is only present in the male inflorescence. We could not detect any expression at corresponding developmental stages in female teosinte inflorescences. Further hybridization experiments with female maize inflorescences showed that zmm6 is also expressed in later stages during spikelet development, but then it is present in both the sessile and the pedicellate spikelet in stamen and gynoecium. According to the observed expression pattern one can assume a possible role for zmm6. Though we do not know yet whether initial expression is located in the pedicellate or the sessile spikelet, the expression correlates with the presence of the pedicellate spikelet which is aborted at an early stage during development of the female teosinte inflorescence. Thus zmm6 could have acted, e.g., as a "reactivator" of the pedicellate spikelet during the transition from teosinte to maize. Furthermore, in investigations of Doebley and co-workers (PNAS 87:9888-9892, 1990) mapping with RI lines showed that zmm6 is located on chromosome 1, in a region that was significantly associated with the morphological trait PEDS (pedicellate spikelet score). This supports the functional role of zmm6 mentioned above. Further experiments are now being carried out to test this hypothesis.

In both the female inflorescences of teosinte and maize, zmm8 is expressed in the undifferentiated flower meristem of each spikelet, and during development expression becomes located to the stamen and gynoecium as well as palea and lemma of the upper flower. zmm8 is not expressed in the lower flower of each spikelet. A comparison of the putative protein sequence downstream of the MADS-domain between zmm8 and OsMADS1 of rice (Chung et al., PMB 26:657-665, 1994), showed 76% sequence identity. The distant relationship between maize and rice, together with the high sequence identity between both genes, makes it conceivable that zmm8 is the ortholog of OsMADS1. The expression pattern of zmm8 closely resembles that reported for OsMADS1, with one remarkable difference: we observed expression also in the stamen primordia of the teosinte ear, while a corresponding expression pattern has not been reported for OsMADS1. Maybe this is due to differences between unisexual and perfect flower development, respectively. For example, expression of a zmm8/OsMADS1-like gene could be incompatible with proper stamen formation. Thus, the gene has to be suppressed in the perfect rice flowers, but not in the female teosinte and maize flower, where stamen primordia are aborted. This hypothesis could be tested by studying the expression of zmm8 in the maize tassel.

zmm7, in contrast to the other analyzed MADS-box genes, shows the latest onset of expression. In the female maize inflorescence, expression starts at a stage when nearly all flower organs are differentiated. It is located in the developing gynoecial ridge, and later overall in the developing silk. Further in situ hybridization experiments will show whether expression of zmm7 is absent in the male inflorescence, as one would expect. Such a result would suggest that zmm7 function is restricted to silk development. Chromosomal localization of this gene and analysis of silkless-mutants should also give more insights. 


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