Germline mutations in BMPR1A/ALK3 cause a subset of cases of juvenile polyposis syndrome and of Cowden and Bannayan-Riley-Ruvalcaba syndromes

Juvenile polyposis syndrome (JPS) is an inherited hamartomatous-polyposis syndrome with a risk for colon cancer. JPS is a clinical diagnosis by exclusion, and, before susceptibility genes were identified, JPS could easily be confused with other inherited hamartoma syndromes, such as Bannayan-Riley-Ruvalcaba syndrome (BRRS) and Cowden syndrome (CS). Germline mutations of MADH4 (SMAD4) have been described in a variable number of probands with JPS. A series of familial and isolated European probands without MADH4 mutations were analyzed for germline mutations in BMPR1A, a member of the transforming growth-factor beta-receptor superfamily, upstream from the SMAD pathway. Overall, 10 (38%) probands were found to have germline BMPR1A mutations, 8 of which resulted in truncated receptors and 2 of which resulted in missense alterations (C124R and C376Y). Almost all available component tumors from mutation-positive cases showed loss of heterozygosity (LOH) in the BMPR1A region, whereas those from mutation-negative cases did not. One proband with CS/CS-like phenotype was also found to have a germline BMPR1A missense mutation (A338D). Thus, germline BMPR1A mutations cause a significant proportion of cases of JPS and might define a small subset of cases of CS/BRRS with specific colonic phenotype.     

Determinants of antagonist binding at the alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor subunit, GluR-D. Role of the conserved arginine 507 and glutamate 727 residues.

Previous structural and mutagenesis studies indicate that the invariant alpha-amino and alpha-carboxyl groups of glutamate receptor agonists are engaged in polar interactions with oppositely charged, conserved arginine and glutamate residues in the ligand-binding domain of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor. To examine the role of these residues (R507 and E727 in the GluR-D subunit) in the discrimination between agonists and antagonists, we analyzed the ligand-binding properties of homomeric GluR-D and its soluble ligand-binding domain with mutations at these positions. Filter-binding assays using [3H]AMPA, an agonist, and [3H]Ro 48-8587, a high-affinity antagonist, as radioligands revealed that even a conservative mutation at R507 (R507K) resulted in the complete loss of both agonist and antagonist binding. In contrast, a negative charge at position 727 was necessary for agonist binding, whereas the isosteric mutation, E727Q, abolished all agonist binding but retained high-affinity binding for [3H]Ro 48-8587, displaceable by 7,8-dinitroquinoxaline-2,3-dione. Competition binding studies with antagonists representing different structural classes in combination with ligand docking experiments suggest that the role of E727 is antagonist-specific, ranging from no interaction to weak electrostatic interactions involving indirect and direct hydrogen bonding with the antagonist molecule. These results underline the importance of ion pair interaction with E727 for agonist activity and suggest that an interaction with R507, but not with E727, is essential for antagonist binding.     

Flavoprotein hydroxylase PgaE catalyzes two consecutive oxygen-dependent tailoring reactions in angucycline biosynthesis

A simplified model system composed of a NADPH-dependent flavoprotein hydroxylase PgaE and a short-chain alcohol dehydrogenase/reductase (SDR) CabV was used to dissect a multistep angucycline modification redox cascade into several subreactions in vitro. We demonstrate that the two enzymes are sufficient for the conversion of angucycline substrate 2,3-dehydro-UWM6 to gaudimycin C. The flavoenzyme PgaE is shown to be responsible for two consecutive NADPH- and O(2)-dependent reactions, consistent with the enzyme-catalyzed incorporation of oxygen atoms at C-12 and C-12b in gaudimycin C. The two reactions do not significantly overlap, and the second catalytic cycle is initiated only after the original substrate 2,3-dehydro-UWM6 is nearly depleted. This allowed us to isolate the product of the first reaction at limiting NADPH concentrations and allowed the study of the qualitative and kinetic properties of the separated reactions. Dissection of the reaction cascade also allowed us to establish that the SDR reductase CabV catalyzes the final biosynthetic step, which is closely coupled to the second PgaE reaction. In the absence of CabV, the complete PgaE reaction leads invariably to product degradation, whereas in its presence, the reaction yields the final product, gaudimycin C. The result implies that the C-6 ketoreduction step catalyzed by CabV is required for stabilization of a reactive intermediate. The close relationship between PgaE and CabV would explain previous in vivo observations: why the absence of a reductase gene may result in the lack of C-12b-oxygenated species and, vice versa, why all C-12b-oxygenated angucyclines appear to have undergone reduction at position C-6.     

Epigenetic analysis of sporadic and Lynch-associated ovarian cancers reveals histology-specific patterns of DNA methylation

Diagnosis and treatment of epithelial ovarian cancer is challenging due to the poor understanding of the pathogenesis of the disease. Our aim was to investigate epigenetic mechanisms in ovarian tumorigenesis and, especially, whether tumors with different histological subtypes or hereditary background (Lynch syndrome) exhibit differential susceptibility to epigenetic inactivation of growth regulatory genes. Gene candidates for epigenetic regulation were identified from the literature and by expression profiling of ovarian and endometrial cancer cell lines treated with demethylating agents. Thirteen genes were chosen for methylation-specific multiplex ligation-dependent probe amplification assays on 104 (85 sporadic and 19 Lynch syndrome-associated) ovarian carcinomas. Increased methylation (i.e., hypermethylation) of variable degree was characteristic of ovarian carcinomas relative to the corresponding normal tissues, and hypermethylation was consistently more prominent in non-serous than serous tumors for individual genes and gene sets investigated. Lynch syndrome-associated clear cell carcinomas showed the highest frequencies of hypermethylation. Among endometrioid ovarian carcinomas, lower levels of promoter methylation of RSK4, SPARC, and HOXA9 were significantly associated with higher tumor grade; thus, the methylation patterns showed a shift to the direction of high-grade serous tumors. In conclusion, we provide evidence of a frequent epigenetic inactivation of RSK4, SPARC, PROM1, HOXA10, HOXA9, WT1-AS, SFRP2, SFRP5, OPCML, and MIR34B in the development of non-serous ovarian carcinomas of Lynch and sporadic origin, as compared to serous tumors. Our findings shed light on the role of epigenetic mechanisms in ovarian tumorigenesis and identify potential targets for translational applications.     

Larval growth in polyphenic salamanders: making the best of a bad lot

Polyphenisms are excellent models for studying phenotypic variation, yet few studies have focused on natural populations. Facultative paedomorphosis is a polyphenism in which salamanders either metamorphose or retain their larval morphology and eventually become paedomorphic. Paedomorphosis can result from selection for capitalizing on favorable aquatic habitats (paedomorph advantage), but could also be a default strategy under poor aquatic conditions (best of a bad lot). We tested these alternatives by quantifying how the developmental environment influences the ontogeny of wild Arizona tiger salamanders (Ambystoma tigrinum nebulosum). Most paedomorphs in our study population arose from slow-growing larvae that developed under high density and size-structured conditions (best of a bad lot), although a few faster-growing larvae also became paedomorphic (paedomorph advantage). Males were more likely to become paedomorphs than females and did so under a greater range of body sizes than females, signifying a critical role for gender in this polyphenism. Our results emphasize that the same phenotype can be adaptive under different environmental and genetic contexts and that studies of phenotypic variation should consider multiple mechanisms of morph production.     

Early-onset renal cell carcinoma as a novel extraparaganglial component of SDHB-associated heritable paraganglioma

Hereditary paraganglioma syndrome has recently been shown to be caused by germline heterozygous mutations in three (SDHB, SDHC, and SDHD) of the four genes that encode mitochondrial succinate dehydrogenase. Extraparaganglial component neoplasias have never been previously documented. In a population-based registry of symptomatic presentations of phaeochromocytoma/paraganglioma comprising 352 registrants, among whom 16 unrelated registrants were SDHB mutation positive, one family with germline SDHB mutation c.847-50delTCTC had two members with renal cell carcinoma (RCC), of solid histology, at ages 24 and 26 years. Both also had paraganglioma. A registry of early-onset RCCs revealed a family comprising a son with clear-cell RCC and his mother with a cardiac tumor, both with the germline SDHB R27X mutation. The cardiac tumor proved to be a paraganglioma. All RCCs showed loss of the remaining wild-type allele. Our observations suggest that germline SDHB mutations can predispose to early-onset kidney cancers in addition to paragangliomas and carry implications for medical surveillance.     

Eleven Candidate Susceptibility Genes for Common Familial Colorectal Cancer

Hereditary factors are presumed to play a role in one third of colorectal cancer (CRC) cases. However, in the majority of familial CRC cases the genetic basis of predisposition remains unexplained. This is particularly true for families with few affected individuals. To identify susceptibility genes for this common phenotype, we examined familial cases derived from a consecutive series of 1514 Finnish CRC patients. Ninety-six familial CRC patients with no previous diagnosis of a hereditary CRC syndrome were included in the analysis. Eighty-six patients had one affected first-degree relative, and ten patients had two or more. Exome sequencing was utilized to search for genes harboring putative loss-of-function variants, because such alterations are likely candidates for disease-causing mutations. Eleven genes with rare truncating variants in two or three familial CRC cases were identified: UACA, SFXN4, TWSG1, PSPH, NUDT7, ZNF490, PRSS37, CCDC18, PRADC1, MRPL3, and AKR1C4. Loss of heterozygosity was examined in all respective cancer samples, and was detected in seven occasions involving four of the candidate genes. In all seven occasions the wild-type allele was lost (P = 0.0078) providing additional evidence that these eleven genes are likely to include true culprits. The study provides a set of candidate predisposition genes which may explain a subset of common familial CRC. Additional genetic validation in other populations is required to provide firm evidence for causality, as well as to characterize the natural history of the respective phenotypes.     

Patterns of size change in late Neogene pocket gophers from the Meade Basin of Kansas and Oklahoma

A dense late Neogene history of pocket gophers from the Meade Basin of south-western Kansas and north-western Oklahoma preserves a detailed record of size change, as indicated by mean length of the lower fourth premolar. A ‘time for space substitution’ interpretation results in the tentative recognition of a pattern analogous to character displacement, in which the small to medium-sized Pliogeomys buisi, P. louderbachi, Geomys minor and G. floralindae are considered ecological analogues and treated as a single ecological entity, Avatar A. The larger G. jacobi and G. quinni are combined as Avatar B. Statistical analyses confirm that populations of Avatar A are of equal size when allochronic from Avatar B, and significantly smaller when synchronic (and sympatric) with Avatar B. However, this pattern is complicated by the observation that Avatar A began dwarfing before the appearance of Avatar B. This historical perspective suggests that static size patterns among modern taxa may have a complex history. Occasional diminutive geomyid transients (G. adamsi, Thomomys cf talpoides) are always paired with significantly larger, medium-sized Geomys. Although a random walk explanation for the pattern of size changes is unlikely, we are currently unable to distinguish between environmental change and competition as the likely forcing mechanisms.     

Interactions between habitat heterogeneity and food affect reproductive output in a top predator

1. Habitat heterogeneity has important repercussions for species abundance, demography and life-history patterns. While habitat effects have been more thoroughly studied in top-down situations (e.g. in association with predation), their role in bottom-up situations (e.g. in association with food abundance) has been less explored and the underlying mechanism(s) behind the ecological patterns have not commonly been identified. 2. With material from 1993 to 2003, we test the hypothesis that the reproduction of Finnish northern goshawks Accipiter gentilis (L.) is bottom-up limited by habitat composition, especially in situations where the density of their main prey (grouse) is low. Special emphasis was placed on identifying the mechanism(s) behind potential habitat effects. 3. While laying date and large-scale variation in the main prey density (but not habitat composition) were related to the number of eggs goshawks laid, small-scale differences in alternative prey density between different territories later influenced how many young were fledged via the mechanism of habitat-dependent partial-brood loss. As a result of this mechanism, a difference in nestling condition also arose between goshawk territories with differing habitat compositions. 4. As the relative proportions of different landscape elements in a given landscape is a function of large-scale differences in geomorphology and land use, this means that the reproductive performance of goshawks as averaged over larger scales can be understood correctly only in respect to the fact that habitat gradients differ across landscapes. 5. In addition to being one of the first papers identifying the mechanism of partial brood loss as being primarily responsible for the habitat-specific differences in the production of young, this study further illustrates the need to identify small-scale mechanisms to correctly understand the large-scale patterns of reproductive performance in territorial species. The repercussions of the observed habitat effect for local population development are discussed.