Nodal signaling: developmental roles and regulation

Nodal-related ligands of the transforming growth factor-beta (TGFbeta) superfamily play central roles in patterning the early embryo during the induction of mesoderm and endoderm and the specification of left-right asymmetry. Additional roles for this pathway in the maintenance of embryonic stem cell pluripotency and in carcinogenesis have been uncovered more recently. Consistent with its crucial developmental functions, Nodal signaling is tightly regulated by diverse mechanisms including the control of ligand processing, utilization of co-receptors, expression of soluble antagonists, as well as positive- and negative-feedback activities.     

The gibberellin biosynthetic genes AtKAO1 and AtKAO2 have overlapping roles throughout Arabidopsis development

Ent‐kaurenoic acid oxidase (KAO), a class of cytochrome P450 monooxygenases of the subfamily CYP88A, catalyzes the conversion of ent‐kaurenoic acid (KA) to gibberellin (GA) GA₁₂, the precursor of all GAs, thereby playing an important role in determining GA concentration in plants. Past work has demonstrated the importance of KAO activity for growth in various plant species. In Arabidopsis, this enzyme is encoded by two genes designated KAO1 and KAO2. In this study, we used various approaches to determine the physiological roles of KAO1 and KAO2 throughout plant development. Analysis of gene expression pattern reveals that both genes are mainly expressed in germinating seeds and young developing organs, thus suggesting functional redundancy. Consistent with this, kao1 and kao2 single mutants are indistinguishable from wild‐type plants. By contrast, the kao1 kao2 double mutant exhibits typical non‐germinating GA‐dwarf phenotypes, similar to those observed in the severely GA‐deficient ga1‐3 mutant. Phenotypic characterization and quantitative analysis of endogenous GA contents of single and double kao mutants further confirm an overlapping role of KAO1 and KAO2 throughout Arabidopsis development.     

Drosophila APC2 and APC1 play overlapping roles in wingless signaling in the embryo and imaginal discs

The regulation of signal transduction plays a key role in cell fate choices, and its disregulation contributes to oncogenesis. This duality is exemplified by the tumor suppressor APC. Originally identified for its role in colon tumors, APC family members were subsequently shown to negatively regulate Wnt signaling in both development and disease. The analysis of the normal roles of APC proteins is complicated by the presence of two APC family members in flies and mice. Previous work demonstrated that, in some tissues, single mutations in each gene have no effect, raising the question of whether there is functional overlap between the two APCs or whether APC-independent mechanisms of Wnt regulation exist. We addressed this by eliminating the function of both Drosophila APC genes simultaneously. We find that APC1 and APC2 play overlapping roles in regulating Wingless signaling in the embryonic epidermis and the imaginal discs. Surprisingly, APC1 function in embryos occurs at levels of expression nearly too low to detect. Further, the overlapping functions exist despite striking differences in the intracellular localization of the two APC family members.     

Drosophila APC2 and APC1 have overlapping roles in the larval brain despite their distinct intracellular localizations

The tumor suppressor APC and its homologs, first identified for a role in colon cancer, negatively regulate Wnt signaling in both oncogenesis and normal development, and play Wnt-independent roles in cytoskeletal regulation. Both Drosophila and mammals have two APC family members. We further explored the functions of the Drosophila APCs using the larval brain as a model. We found that both proteins are expressed in the brain. APC2 has a highly dynamic, asymmetric localization through the larval neuroblast cell cycle relative to known mediators of embryonic neuroblast asymmetric divisions. Adherens junction proteins also are asymmetrically localized in neuroblasts. In addition they accumulate with APC2 and APC1 in nerves formed by axons of the progeny of each neuroblast-ganglion mother cell cluster. APC2 and APC1 localize to very different places when expressed in the larval brain: APC2 localizes to the cell cortex and APC1 to centrosomes and microtubules. Despite this, they play redundant roles in the brain; while each single mutant is normal, the zygotic double mutant has severely reduced numbers of larval neuroblasts. Our experiments suggest that this does not result from misregulation of Wg signaling, and thus may involve the cytoskeletal or adhesive roles of APC proteins.     

Distinct Wnt signaling pathways have opposing roles in appendage regeneration

In contrast to mammals, lower vertebrates have a remarkable capacity to regenerate complex structures damaged by injury or disease. This process, termed epimorphic regeneration, involves progenitor cells created through the reprogramming of differentiated cells or through the activation of resident stem cells. Wnt/beta-catenin signaling regulates progenitor cell fate and proliferation during embryonic development and stem cell function in adults, but its functional involvement in epimorphic regeneration has not been addressed. Using transgenic fish lines, we show that Wnt/beta-catenin signaling is activated in the regenerating zebrafish tail fin and is required for formation and subsequent proliferation of the progenitor cells of the blastema. Wnt/beta-catenin signaling appears to act upstream of FGF signaling, which has recently been found to be essential for fin regeneration. Intriguingly, increased Wnt/beta-catenin signaling is sufficient to augment regeneration, as tail fins regenerate faster in fish heterozygous for a loss-of-function mutation in axin1, a negative regulator of the pathway. Likewise, activation of Wnt/beta-catenin signaling by overexpression of wnt8 increases proliferation of progenitor cells in the regenerating fin. By contrast, overexpression of wnt5b (pipetail) reduces expression of Wnt/beta-catenin target genes, impairs proliferation of progenitors and inhibits fin regeneration. Importantly, fin regeneration is accelerated in wnt5b mutant fish. These data suggest that Wnt/beta-catenin signaling promotes regeneration, whereas a distinct pathway activated by wnt5b acts in a negative-feedback loop to limit regeneration.     

CGR2 and CGR3 have critical overlapping roles in pectin methylesterification and plant growth in Arabidopsis thaliana

Pectins are critical polysaccharides of the cell wall that are involved in key aspects of a plant's life, including cell‐wall stiffness, cell‐to‐cell adhesion, and mechanical strength. Pectins undergo methylesterification, which affects their cellular roles. Pectin methyltransferases are believed to methylesterify pectins in the Golgi, but little is known about their identity. To date, there is only circumstantial evidence to support a role for QUASIMODO2 (QUA2)‐like proteins and an unrelated plant‐specific protein, cotton Golgi‐related 3 (CGR3), in pectin methylesterification. To add to the knowledge of pectin biosynthesis, here we characterized a close homolog of CGR3, named CGR2, and evaluated the effect of loss‐of‐function mutants and over‐expression lines of CGR2 and CGR3 in planta. Our results show that, similar to CGR3, CGR2 is a Golgi protein whose enzyme active site is located in the Golgi lumen where pectin methylesterification occurs. Through phenotypical analyses, we also established that simultaneous loss of CGR2 and CGR3 causes severe defects in plant growth and development, supporting critical but overlapping functional roles of these proteins. Qualitative and quantitative cell‐wall analytical assays of the double knockout mutant demonstrated reduced levels of pectin methylesterification, coupled with decreased microsomal pectin methyltransferase activity. Conversely, CGR2 and CGR3 over‐expression lines have markedly opposite phenotypes to the double knockout mutant, with increased cell‐wall methylesterification levels and microsomal pectin methyltransferase activity. Based on these findings, we propose that CGR2 and CGR3 are critical proteins in plant growth and development that act redundantly in pectin methylesterification in the Golgi apparatus.     

Class III homeodomain-leucine zipper gene family members have overlapping, antagonistic, and distinct roles in Arabidopsis development

The Arabidopsis thaliana genome contains five class III homeodomain-leucine zipper genes. We have isolated loss-of-function alleles for each family member for use in genetic analysis. This gene family regulates apical embryo patterning, embryonic shoot meristem formation, organ polarity, vascular development, and meristem function. Genetic analyses revealed a complex pattern of overlapping functions, some of which are not readily inferred by phylogenetic relationships or by gene expression patterns. The PHABULOSA and PHAVOLUTA genes perform overlapping functions with REVOLUTA, whereas the PHABULOSA, PHAVOLUTA, and CORONA/ATHB15 genes perform overlapping functions distinct from REVOLUTA. Furthermore, ATHB8 and CORONA encode functions that are both antagonistic to those of REVOLUTA within certain tissues and overlapping with REVOLUTA in other tissues. Differences in expression patterns explain some of these genetic interactions, whereas other interactions are likely attributable to differences in protein function as indicated by cross-complementation studies.