Mutational Analysis of a Wheat O-methyltransferase Involved in Flavonoid Metabolism

Methylated flavones, and tricin in particular, have been implicated in protecting wheat plants against a variety of biotic and abiotic stresses. Methylated flavones are produced via O-methylation of the hydroxyl groups in flavones, which is catalyzed by O-methyltransferases (OMTs). To examine the role of wheat OMT2 in methylated flavone biosynthesis and facilitate interrogation of tricin functions in wheat-environment interactions, loss-of-function mutants of OMT2 homoeologs, omt-A2 and omt-B2, were identified from a tetraploid wheat Targeting Induced Local Lesions in Genomes (TILLING) mutant population and crossed to generate the omt-A2omt-B2 double mutant. Although tricin and most other soluble phenolics did not differ in leaves and glumes of TILLING control and the omt-A2, omt-B2, and omt-A2 omt-B2 mutants, chlorogenic acid was increased in glumes of omt-A2 omt-B2 relative to TILLING control, suggesting that it might serve as a substrate for OMT2. The omt2 mutant lines showed similar growth phenotypes as well as comparable lignin deposition in cell walls of stems compared to TILLING control. These results collectively suggest that OMT2 and its close homolog OMT1 may possess overlapping activities in tricin production, with OMT1 compensating for the missing OMT2 activities in the omt2 mutant lines.

Interaction of two MADS-box genes leads to growth phenotype divergence of all-flesh type of tomatoes

All-flesh tomato cultivars are devoid of locular gel and exhibit enhanced firmness and improved postharvest storage. Here, we show that SlMBP3 is a master regulator of locular tissue in tomato fruit and that a deletion at the gene locus underpins the All-flesh trait. Intriguingly, All-flesh varieties lack the deleterious phenotypes reported previously for SlMBP3 under-expressing lines and which preclude any potential commercial use. We resolve the causal factor for this phenotypic divergence through the discovery of a natural mutation at the SlAGL11 locus, a close homolog of SlMBP3. Misexpressing SlMBP3 impairs locular gel formation through massive transcriptomic reprogramming at initial phases of fruit development. SlMBP3 influences locule gel formation by controlling cell cycle and cell expansion genes, indicating that important components of fruit softening are determined at early pre-ripening stages. Our findings define potential breeding targets for improved texture in tomato and possibly other fleshy fruits.

The maize gene maternal depression of r1 (mdr1) encodes a DNA glycosylase with maternal and paternal fertility functions

Demethylation of transposons can activate expression of nearby genes and cause imprinted gene expression in endosperm, and it is hypothesized to lead to expression of transposon siRNAs that reinforce silencing in the next generation through transfer either into egg or embryo. Here we describe maternal derepression of R1 (mdr1), a DNA glycosylase with homology to Arabidopsis DEMETER that is partially responsible for demethylation of thousands of regions in endosperm. Maternally-expressed imprinted genes were enriched strongly enriched for overlap with demethylated regions, but the majority of genes that overlapped demethylated regions were not imprinted. Demethylated regions were depleted from the majority of repetitive DNA in the genome but enriched in a set of transposon families accounting for about a tenth of the total demethylated regions. Demethylated regions produced few siRNAs and were not associated with excess CHH methylation in endosperm or other tissues. mdr1 and its close homolog dng102 are essential factors in maternal and paternal fertility in maize, as neither double mutant microgametophytes nor megagametophytes gave rise to seeds. These data establish DNA demethylation by glycosylases as essential in maize endosperm and pollen and suggest that neither transposon regulation nor genomic imprinting are its main function.

The DREAM complex represses growth in response to DNA damage in Arabidopsis

The DNA of all organisms is constantly damaged by physiological processes and environmental conditions. Upon persistent damage, plant growth and cell proliferation are reduced. Based on previous findings that RBR1, the only Arabidopsis homolog of the mammalian tumor suppressor gene retinoblastoma, plays a key role in the DNA damage response in plants, we unravel here the network of RBR1 interactors under DNA stress conditions. This led to the identification of homologs of every DREAM component in Arabidopsis, including previously not recognized homologs of LIN52. Interestingly, we also discovered NAC044, a mediator of DNA damage response in plants and close homolog of the major DNA damage regulator SOG1, to directly interact with RBR1 and the DREAM component LIN37B. Consistently, not only mutants in NAC044 but also the double mutant of the two LIN37 homologs and mutants for the DREAM component E2FB showed reduced sensitivities to DNA-damaging conditions. Our work indicates the existence of multiple DREAM complexes that work in conjunction with NAC044 to mediate growth arrest after DNA damage.

Hair (H) interacts with SlZFP8-like to regulate the initiation and elongation of trichomes by modulating SlZFP6 expression in tomato

Trichomes are specialized glandular or non-glandular structures that provide physical or chemical protection against insect and pathogens attack. Trichomes in Arabidopsis, as typical non-glandular structures, have been extensively studied. However, the molecular mechanism underlying glandular trichome formation and elongation still remains largely unknown. We previously demonstrated that Hair (H) is essential for the formation of type I and type VI trichomes. Here, we found that overexpression of H increased the density and length of tomato trichomes. We revealed that H physically interacts with its close homolog SlZFP8-like (SlZFP8L) and SlZFP8L also directly interacts with Woolly (Wo) by biochemical assays. SlZFP8L overexpression plants showed increased trichome density and length. We further found that the expression of SlZFP6, encoding a C2H2 zinc finger protein, is positively regulated by H. We identified that SlZFP6, is a direct target of H through ChIP-qPCR, Y1H, and LUC assays. Similar to H and SlZFP8L, the overexpression of SlZFP6 also increased the density and length of tomato trichomes. Taken together, our results suggest that H interacts with SlZFP8-like to regulate the initiation and elongation of trichomes by modulating SlZFP6 expression in tomato.

The structural mechanism underlying the antithetic effect of homologous RND1 and RhoD GTPases in plexin regulation

Plexins are semaphorin receptors that play essential roles in neuronal axon guidance and in many other important biological processes. Plexin signaling depends on a semaphorin-induced dimerization mechanism, and is modulated by small signaling GTPases of the Rho family, of which RND1 serves as a plexin activator yet its close homolog RhoD an inhibitor. Using molecular dynamics (MD) simulations we showed that RND1 reinforces plexin dimerization interface whereas RhoD destabilizes it due to their differential interaction with cell membrane. Upon binding plexin dimers at the Rho-GTPase binding (RBD) domains, RND1 and RhoD interact differently with the inner leaflet of cell membrane, and exert opposite effects on the dimerization interface via an allosteric network involving the RBD domain, RBD linkers, and a buttress segment adjacent to the dimerization interface. The differential membrane interaction is attributed to the fact that, unlike RND1, RhoD features a short C-terminal tail and a positively-charged membrane interface.

Molecular variation in a functionally divergent homolog of FCA regulates flowering time in Arabidopsis thaliana

The identification and functional characterization of natural variants in plants are essential for understanding phenotypic adaptation. Here we identify a molecular variation in At2g47310 that contributes to the natural variation in flowering time in Arabidopsis thaliana accessions. This gene, which we term SISTER of FCA (SSF), functions in an antagonistic manner to its close homolog FCA. Genome-wide association analysis screens two major haplotypes of SSF associated with the natural variation in FLC expression, and a single polymorphism, SSF-N414D, is identified as a main contributor. The SSF414N protein variant interacts more strongly with CUL1, a component of the E3 ubiquitination complex, than the SSF414D form, mediating differences in SSF protein degradation and FLC expression. FCA and SSF appear to have arisen through gene duplication after dicot-monocot divergence, with the SSF-N414D polymorphism emerging relatively recently within A. thaliana. This work provides a good example for deciphering the functional importance of natural polymorphisms in different organisms.