Fluvial processes and landforms

The period 1965-2000 saw a sustained increase in research and publication on fluvial processes and landforms. The trend toward generalisation and/or mechanistic understanding, rather than site-specific history, continued. Research was multi-disciplinary, with important contributions from hydraulic engineers, geologists and physical geographers and from experimental and theoretical approaches as well as geomorphological and sedimentological fieldwork. Rapidly increasing computer power underpinned new measurement methods and greatly increased the scope of data analysis and numerical modelling. There were major advances in understanding the interaction of river process and form at reach scale, with growing recognition of differences between sand-bed and coarse-bed rivers. Field studies outside Europe and North America led to greater awareness of the diversity of river planforms and deposition landforms. Conceptual models of how rivers respond to natural or anthropogenic change in boundary conditions at different timescales were refined, taking advantage of studies of response to land use change, major floods, and volcanic eruptions. Dating of sediments allowed greater appreciation of fluctuations in the incidence of extreme driving events over centuries and thousands of years. Towards the end of the period research on bedrock rivers began to take off.

How well can we predict earthquake site response so far? Site-specific approaches

Earthquake site responses or site effects are the modifications of surface geology to seismic waves. How well can we predict the site effects (average over many earthquakes) at individual sites so far? To address this question, we tested and compared the effectiveness of different estimation techniques in predicting the outcrop Fourier site responses separated using the general inversion technique (GIT) from recordings. Techniques being evaluated are (a) the empirical correction to the horizontal-to-vertical spectral ratio of earthquakes (c-HVSR), (b) one-dimensional ground response analysis (GRA), and (c) the square-root-impedance (SRI) method (also called the quarter-wavelength approach). Our results show that c-HVSR can capture significantly more site-specific features in site responses than both GRA and SRI in the aggregate, especially at relatively high frequencies. c-HVSR achieves a “good match” in spectral shape at ∼80%–90% of 145 testing sites, whereas GRA and SRI fail at most sites. GRA and SRI results have a high level of parametric and/or modeling errors which can be constrained, to some extent, by collecting on-site recordings.

Limb Specific Failure of Proliferation and Translation in the Mesenchyme Leads to Skeletal Defects in Diamond Blackfan Anemia

Ribosomopathies are a class of disorders caused by defects in the structure or function of the ribosome and characterized by tissue-specific abnormalities. Diamond Blackfan anemia (DBA) arises from different mutations, predominantly in genes encoding ribosomal proteins (RPs). Apart from the anemia, skeletal defects are among the most common anomalies observed in patients with DBA, but they are virtually restricted to radial ray and other upper limb defects. What leads to these site-specific skeletal defects in DBA remains a mystery. Using a novel mouse model for RP haploinsufficiency, we observed specific, differential defects of the limbs. Using complementary in vitro and in vivo approaches, we demonstrate that reduced WNT signaling and subsequent increased β-catenin degradation in concert with increased expression of p53 contribute to mesenchymal lineage failure. We observed differential defects in the proliferation and differentiation of mesenchymal stem cells (MSCs) from the forelimb versus the hind limbs of the RP haploinsufficient mice that persisted after birth and were partially rescued by allelic reduction of Trp53. These defects are associated with a global decrease in protein translation in RP haploinsufficient MSCs, with the effect more pronounced in cells isolated from the forelimbs. Together these results demonstrate translational differences inherent to the MSC, explaining the site-specific skeletal defects observed in DBA.

Increased potential for shell competition among hermit crabs (Pagurus beringanus and Pagurus granosimanus) in the rocky intertidal

We examined the prevalence and shell use of two species of hermit crabs (Pagurus granosimanus and Pagurus beringanus) in exposed and protected microhabitats at five sites in the rocky temperate intertidal on San Juan Island, Washington, to compare present habitat partitioning and potential interspecific competition to that reported nearly 50 years ago. We found that, in contrast to previous findings, the two species of hermit crabs overlapped extensively at some sites, typically those with less wave action. While the hermit crabs typically inhabited certain types of shells significantly more than others, and that use was congruent across microhabitats and species of hermit crabs at the same site, the dominant domicile differed substantially across sites. We provide a more complete ranking of shell use than previous authors and note site-specific dominant shell use. We conclude that previous habitat partitioning by depth may have weakened at protected sites. We hypothesize that increasing temperatures have caused P. granosimanus to expand its range deeper into the intertidal, which may increase the degree of interspecific competition for shells at the edge of the species’ tidal height range, where they overlap. Whether the habitat shift by this hermit crab is due to recent alterations in climate (particularly elevated temperatures, ocean acidification and lower local open ocean salinity) is unknown, but warrants further study.

Application of RtcB ligase to monitor self-cleaving ribozyme activity by RNA-seq

Self-cleaving ribozymes are catalytic RNAs and can be found in all domains of life. They catalyze a site-specific cleavage that results in a 5′ fragment with a 2′,3′ cyclic phosphate (2′,3′ cP) and a 3′ fragment with a 5′ hydroxyl (5′ OH) end. Recently, several strategies to enrich self-cleaving ribozymes by targeted biochemical methods have been introduced by us and others. Here, we develop an alternative strategy in which 5ʹ OH RNAs are specifically ligated by RtcB ligase, which first guanylates the 3′ phosphate of the adapter and then ligates it directly to RNAs with 5′ OH ends. Our results demonstrate that adapter ligation to highly structured ribozyme fragments is much more efficient using the thermostable RtcB ligase from Pyrococcus horikoshii than the broadly applied Escherichia coli enzyme. Moreover, we investigated DNA, RNA and modified RNA adapters for their suitability in RtcB ligation reactions. We used the optimized RtcB-mediated ligation to produce RNA-seq libraries and captured a spiked 3ʹ twister ribozyme fragment from E. coli total RNA. This RNA-seq-based method is applicable to detect ribozyme fragments as well as other cellular RNAs with 5ʹ OH termini from total RNA.

Floxed exon (Flexon): A flexibly positioned stop cassette for recombinase-mediated conditional gene expression

Conditional gene expression is a powerful tool for genetic analysis of biological phenomena. In the widely used “lox-stop-lox” approach, insertion of a stop cassette consisting of a series of stop codons and polyadenylation signals flanked by lox sites into the 5′ untranslated region (UTR) of a gene prevents expression until the cassette is excised by tissue-specific expression of Cre recombinase. Although lox-stop-lox and similar approaches using other site-specific recombinases have been successfully used in many experimental systems, this design has certain limitations. Here, we describe the Floxed exon (Flexon) approach, which uses a stop cassette composed of an artificial exon flanked by artificial introns, designed to cause premature termination of translation and nonsense-mediated decay of the mRNA and allowing for flexible placement into a gene. We demonstrate its efficacy in Caenorhabditis elegans by showing that, when promoters that cause weak and/or transient cell-specific expression are used to drive Cre in combination with a gfp(flexon) transgene, strong and sustained expression of green fluorescent protein (GFP) is obtained in specific lineages. We also demonstrate its efficacy in an endogenous gene context: we inserted a flexon into the Argonaute gene rde-1 to abrogate RNA interference (RNAi), and restored RNAi tissue specifically by expression of Cre. Finally, we describe several potential additional applications of the Flexon approach, including more precise control of gene expression using intersectional methods, tissue-specific protein degradation, and generation of genetic mosaics. The Flexon approach should be feasible in any system where a site-specific recombination-based method may be applied.