Genome-wide identification, interaction of the MADS-box proteins in Zanthoxylum armatum and functional characterization of ZaMADS80 in floral development

Abstract

As a typical dioecious species, <i>Zanthoxylum armatum</i> establishes apomictic reproduction, hence only female trees are cultivated. However, male and hermaphrodite flowers have recently appeared in female plants, resulting in a dramatic yield reduction. To date, the genetic basis underlying sex determination and apomixis in <i>Z. armatum</i> has been largely unknown. Here, we observed abortion of the stamen or carpel prior to primordium initiation, thus corroborating the potential regulation of MADS-box in sex determination. In <i>Z. armatum</i>, a total of 105 MADS-box genes were identified, harboring 86 MIKC-type MADSs with lack of FLC orthologues. Transcriptome analysis revealed candidate <i>MADSs</i> involved in floral organ identity, including ten male-biased <i>MADSs</i>, represented by <i>ZaMADS92/81/75</i>(AP3/PI-like), and twenty-six female-specified, represented by <i>ZaMADS80/49</i> (STK/AGL11-like) and <i>ZaMADS42</i> (AG-like). Overexpressing <i>ZaMADS92</i> resulted in earlier flowering, while <i>ZaMADS80</i> overexpression triggered precocious fruit set and parthenocarpy as well as dramatic modifications in floral organs. To characterize their regulatory mechanisms, a comprehensive protein-protein interaction network of the represented MADSs was constructed based on yeast two-hybrid and bimolecular fluorescence complementation assays. Compared with model plants, the protein interaction patterns in <i>Z. armatum</i> exhibited both conservation and divergence. ZaMADS70 (SEP3-like) interacted with ZaMADS42 and ZaMADS48 (AP3-like) but not ZaMADS40 (AP1-like), facilitating the loss of petals in <i>Z. armatum</i>. The ZaMADS92/ZaMADS40 heterodimer could be responsible for accelerating flowering in <i>ZaMADS92</i>-OX lines. Moreover, the interactions between ZaMADS80 and ZaMADS67(AGL32-like) might contribute to apomixis. This work provides new insight into the molecular mechanisms of MADS-boxes in sex organ identity in <i>Z. armatum</i>.