Inhibition of myoblast differentiation by Sfrp1 and Sfrp2

Secreted Frizzled-related proteins (Sfrps) are extracellular regulators of Wnt signalling and play important roles in developmental and oncogenic processes. They are known to be upregulated in regenerating muscle and in myoblast cultures but their function is unknown. Here, we show that the addition of recombinant Sfrp1 or Sfrp2 to C2C12 cell line cultures or to primary cultures of satellite cells results in the inhibition of myotube formation with no significant effect on the cell cycle or apoptosis. Even though at confluence, treated and untreated cultures are identical in appearance, analyses have shown that, for maximum effect, the cells have to be treated while they are proliferating. Furthermore, removal of Sfrp from the culture medium during differentiation restores normal myotube formation. We conclude that Sfrp1 and Sfrp2 act to prevent myoblasts from entering the terminal differentiation process. S. Descamps and J. Levin contributed equally to this work.     

Sfrp1 and Sfrp2 regulate anteroposterior axis elongation and somite segmentation during mouse embryogenesis

Regulation of Wnt signaling is essential for embryonic patterning. Sfrps are secreted Wnt antagonists that directly interact with the Wnt ligand to inhibit signaling. Here, we show that Sfrp1 and Sfrp2 are required for anteroposterior (AP) axis elongation and somitogenesis in the thoracic region during mouse embryogenesis. Double homozygous mutations in Sfrp1 and Sfrp2 lead to severe shortening of the thoracic region. By contrast, a homozygous mutation in one or the other exerts no effect on embryogenesis, indicating that Sfrp1 and Sfrp2 are functionally redundant. The defect of a shortened thoracic region appears to be the consequence of AP axis reduction and incomplete somite segmentation. The reduction in the AP axis is partially due to abnormalities in cell migration of pre-somitic mesoderm from the end of gastrulation. Aberrant somite segmentation is associated with altered oscillations of Notch signaling, as evidenced by abnormal Lfng and Hes7 expression during somitogenesis in the thoracic region. This study suggests that Wnt regulation by Sfrp1 and Sfrp2 is required for embryonic patterning.     

Sfrp1a and Sfrp5 function as positive regulators of Wnt and BMP signaling during early retinal development

Axial patterning of the developing eye is critically important for proper axonal pathfinding as well as for key morphogenetic events, such as closure of the optic fissure. The dorsal retina is initially specified by the actions of Bone Morphogenetic Protein (BMP) signaling, with such identity subsequently maintained by the Wnt-β catenin pathway. Using zebrafish as a model system, we demonstrate that Secreted frizzled-related protein 1a (Sfrp1a) and Sfrp5 work cooperatively to pattern the retina along the dorso-ventral axis. Sfrp1a/5 depleted embryos display a reduction in dorsal marker gene expression that is consistent with defects in BMP- and Wnt-dependent dorsal retina identity. In accord with this finding, we observe a marked reduction in transgenic reporters of BMP and Wnt signaling within the dorsal retina of Sfrp1a/5 depleted embryos. In contrast to studies in which canonical Wnt signaling is blocked, we note an increase in BMP ligand expression in Sfrp1a/5 depleted embryos, a phenotype similar to that seen in embryos with inhibited BMP signaling. Overexpression of a low dose of sfrp5 mRNA causes an increase in dorsal retina marker gene expression. We propose a model in which Sfrp proteins function as facilitators of both BMP and Wnt signaling within the dorsal retina.     

Sfrp1, Sfrp2, and Sfrp5 regulate the Wnt/beta-catenin and the planar cell polarity pathways during early trunk formation in mouse

Sfrp is a secreted Wnt antagonist that directly interacts with Wnt ligand. We show here that inactivation of Sfrp1, Sfrp2, and Sfrp5 leads to fused somites formation in early-somite mouse embryos, simultaneously resulting in defective convergent extension (CE), which causes severe shortening of the anteroposterior axis. These observations indicate the redundant roles of Sfrp1, Sfrp2, and Sfrp5 in early trunk formation. The roles of the Sfrps were genetically distinguished in terms of the regulation of Wnt pathways. Genetic analysis combining Sfrps mutants and Loop-tail mice revealed the involvement of Sfrps in CE through the regulation of the planar cell polarity pathway. Furthermore, Dkk1-deficient embryos carrying Sfrp1 homozygous and Sfrp2 heterozygous mutations display irregular somites and indistinct intersomitic boundaries, which indicates that Sfrps-mediated inhibition of the Wnt/beta-catenin pathway is necessary for somitogenesis. Our results suggest that Sfrps regulation of the canonical and noncanonical pathways is essential for proper trunk formation.     

Breakdown of self-incompatibility in a natural population of Petunia axillaris caused by loss of pollen function

Although Petunia axillaris subsp. axillaris is described as a self-incompatible taxon, some of the natural populations we have identified in Uruguay are composed of both self-incompatible and self-compatible plants. Here, we studied the self-incompatibility (SI) behavior of 50 plants derived from such a mixed population, designated U83, and examined the cause of the breakdown of SI. Thirteen plants were found to be self-incompatible, and the other 37 were found to be self-compatible. A total of 14 S-haplotypes were represented in these 50 plants, including two that we had previously identified from another mixed population, designated U1. All the 37 self-compatible plants carried either an S(C1)- or an S(C2)-haplotype. S(C1)S(C1) and S(C2)S(C2) homozygotes were generated by self-pollination of two of the self-compatible plants, and they were reciprocally crossed with 40 self-incompatible S-homozygotes (S(1)S(1) through S(40)S(40)) generated from plants identified from three mixed populations, including U83. The S(C1)S(C1) homozygote was reciprocally compatible with all the genotypes examined. The S(C2)S(C2) homozygote accepted pollen from all but the S(17)S(17) homozygote (identified from the U1 population), but the S(17)S(17) homozygote accepted pollen from the S(C2)S(C2) homozygote. cDNAs encoding S(C2)- and S(17)-RNases were cloned and sequenced, and their nucleotide sequences were completely identical. Analysis of bud-selfed progeny of heterozygotes carrying S(C1) or S(C2) showed that the SI behavior of S(C1) and S(C2) was identical to that of S(C1) and S(C2) homozygotes, respectively. All these results taken together suggested that the S(C2)-haplotype was a mutant form of the S(17)-haplotype, with the defect lying in the pollen function. The possible nature of the mutation is discussed.     

Minute mosaics caused by early chromosome loss

Summary Two genetic operations have been combined in order to ascertain whether there are differential proliferation rates in the syncytial nuclei and the blastoderm cells prior to the formation of the imaginal disc anlagen. Early chromosome loss caused by the mutantca ^nd has been associated with the generation ofMinute (M/M ⁺) genotypes in normal (M ⁺/M ⁺) zygotes or of non-Minute genotypes inMinute zygotes. The results indicate that there is no growth competition betweenMinute and non-Minute cells prior to the formation of the imaginal discs. Growth competition, however, leads later, during the proliferation phase of the discs, to the demarcation of compartment boundaries within imaginal discs.     

Profound urinary protein loss and acute renal failure caused by cyclooxygenase-2 inhibitor

Cumulative evidence has shown that nonsteroidal anti-inflammatory drugs (NSAIDs) can induce acute renal failure and nephrotic-range proteinuria. Cyclooxygenase-2 (COX-2) inhibitors have less nephrotoxicity; however, recent data indicate that they may cause the same renal problems as NSAIDs do. Herein, we present a case of celecoxib-associated minimal change disease (MCD) with profound urinary protein loss and acute renal failure. Renal function and nephrotic syndrome in this patient resolved completely after discontinuation of celecoxib and treatment with methylprednisolone. Clinicians should keep high index of suspicions in patients developing nephrotic syndrome and acute renal failure after taking COX-2 inhibitors since secondary MCD responds well to timely cessation of COX-2 inhibitors and administration of steroid therapy.     

Rescue of a lysosomal storage disorder caused by Grn loss of function with a brain penetrant progranulin biologic

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Rapid evolution caused by pollinator loss in Mimulus guttatus

Anthropogenic perturbations including habitat loss and emerging disease are changing pollinator communities and generating novel selection pressures on plant populations. Disruption of plant-pollinator relationships is predicted to cause plant mating system evolution, although this process has not been directly observed. This study demonstrates the immediate evolutionary effects of pollinator loss within experimental populations of a predominately outcrossing wildflower. Initially equivalent populations evolved for five generations within two pollination treatments: abundant bumblebee pollinators versus no pollinators. The populations without pollinators suffered greatly reduced fitness in early generations but rebounded as they evolved an improved ability to self-fertilize. All populations diverged in floral, developmental, and life-history traits, but only a subset of characters showed clear association with pollination treatment. Pronounced treatment effects were noted for anther-stigma separation and autogamous seed set. Dramatic allele frequency changes at two chromosomal polymorphisms occurred in the no pollinator populations, explaining a large fraction of divergence in pollen viability. The pattern of phenotypic and genetic changes in this experiment favors a sequential model for the evolution of the multitrait "selfing syndrome" observed throughout angiosperms.     

Degenerative disorders caused by Bcl-2 deficiency prevented by loss of its BH3-only antagonist Bim.

Apoptosis is triggered when proapoptotic members of the Bcl-2 protein family bearing only the BH3 association domain bind to Bcl-2 or its homologs and block their antiapoptotic activity. To test whether loss of the BH3-only protein Bim could prevent the cellular attrition caused by Bcl-2 deficiency, we generated mice lacking both genes. Mice without Bcl-2 have a fragile lymphoid system, become runted, turn gray, and succumb to polycystic kidney disease. Concomitant absence of Bim prevented all these disorders. Indeed, loss of even one bim allele restored normal kidney development, growth, and health. These results demonstrate that Bim levels set the threshold for initiation of apoptosis in several tissues and suggest that degenerative diseases might be alleviated by blocking BH3-only proteins.     

Naturally occurring broad-spectrum powdery mildew resistance in a Central American tomato accession is caused by loss of mlo function

The resistant cherry tomato (Solanum lycopersicum var. cerasiforme) line LC-95, derived from an accession collected in Ecuador, harbors a natural allele (ol-2) that confers broad-spectrum and recessively inherited resistance to powdery mildew (Oidium neolycopersici). As both the genetic and phytopathological characteristics of ol-2-mediated resistance are reminiscent of powdery mildew immunity conferred by loss-of-function mlo alleles in barley and Arabidopsis, we initiated a candidate-gene approach to clone Ol-2. A tomato Mlo gene (SlMlo1) with high sequence-relatedness to barley Mlo and Arabidopsis AtMLO2 mapped to the chromosomal region harboring the Ol-2 locus. Complementation experiments using transgenic tomato lines as well as virus-induced gene silencing assays suggested that loss of SlMlo1 function is responsible for powdery mildew resistance conferred by ol-2. In progeny of a cross between a resistant line bearing ol-2 and the susceptible tomato cultivar Moneymaker, a 19-bp deletion disrupting the SlMlo1 coding region cosegregated with resistance. This polymorphism results in a frameshift and, thus, a truncated nonfunctional SlMlo1 protein. Our findings reveal the second example of a natural mlo mutant that possibly arose post-domestication, suggesting that natural mlo alleles might be evolutionarily short-lived due to fitness costs related to loss of mlo function.