C. elegans ksr-1 and ksr-2 have both unique and redundant functions and are required for MPK-1 ERK phosphorylation

Kinase Suppressor of Ras (KSR) is a conserved protein that positively regulates Ras signaling and may function as a scaffold for Raf, MEK, and ERK. However, the precise role of KSR is not well understood, and some observations have suggested that KSR might act in a parallel pathway. In C. elegans, ksr-1 is only required for a specific Ras-mediated process (sex myoblast migration) and is a nonessential positive regulator of other Ras-mediated developmental events. We report the existence of a second C. elegans ksr gene, ksr-2, which is required for Ras-mediated signaling during germline meiotic progression and functions redundantly with ksr-1 during development of the excretory system, hermaphrodite vulva, and male spicules. Thus, while the ksr-1 and ksr-2 genes are individually required only for specific Ras-dependent processes, together these two genes appear necessary for most aspects of Ras-mediated signaling in C. elegans. The finding that ksr-2; ksr-1 double mutants have strong ras-like phenotypes and severely reduced or absent levels of diphosphorylated MPK-1 ERK strongly supports models where KSR acts to promote the activation or maintenance of the Raf/MEK/ERK kinase cascade.     

AXL and AXR1 have redundant functions in RUB conjugation and growth and development in Arabidopsis

Cullin-RING ubiquitin-protein ligases such as the Skp1, cullin, F-box protein (SCF) have been implicated in many growth and developmental processes in plants. Normal SCF function requires that the CUL1 subunit be post-translationally modified by related to ubiquitin (RUB), a protein related to ubiquitin. This process is mediated by two enzymes: the RUB-activating and RUB-conjugating enzymes. In Arabidopsis, the RUB-activating enzyme is a heterodimer consisting of AXR1 and ECR1. Mutations in the AXR1 gene result in a pleiotropic phenotype that includes resistance to the plant hormone auxin. Here we report that the AXL (AXR1-like) gene also functions in the RUB conjugation pathway. Overexpression of AXL in the axr1-3 background complements the axr1-3 phenotype. Biochemical analysis indicates that AXL overexpression restores CUL1 modification to the wild-type level, indicating that AXR1 and AXL have the same biochemical activity. Although the axl mutant resembles wild-type plants, the majority of axr1 axl-1 double mutants are embryo or seedling lethal. Furthermore, the axl-1 mutation reveals novel RUB-dependent processes in embryo development. We conclude that AXR1 and AXL function redundantly in the RUB conjugating pathway.     

Pkd1 is required for male reproductive tract development

Reproductive tract abnormalities and male infertility have higher incidence in ADPKD patients than in general populations. In this work, we reveal that Pkd1, whose mutations account for 85% of ADPKD cases, is essential for male reproductive tract development. Disruption of Pkd1 caused multiple organ defects in the murine male reproductive tract. The earliest visible defect in the Pkd1^?/? reproductive tract was cystic dilation of the efferent ducts, which are derivatives of the mesonephric tubules. Epididymis development was delayed or arrested in the Pkd1^?/? mice. No sign of epithelial coiling was seen in the null mutants. Disruption of Pkd1 in epithelium alone using the Pax2-cre mice was sufficient to cause efferent duct dilation and coiling defect in the epididymis, suggesting that Pkd1 is critical for epithelium development and maintenance in male reproductive tract. In-depth analysis showed that Pkd1 is required to maintain tubulin cytoskeleton and important for Tgf-β/Bmp signal transduction in epithelium of male reproductive tract. Altogether, our results for the first time provide direct evidence for developmental roles of Pkd1 in the male reproductive tract and provide new insights in reproductive tract abnormalities and infertility in ADPKD patients.     

Conifer reproductive development involves B-type MADS-box genes with distinct and different activities in male organ primordia

The Norway spruce MADS-box genes DAL11, DAL12 and DAL13 are phylogenetically related to the angiosperm B-function MADS-box genes: genes that act together with A-function genes in specifying petal identity and with C-function genes in specifying stamen identity to floral organs. In this report we present evidence to suggest that the B-gene function in the specification of identity of the pollen-bearing organs has been conserved between conifers and angiosperms. Expression of DAL11 or DAL12 in transgenic Arabidopsis causes phenotypic changes which partly resemble those caused by ectopic expression of the endogenous B-genes. In similar experiments, flowers of Arabidopsis plants expressing DAL13 showed a different homeotic change in that they formed ectopic anthers in whorls one, two or four. We also demonstrate the capacity of the spruce gene products to form homodimers, and that DAL11 and DAL13 may form heterodimers with each other and with the Arabidopsis B-protein AP3, but not with PI, the second B-gene product in Arabidopsis. In situ hybridization experiments show that the conifer B-like genes are expressed specifically in developing pollen cones, but differ in both temporal and spatial distribution patterns. These results suggest that the B-function in conifers is dual and is separated into a meristem identity and an organ identity function, the latter function possibly being independent of an interaction with the C-function. Thus, even though an ancestral B-function may have acted in combination with C to specify micro- and megasporangia, the B-function has evolved differently in conifers and angiosperms.     

The Aspergillus nidulans genes chsA and chsD encode chitin synthases which have redundant functions in conidia formation

 We previously isolated three chitin synthase genes (chsA, chsB, and chsC) from Aspergillus nidulans. In the present work, we describe the isolation and characterization of another chitin synthase gene, named chsD, from A. nidulans. Its deduced amino acid sequence shows 56.7% and 55.9% amino acid identity, respectively, with Cal1 of Saccharomyces cerevisiae and Chs3 of Candida albicans. Disruption of chsD caused no defect in cell growth or morphology during the asexual cycle and caused no decrease in chitin content in hyphae. However, double disruption of chsA and chsD caused a remarkable decrease in the efficiency of conidia formation, while double disruption of chsC and chsD caused no defect. Thus it appears that chsA and chsD serve redundant functions in conidia formation.     

The Aspergillus nidulans genes chsA and chsD encode chitin synthases which have redundant functions in conidia formation

We previously isolated three chitin synthase genes (chsA, chsB, and chsC) from Aspergillus nidulans. In the present work, we describe the isolation and characterization of another chitin synthase gene, named chsD, from A. nidulans. Its deduced amino acid sequence shows 56.7% and 55.9% amino acid identity, respectively, with Cal1 of Saccharomyces cerevisiae and Chs3 of Candida albicans. Disruption of chsD caused no defect in cell growth or morphology during the asexual cycle and caused no decrease in chitin content in hyphae. However, double disruption of chsA and chsD caused a remarkable decrease in the efficiency of conidia formation, while double disruption of chsC and chsD caused no defect. Thus it appears that chsA and chsD serve redundant functions in conidia formation.     

Arabidopsis MSI1 is required for epigenetic maintenance of reproductive development

WD40 repeat proteins similar to yeast MSI1 are conserved in animals and plants, in which they participate in complexes involved in chromatin metabolism. Although MSI1-like proteins are well characterised biochemically, their function in the development of multicellular eukaryotes is not well understood. We constructed Arabidopsis plants in which the AtMSI1 protein level was altered. Strong ectopic expression of AtMSI1 produced no visible altered phenotype, but reduction of AtMSI1 dramatically affected development. The primary shoot apical meristem was unable to develop organs after the transition to flowering. Flowers that developed on floral shoots from axillary meristems experienced a progressive loss of floral morphology, including a reduction in size of the petals and stamens and the development of carpel-like sepals. Ovule development was disrupted in all flowers, resulting in complete female sterility. Molecular analysis of the mutant plants revealed that AtMSI1 is required to maintain the correct temporal and organ-specific expression of homeotic genes, including AGAMOUS and APETALA2. In contrast, FAS1 and FAS2, which together with AtMSI1 form the chromatin assembly complex CAF-1, are not required for repression of these genes. Therefore, AtMSI1 has specific functions in addition to CAF-1-mediated chromatin assembly. Efficient formation of heterochromatin, but not methylation of centromeric DNA repeats, depends on AtMSI1 presence demonstrating a key role of AtMSI1 in maintenance of chromatin structure.     

Multiple and integrated functions of floral C-class MADS-box genes in flower and fruit development of Physalis floridana

Plant Molecular Biology - This work reveals potentially multiple and integrated roles in flower and fruit development of floral C-class MADS-box genes in Physalis. The Physalis fruit features a...     

SPATULA and ALCATRAZ, are partially redundant, functionally diverging bHLH genes required for Arabidopsis gynoecium and fruit development

The Arabidopsis gynoecium is a complex organ that facilitates fertilization, later developing into a dehiscent silique that protects seeds until their dispersal. Identifying genes important for development is often hampered by functional redundancy. We report unequal redundancy between two closely related genes, SPATULA (SPT) and ALCATRAZ (ALC), revealing previously unknown developmental roles for each. SPT is known to support septum, style and stigma development in the flower, whereas ALC is involved in dehiscence zone development in the fruit. ALC diverged from a SPT-like ancestor following gene duplication coinciding with the At-β polyploidy event. Here we show that ALC is also involved in early gynoecium development, and SPT in later valve margin generation in the silique. Evidence includes the increased severity of early gynoecium disruption, and of later valve margin defects, in spt-alc double mutants. In addition, a repressive version of SPT (35S:SPT-SRDX) disrupts both structures. Consistent with redundancy, ALC and SPT expression patterns overlap in these tissues, and the ALC promoter carries two atypical E-box elements identical to one in SPT required for valve margin expression. Further, SPT can heterodimerize with ALC, and 35S:SPT can fully complement dehiscence defects in alc mutants, although 35S:ALC can only partly complement spt gynoecium disruptions, perhaps associated with its sequence simplification. Interactions with FRUITFULL and SHATTERPROOF genes differ somewhat between SPT and ALC, reflecting their different specializations. These two genes are apparently undergoing subfunctionalization, with SPT essential for earlier carpel margin tissues, and ALC specializing in later dehiscence zone development.     

C- and D-class MADS-box genes from Phalaenopsis equestris (Orchidaceae) display functions in gynostemium and ovule development

Gynostemium and ovule development in orchid are unique developmental processes in the plant kingdom. Characterization of C- and D-class MADS-box genes could help reveal the molecular mechanisms underlying gynostemium and ovule development in orchids. In this study, we isolated and characterized a C- and a D-class gene, PeMADS1 and PeMADS7, respectively, from Phalaenopsis equestris. These two genes showed parallel spatial and temporal expression profiles, which suggests their cooperation in gynostemium and ovule development. Furthermore, only PeMADS1 was ectopically expressed in the petals of the gylp (gynostemium-like petal) mutant, whose petals were transformed into gynostemium-like structures. Protein-protein interaction analyses revealed that neither PeMADS1 and PeMADS7 could form a homodimer or a heterodimer. An E-class protein was needed to bridge the interaction between these two proteins. A complementation test revealed that PeMADS1 could rescue the phenotype of the AG mutant. Overexpression of PeMADS7 in Arabidopsis caused typical phenotypes of the D-class gene family. Together, these results indicated that both C-class PeMADS1 and D-class PeMADS7 play important roles in orchid gynostemium and ovule development.     

Overexpression of tomato LeAGP-1 arabinogalactan-protein promotes lateral branching and hampers reproductive development

LeAGP-1 is a glycosylphosphatidylinositol (GPI)-anchored arabinogalactan-protein (AGP) in tomato (Lycopersicon esculentum). Patterns of mRNA expression and protein localization for LeAGP-1 indicate that it likely functions in certain aspects of plant growth and development. To elucidate LeAGP-1 function(s), transgenic tomato plants expressing enhanced green fluorescent protein (GFP) fused to LeAGP-1 [GFP-LeAGP-1] or two LeAGP-1 variants, one lacking the C-terminal GPI-anchor domain [GFP-LeAGP-1DeltaC] and the other lacking the lysine-rich domain [GFP-LeAGP-1DeltaK], under the control of the CaMV35S promoter were produced using Agrobacterium-mediated transformation. Transgenic T0 and T1 lines with high levels of both GFP-LeAGP-1 mRNA and protein: (i) were significantly shorter; (ii) were highly branched; (iii) produced more flower buds, but most of these flowers did not mature, resulting in less fruit production; and (iv) produced seeds that were significantly smaller than normal seeds. Overexpression of LeAGP-1DeltaK had a similar or even more pronounced effect on plant vegetative and reproductive growth, while the effect of LeAGP-1DeltaC overexpression on plant reproduction was minimal. These results indicate that the GPI anchor is critical for LeAGP-1 function. As the phenotype of GFP-LeAGP-1 overexpressing transgenic plants is similar to that of cytokinin-overproducing plants, mRNA expression patterns of LeAGP-1 under different hormone treatments were examined. Cytokinins upregulated LeAGP-1 mRNA expression, while auxins and ABA inhibited LeAGP-1 mRNA expression. Based on these results, GPI-anchored LeAGP-1 most likely functions in plant growth and development in concert with auxin/cytokinin signaling.