A role for Alström syndrome protein, alms1, in kidney ciliogenesis and cellular quiescence

Premature truncation alleles in the ALMS1 gene are a frequent cause of human Alstr?m syndrome. Alstr?m syndrome is a rare disorder characterized by early obesity and sensory impairment, symptoms shared with other genetic diseases affecting proteins of the primary cilium. ALMS1 localizes to centrosomes and ciliary basal bodies, but truncation mutations in Alms1/ALMS1 do not preclude formation of cilia. Here, we show that in vitro knockdown of Alms1 in mice causes stunted cilia on kidney epithelial cells and prevents these cells from increasing calcium influx in response to mechanical stimuli. The stunted-cilium phenotype can be rescued with a 5' fragment of the Alms1 cDNA, which resembles disease-associated alleles. In a mouse model of Alstr?m syndrome, Alms1 protein can be stably expressed from the mutant allele and is required for cilia formation in primary cells. Aged mice developed specific loss of cilia from the kidney proximal tubules, which is associated with foci of apoptosis or proliferation. As renal failure is a common cause of mortality in Alstr?m syndrome patients, we conclude that this disease should be considered as a further example of the class of renal ciliopathies: wild-type or mutant alleles of the Alstr?m syndrome gene can support normal kidney ciliogenesis in vitro and in vivo, but mutant alleles are associated with age-dependent loss of kidney primary cilia.     

Expression of NEP2, a soluble neprilysin-like endopeptidase, during embryogenesis in Drosophila melanogaster

Members of the neprilysin family of neutral endopeptidases (M13) are typically membrane-bound enzymes known to be involved in the extra-cellular metabolism of signalling peptides and have important roles during mammalian embryogenesis. In this study we show that membranes prepared from embryos of Drosophila melanogaster possess neprilysin-like activity that is inhibited by phosphoramidon and thiorphan, both inhibitors of mammalian neprilysin. Unexpectedly, we also found strong neprilysin-like neutral endopeptidase activity in a soluble embryo fraction, which we identify as NEP2 by Western blot and immunoprecipitation experiments using NEP2 specific antibodies. NEP2 is a soluble secreted member of the neprilysin family that has been shown previously to be expressed in larval and adult Malpighian tubules and in the testes of adult males. In situ hybridization studies reveal expression at stage 10-11 in a pattern similar to that previously described for stellate cell progenitors of the caudal visceral mesoderm. In later stages of embryogenesis, some of these cells appear to migrate into the growing Malpighian tubule. Recombinant NEP2 protein is N-glycosylated and displays optimum endopeptidase activity at neutral pH, consistent with a role as an extracellular peptidase. The recombinant enzyme hydrolyses Drosophila tachykinin peptides (DTK) at peptide bonds N-terminal to hydrophobic residues. DTK2, like Locusta tachykinin-1, was cleaved at the penultimate peptide bond (Gly(7)-Leu(8)), whereas the other Drosophila peptides were cleaved centrally at Xxx-Phe bonds. However, the rates of hydrolysis of the latter substrates were much slower than the hydrolysis rates of DTK2 and Locusta tachykinin-1, suggesting that the interaction of the bulky side-chain of phenylalanine at the S'(1) sub-site is less favorable for peptide bond hydrolysis. The secretion of NEP2 from tissues during embryogenesis suggests a possible developmental role for this endopeptidase in peptide signalling in D. melanogaster.     

Molecular dynamics simulations and coupled nucleotide substitution experiments indicate the nature of A·A base pairing and a putative structure of the coralyne-induced homo-adenine duplex

Coralyne is an alkaloid drug that binds homo-adenine DNA (and RNA) oligonucleotides more tightly than it does Watson–Crick DNA. Hud’s laboratory has shown that poly(dA) in the presence of coralyne forms an anti-parallel duplex, however attempts to determine the structure by NMR spectroscopy and X-ray crystallography have been unsuccessful. Assuming adenine–adenine hydrogen bonding between the two poly(dA) strands, we constructed 40 hypothetical homo-(dA) anti-parallel duplexes and docked coralyne into the six most favorable duplex structures. The two most stable structures had trans glycosidic bonds, but distinct pairing geometries, i.e. either Watson–Crick Hoogsteen (transWH) or Watson–Crick Watson–Crick (transWW) with stability of transWH > transWW. To narrow down the possibilities, 7-deaza adenine base substitutions (dA→7) were engineered into homo-(dA) sequences. These substitutions significantly reduced the thermal stability of the coralyne-induced homo-(dA) structure. These experiments strongly suggest the involvement of N7 in the coralyne-induced A·A base pairs. Moreover, due to the differential effect on melting as a function of the location of the dA→7 mutations, these results are consistent with the N1–N7 base pairing of the transWH pairs. Together, the simulation and base substitution experiments predict that the coralyne-induced homo-(dA) duplex structure adopts the transWH geometry.     

Structural models of primary cell walls in flowering plants: consistency of molecular structure with the physical properties of the walls during growth

Advances in determination of polymer structure and in preservation of structure for electron microscopy provide the best view to date of how polysaccharides and structural proteins are organized into plant cell walls. The walls that form and partition dividing cells are modified chemically and structurally from the walls expanding to provide a cell with its functional form. In grasses, the chemical structure of the wall differs from that of all other flowering plant species that have been examined. Nevertheless, both types of wall must conform to the same physical laws. Cell expansion occurs via strictly regulated reorientation of each of the wall's components that first permits the wall to stretch in specific directions and then lock into final shape. This review integrates information on the chemical structure of individual polymers with data obtained from new techniques used to probe the arrangement of the polymers within the walls of individual cells. We provide structural models of two distinct types of walls in flowering plants consistent with the physical properties of the wall and its components.     

Interrelatedness of 5S RNA sequences investigated by correspondence analysis

Summary Correspondence analysis (a form of multivariate statistics) applied to 74 5S ribosomal RNA sequences indicates that the sequences are interrelated in a systematic, nonrandom fashion. Aligned sequences are represented as vectors in a 5N-dimensional space, where N is the number of base positions in the 5S RNA molecule. Mutually orthogonal directions (called factor axes) along which intersequence variance is greatest are defined in this hyperspace. Projection of the sequences onto planes defined by these factorial directions reveals clustering of species that is suggestive of phylogenetic relationships. For each factorial direction, correspondence analysis points to regions of importance, i.e., those base positions at which the systematic changes occur that define that particular direction. In effect, the technique provides a rapid determination of group-specific signatures. In several instances, similarities between sequences are indicated that have only recently been inferred from visual base-to-base comparisons. These results suggest that correspondence analysis may provide a valuable starting point from which to uncover the patterns of change underlying the evolution of a macromolecule, such as 5S RNA.     

Assessing the ecological risks from the persistence and spread of feral populations of insect-resistant transgenic maize

One source of potential harm from the cultivation of transgenic crops is their dispersal, persistence and spread in non-agricultural land. Ecological damage may result from such spread if the abundance of valued species is reduced. The ability of a plant to spread in non-agricultural habitats is called its invasiveness potential. The risks posed by the invasiveness potential of transgenic crops are assessed by comparing in agronomic field trials the phenotypes of the crops with the phenotypes of genetically similar non-transgenic crops known to have low invasiveness potential. If the transgenic and non-transgenic crops are similar in traits believed to control invasiveness potential, it may be concluded that the transgenic crop has low invasiveness potential and poses negligible ecological risk via persistence and spread in non-agricultural habitats. If the phenotype of the transgenic crop is outside the range of the non-transgenic comparators for the traits controlling invasiveness potential, or if the comparative approach is regarded as inadequate for reasons of risk perception or risk communication, experiments that simulate the dispersal of the crop into non-agricultural habitats may be necessary. We describe such an experiment for several commercial insect-resistant transgenic maize events in conditions similar to those found in maize-growing regions of Mexico. As expected from comparative risk assessments, the transgenic maize was found to behave similarly to non-transgenic maize and to be non-invasive. The value of this experiment in assessing and communicating the negligible ecological risk posed by the low invasiveness potential of insect-resistant transgenic maize in Mexico is discussed.     

Fast parallel Markov clustering in bioinformatics using massively parallel computing on GPU with CUDA and ELLPACK-R sparse format

Markov clustering (MCL) is becoming a key algorithm within bioinformatics for determining clusters in networks. However,with increasing vast amount of data on biological networks, performance and scalability issues are becoming a critical limiting factor in applications. Meanwhile, GPU computing, which uses CUDA tool for implementing a massively parallel computing environment in the GPU card, is becoming a very powerful, efficient, and low-cost option to achieve substantial performance gains over CPU approaches. The use of on-chip memory on the GPU is efficiently lowering the latency time, thus, circumventing a major issue in other parallel computing environments, such as MPI. We introduce a very fast Markov clustering algorithm using CUDA (CUDA-MCL) to perform parallel sparse matrix-matrix computations and parallel sparse Markov matrix normalizations, which are at the heart of MCL. We utilized ELLPACK-R sparse format to allow the effective and fine-grain massively parallel processing to cope with the sparse nature of interaction networks data sets in bioinformatics applications. As the results show, CUDA-MCL is significantly faster than the original MCL running on CPU. Thus, large-scale parallel computation on off-the-shelf desktop-machines, that were previously only possible on supercomputing architectures, can significantly change the way bioinformaticians and biologists deal with their data.     

Grandparentage assignments identify unexpected adfluvial life history tactic contributing offspring to a reintroduced population

Diversity in life history tactics contributes to the persistence of a population because it helps to protect against stochastic environments by varying individuals in space and time. However, some life history tactics may not be accounted for when assessing the demographic viability of a population. One important factor in demographic viability assessments is cohort replacement rate (CRR), which is defined as the number of future adults produced by an adult. We assessed if precocial resident males (<age‐3) and adfluvial Chinook salmon (Oncorhynchus tshawytscha), adults that reside in freshwater their entire lives, contributed offspring to a reintroduced population from 2008 to 2013. We found that 9 ± 5% of offspring with an unassigned parent remained unexplained after accounting for sources of human error. Using grandparentage assignments, we identified 31 precocial resident males and 48 probable adfluvial Chinook salmon produced by anadromous mate pairs from 2007 to 2012. Previously published CRR estimates for the 2007 and 2008 reintroduced adults, based on only anadromous returning adult offspring, were 0.40 and 0.31, respectively. By incorporating adfluvial females, we found CRR estimates increased by 17% (CRR: 0.46) and 13% (CRR: 0.35) for the 2007 and 2008 cohorts, respectively.