The Arabidopsis TMS1 encodes a heat shock protein identical to the Hsp40 protein AtERdj3A and plays important roles in the thermotolerance of pollen tubes and other plant tissues. Despite its importance to plant growth and reproduction, little has been known about its mechanisms underlying thermotolerance of plants. In this study, the relationship between TMS1 and the Hsp70 proteins, Binding Immunoglobulin Proteins (BiPs) was explored to understand the molecular mechanisms of TMS1 in thermotolerance of plants. The expression of TMS1 was induced not only by heat shock, but also by dithiothreitol (DTT) and L-azetidine-2-carboxylic acid (AZC), similarly to the three BiP genes, indicating that TMS1 may be involved in unfolded protein response (UPR). The firefly luciferase complementary imaging (LCI), GST pull-down and ATPase enzyme activity assays demonstrated that the DnaJ domain of TMS1 could interact with BiP1 and BiP3, and could stimulate their ATPase enzyme activities. In addition, the expression level of TMS1 was reduced in the bzip28 bzip60 double mutant. These results suggest that TMS1 may function at the downstream of bZIP28 and bZIP60 and be involved in termotolerance of plants, possibly by participating in refolding or degradation of unfolded and misfolded proteins through interaction with the BiPs.

Pollen formation and pollen tube growth are essential for the delivery of male gametes into the female embryo sac for double fertilization. Little is known about the mechanisms that regulate the late developmental process of pollen formation and pollen germination. In this study, we characterized a group of Arabidopsis AGC kinase proteins, NDR2/4/5, involved in pollen development and pollen germination. The NDR2/4/5 genes are mainly expressed in pollen grains at the late developmental stages and in pollen tubes. They function redundantly in pollen formation and pollen germination. At the tricellular stages, the ndr2 ndr4 ndr5 mutant pollen grains exhibit an abnormal accumulation of callose, precocious germination and burst in anthers, leading to a drastic reduction in fertilization and a reduced seed set. NDR2/4/5 proteins can interact with another group of proteins (MOB1A/1B) homologous to the MOB proteins from the Hippo signaling pathway in yeast and animals. The Arabidopsis mob1a mob1b mutant pollen grains also have a phenotype similar to that of ndr2 ndr4 ndr5 pollen grains. These results provide new evidence demonstrating that the Hippo signaling components are conserved in plants and play important roles in sexual plant reproduction.

Restriction of pollen germination before the pollen grain is pollinated to stigma is essential for successful fertilization in angiosperms. However, the mechanisms underlying the process remain poorly understood. Here, we report functional characterization of the MAPKKK kinases, MAP3Kε1 and MAP3Kε2, involve in control of pollen germination in Arabidopsis. The two genes were expressed in different tissues with higher expression levels in the tricellular pollen grains. The map3kε1 map3kε2 double mutation caused abnormal callose accumulation, increasing level of JA and precocious pollen germination, resulting in significantly reduced seed set. Furthermore, the map3kε1 map3kε2 double mutations obviously upregulated the expression levels of genes in JA biosynthesis and signaling. The MAP3Kε1/2 interacted with MOB1A/1B which shared homology with the core components of Hippo singling pathway in yeast. The Arabidopsis mob1a mob1b mutant also exhibited a similar phenotype of precocious pollen germination to that in map3kε1 map3kε2 mutants. Taken together, these results suggested that the MAP3Kεs interacted with MOB1s and played important role in restriction of the precocious pollen germination, possibly through crosstalk with JA signaling and influencing callose accumulation in Arabidopsis.