The anterior Hox gene ceh-13 and elt-1/GATA activate the posterior Hox genes nob-1 and php-3 to specify posterior lineages in the C. elegans embryo.

Hox transcription factors play a conserved role in specifying positional identity during animal development, with posterior Hox genes typically repressing the expression of more anterior Hox genes. Here, we dissect the regulation of the posterior Hox genes nob-1 and php-3 in the nematode C. elegans. We show that nob-1 and php-3 are co-expressed in gastrulation-stage embryos in cells that previously expressed the anterior Hox gene ceh-13. This expression is controlled by several partially redundant transcriptional enhancers. These enhancers act in a ceh-13-dependant manner, providing a striking example of an anterior Hox gene positively regulating a posterior Hox gene. Several other regulators also act positively through nob-1/php-3 enhancers, including elt-1/GATA, ceh-20/ceh-40/Pbx, unc-62/Meis, pop-1/TCF, ceh-36/Otx, and unc-30/Pitx. We identified defects in both cell position and cell division patterns in ceh-13 and nob-1;php-3 mutants, suggesting that these factors regulate lineage identity in addition to positional identity. Together, our results highlight the complexity and flexibility of Hox gene regulation and function and the ability of developmental transcription factors to regulate different targets in different stages of development.

Members of the ELMOD protein family specify formation of distinct aperture domains on the Arabidopsis pollen surface.

Pollen apertures, the characteristic gaps in pollen wall exine, have emerged as a model for studying the formation of distinct plasma membrane domains. In each species, aperture number, position, and morphology are typically fixed; across species they vary widely. During pollen development, certain plasma membrane domains attract specific proteins and lipids and become protected from exine deposition, developing into apertures. However, how these aperture domains are selected is unknown. Here, we demonstrate that patterns of aperture domains in Arabidopsis are controlled by the members of the ancient ELMOD protein family, which, although important in animals, has not been studied in plants. We show that two members of this family, MACARON (MCR) and ELMOD_A, act upstream of the previously discovered aperture proteins and that their expression levels influence the number of aperture domains that form on the surface of developing pollen grains. We also show that a third ELMOD family member, ELMOD_E, can interfere with MCR and ELMOD_A activities, changing aperture morphology and producing new aperture patterns. Our findings reveal key players controlling early steps in aperture domain formation, identify residues important for their function, and open new avenues for investigating how diversity of aperture patterns in nature is achieved.

Zooming in on cells reveals patterns on their outer surfaces. These patterns are actually a collection of distinct areas of the cell surface, each containing specific combinations of molecules. The outer layers of pollen grains consist of a cell wall, and a softer cell membrane that sits underneath. As a pollen grain develops, it recruits certain fats and proteins to specific areas of the cell membrane, known as 'aperture domains'. The composition of these domains blocks the cell wall from forming over them, leading to gaps in the wall called 'pollen apertures'. Pollen apertures can open and close, aiding reproduction and protecting pollen grains from dehydration. The number, location, and shape of pollen apertures vary between different plant species, but are consistent within the same species. In the plant species Arabidopsis thaliana, pollen normally develops three long and narrow, equally spaced apertures, but it remains unclear how pollen grains control the number and location of aperture domains. Zhou et al. found that mutations in two closely related A. thaliana proteins ­ ELMOD_A and MCR ­ alter the number and positions of pollen apertures. When A. thaliana plants were genetically modified so that they would produce different levels of ELMOD_A and MCR, Zhou et al. observed that when more of these proteins were present in a pollen grain, more apertures were generated on the pollen surface. This finding suggests that the levels of these proteins must be tightly regulated to control pollen aperture numbers. Further tests revealed that another related protein, called ELMOD_E, also has a role in domain formation. When artificially produced in developing pollen grains, it interfered with the activity of ELMOD_A and MCR, changing pollen aperture shape, number, and location. Zhou et al. identified a group of proteins that help control the formation of domains in the cell membranes of A. thaliana pollen grains. Further research will be required to determine what exactly these proteins do to promote formation of aperture domains and whether similar proteins control domain development in other organisms.

A species-specific functional module controls formation of pollen apertures.

Pollen apertures are an interesting model for the formation of specialized plasma-membrane domains. The plant-specific protein INP1 serves as a key aperture factor in such distantly related species as Arabidopsis, rice and maize. Although INP1 orthologues probably play similar roles throughout flowering plants, they show substantial sequence divergence and often cannot substitute for each other, suggesting that INP1 might require species-specific partners. Here, we present a new aperture factor, INP2, which satisfies the criteria for being a species-specific partner for INP1. Both INP proteins display similar structural features, including the plant-specific DOG1 domain, similar patterns of expression and mutant phenotypes, as well as signs of co-evolution. These proteins interact with each other in a species-specific manner and can restore apertures in a heterologous system when both are expressed but not when expressed individually. Our findings suggest that the INP proteins form a species-specific functional module that underlies formation of pollen apertures.

Changes in morphogen kinetics and pollen grain size are potential mechanisms of aberrant pollen aperture patterning in previously observed and novel mutants of Arabidopsis thaliana.

Pollen provides an excellent system to study pattern formation at the single-cell level. Pollen surface is covered by the pollen wall exine, whose deposition is excluded from certain surface areas, the apertures, which vary between the species in their numbers, positions, and morphology. What determines aperture patterns is not understood. Arabidopsis thaliana normally develops three apertures, equally spaced along the pollen equator. However, Arabidopsis mutants whose pollen has higher ploidy and larger volume develop four or more apertures. To explore possible mechanisms responsible for aperture patterning, we developed a mathematical model based on the Gierer-Meinhardt system of equations. This model was able to recapitulate aperture patterns observed in the wild-type and higher-ploidy pollen. We then used this model to further explore geometric and kinetic factors that may influence aperture patterns and found that pollen size, as well as certain kinetic parameters, like diffusion and decay of morphogens, could play a role in formation of aperture patterns. In conjunction with mathematical modeling, we also performed a forward genetic screen in Arabidopsis and discovered two mutants with aperture patterns that had not been previously observed in this species but were predicted by our model. The macaron mutant develops a single ring-like aperture, matching the unusual ring-like pattern produced by the model. The doughnut mutant forms two pore-like apertures at the poles of the pollen grain. Further tests on these novel mutants, motivated by the modeling results, suggested the existence of an area of inhibition around apertures that prevents formation of additional apertures in their vicinity. This work demonstrates the ability of the theoretical model to help focus experimental efforts and to provide fundamental insights into an important biological process.