A road map for molecular ecology

The discipline of molecular ecology has undergone enormous changes since the journal bearing its name was launched approximately two decades ago. The field has seen great strides in analytical methods development, made groundbreaking discoveries and experienced a revolution in genotyping technology. Here, we provide brief perspectives on the main subdisciplines of molecular ecology, describe key questions and goals, discuss common challenges, predict future research directions and suggest research priorities for the next 20 years.     

Integrin alpha1beta1 controls reactive oxygen species synthesis by negatively regulating epidermal growth factor receptor-mediated Rac activation

Integrins control many cell functions, including generation of reactive oxygen species (ROS) and regulation of collagen synthesis. Mesangial cells, found in the glomerulus of the kidney, are able to produce large amounts of ROS via the NADPH oxidase. We previously demonstrated that integrin alpha1-null mice develop worse fibrosis than wild-type mice following glomerular injury and this is due, in part, to excessive ROS production by alpha1-null mesangial cells. In the present studies, we describe the mechanism whereby integrin alpha1-null mesangial cells produce excessive ROS. Integrin alpha1-null mesangial cells have constitutively increased basal levels of activated Rac1, which result in its increased translocation to the cell membrane, excessive ROS production, and consequent collagen IV deposition. Basal Rac1 activation is a direct consequence of ligand-independent increased epidermal growth factor receptor (EGFR) phosphorylation in alpha1-null mesangial cells. Thus, our study demonstrates that integrin alpha1beta1-EGFR cross talk is a key step in negatively regulating Rac1 activation, ROS production, and excessive collagen synthesis, which is a hallmark of diseases characterized by irreversible fibrosis.     

MMP-13 selective alpha-sulfone hydroxamates: identification of selective P1' amides

Continuing our interest in designing compounds preferentially potent and selective for MMP-13, we report on a series of hydroxamic acids with a flexible amide P1′ substituents. We identify an amide which spares both MMP-1 and -14, and shows >500 fold selectivity for MMP-13 versus MMP-2 and -8.     

Underwater application of quantitative PCR on an ocean mooring

The Environmental Sample Processor (ESP) is a device that allows for the underwater, autonomous application of DNA and protein probe array technologies as a means to remotely identify and quantify, in situ, marine microorganisms and substances they produce. Here, we added functionality to the ESP through the development and incorporation of a module capable of solid-phase nucleic acid extraction and quantitative PCR (qPCR). Samples collected by the instrument were homogenized in a chaotropic buffer compatible with direct detection of ribosomal RNA (rRNA) and nucleic acid purification. From a single sample, both an rRNA community profile and select gene abundances were ascertained. To illustrate this functionality, we focused on bacterioplankton commonly found along the central coast of California and that are known to vary in accordance with different oceanic conditions. DNA probe arrays targeting rRNA revealed the presence of 16S rRNA indicative of marine crenarchaea, SAR11 and marine cyanobacteria; in parallel, qPCR was used to detect 16S rRNA genes from the former two groups and the large subunit RuBisCo gene (rbcL) from Synecchococcus. The PCR-enabled ESP was deployed on a coastal mooring in Monterey Bay for 28 days during the spring-summer upwelling season. The distributions of the targeted bacterioplankon groups were as expected, with the exception of an increase in abundance of marine crenarchaea in anomalous nitrate-rich, low-salinity waters. The unexpected co-occurrence demonstrated the utility of the ESP in detecting novel events relative to previously described distributions of particular bacterioplankton groups. The ESP can easily be configured to detect and enumerate genes and gene products from a wide range of organisms. This study demonstrated for the first time that gene abundances could be assessed autonomously, underwater in near real-time and referenced against prevailing chemical, physical and bulk biological conditions.     

Thermal biology and immersion tolerance of the Beringian pseudoscorpion <Emphasis Type="Italic">Wyochernes asiaticus</Emphasis>

Wyochernes asiaticus (Arachnida: Pseudoscorpiones: Chernetidae) is a pseudoscorpion distributed across Beringia, the areas of Yukon, Alaska and Siberia that remained unglaciated at the last glacial maximum. Along with low temperatures, its streamside habitat suggests that submergence during flood events is an important physiological challenge for this species. We collected W. asiaticus in midsummer from 66.8°N Yukon Territory, Canada, and measured thermal and immersion tolerance. Wyochernes asiaticus is freeze-avoidant, with a mean supercooling point of ?6.9 °C. It remains active at low temperatures (mean critical thermal minimum, CT_min, is ?3.6 °C) and has a critical thermal maximum (CT_max) of 37.8 °C, which is lower than other arachnids and consistent with its restriction to high latitudes. Fifty per cent of W. asiaticus individuals survived immersion in oxygen-depleted water for 17 days, suggesting that this species has high tolerance to immersion during flooding events. To our knowledge, these are the first data on the environmental physiology of any pseudoscorpion and a new addition to our understanding of the biology of polar microarthropods.     

Hormonal correlates of reproductive status in the queenless ponerine ant, Streblognathus peetersi

In colonies of the queenless ant Streblognathus peetersi, dominance interactions produce a reproductive hierarchy in which one individual, the alpha, is capable of producing offspring while her subordinates remain infertile. Based on differences between behaviour and cuticular hydrocarbon profiles, the subordinates can be further divided into high and low ranking workers. Although it had been shown previously that alphas treated with a juvenile hormone analog lose their reproductive status, little was known of the endocrinological basis of dominance in this species. To elucidate the underlying endocrinology of these three ranks, we measured the individual in vitro rate of juvenile hormone (JH) production of excised corpora allata, and the ecdysteroid titer of pooled hemolymph samples. Production of JH was highest in low-ranking workers, intermediate in high rankers, and almost undetectable in alphas. Ecdysteroid titers were low for low rankers, but were more than twice as high for both high rankers and alphas. The results support the hypothesis that JH suppresses ovarian function in these queenless ants, and suggest that ecdysteroids may be responsible for stimulating vitellogenin production. The possible role of these hormones as behavioural modulators is also discussed.     

Oligomerization of signaling complexes by the multipoint binding of GRB2 to both LAT and SOS1

Receptor oligomerization is vital for activating intracellular signaling, in part by initiating events that recruit effector and adaptor proteins to sites of active signaling. Whether these distal molecules themselves oligomerize is not well appreciated. In this study, we examined the molecular interactions of the adaptor protein GRB2. In T cells, the SH2 domain of GRB2 binds phosphorylated tyrosines on the adaptor protein LAT and the GRB2 SH3 domains associate with the proline-rich regions of SOS1 and CBL. Using biochemical and biophysical techniques in conjunction with confocal microscopy, we observed that the simultaneous association of GRB2, via its SH2 and SH3 domains, with multivalent ligands led to the oligomerization of these ligands, which affected signaling. These data suggest that multipoint binding of distal adaptor proteins mediates the formation of oligomeric signaling clusters vital for intracellular signaling.     

Genetic and Biophysical Study of R Plasmids Conferring Sulfonamide Resistance in Shigella Strains Isolated in 1952 and 1956

The conjugative plasmids determining sulfonamide resistance in five Shigella strains, each isolated from a different patient, have been characterized. One S. flexneri 2a strain, isolated in 1952, harbored an fi(+) plasmid of molecular weight 53 x 10(6), which specified synthesis of F-like pili and bore determinants for sulfonamide resistance (Su) and bacteriocinogeny (Col). This plasmid was compatible with plasmids of groups F(I), F(II), I(alpha), and P. A second S. flexneri 2a strain isolated in 1952 harbored an fi(-) plasmid of molecular weight 59 x 10(6), bearing the Su determinant and compatible with all plasmids tested. This strain also harbored an fi(+) group-F(II) plasmid of molecular weight 42 x 10(6), which bore the Col determinant and specified synthesis of F-like pili. Three S. dysenteriae 2 strains isolated in 1956 carried apparently identical fi(-) plasmids of molecular weight 58 x 10(6), which bore the Su determinant, could form transconjugants in Pseudomonas but not in Proteus, and were incompatible with the P-group plasmid RP4.