Adaptation to variation in growing season length in arctic populations of Saxifraga oppositifolia L.

Summary

At high latitudes Saxifraga oppositifolia exists with distinct ecotypes adapted to differences in growing season length. In areas with late snow-lie and cold, wet soils increased metabolic rates and rapid shoot production compensate for ultra-short growing seasons but do not conserve carbohydrate or water for adverse periods. An opposing strategy is evident in ecotypes living in sites with an earlier resumption of growth where soils are warmer and drier and the growing season longer. Here metabolic rates are lower and result in a greater ability to conserve both carbohydrate and water. The existence of opposing strategies for survival in warm and cold habitats, suggests that even in the minimal thermal conditions of the high Arctic a high degree of biodiversity gives the species as a whole a wide ecological amplitude. This degree of diversity not only increases the sites in which the species can survive but confers an ability to adapt to climate change by changing ecotype frequencies to accommodate climatic fluctuations. This facility may have contributed to the survival of populations of this and other species in the high Arctic during the Weichselian glaciation.     

DNA fingerprinting analysis of Booroola pedigrees: a search for linkage to the Booroola gene

Seven minisatellite probes from a variety of sources were used to analyse 11 paternal half-sib families in which the Booroola gene was segregating. A total of 402 bands that showed segregation in the pedigrees were examined for linkage to the Booroola gene. None of the bands showed segregation with the Booroola gene. The most likely evidence for a linked band was produced by the HaRas HVR probe in Family 902 (=0.0; LOD 2.3). The conclusion, however, is that the minisatellite probes used in this study could not be used as markers for the Booroola gene. The study highlighted problems associated with the use of minisatellite probes in linkage studies in half-sib families. The complex banding patterns found on fingerprinting gels was a major source of scoring error. In a few cases both of the sire's alleles could be identified at a particular locus, but in most cases only one of the alleles could be identified. For the most part, the bands had to be treated as dominant alleles. The contribution of dam alleles to the banding pattern could only be estimated. There was an indication that minisatellite loci in sheep are clustered in particular regions of the sheep genome as the rate at which bands segregated with each other was higher than one would expect from loci randomly distributed throughout the genome.     

An ultrasensitive high-throughput electrochemiluminescence immunoassay for the Cdc42-associated protein tyrosine kinase ACK1

Several drugs inhibiting protein kinases have been launched successfully, demonstrating the attractiveness of protein kinases as therapeutic targets. Functional genomics research within both academia and industry has led to the identification of many more kinases as potential drug targets. Although a number of well-known formats are used for measuring protein kinase activity, some less well-characterized protein kinases identified through functional genomics present particular challenges for existing assay formats when there is limited knowledge of the endogenous substrates or activation mechanisms for these novel kinase targets. This is especially the case when a very sensitive assay is required to differentiate often highly potent inhibitors developed by late-stage medicinal chemistry programs. ACK1 is a non-receptor tyrosine kinase that has been shown to be involved in tumorigenesis and metastasis. Here we describe the development of an extremely sensitive high-throughput assay for ACK1 capable of detecting 240 fmol per well of the kinase reaction product employing a BV-tag-based electrochemiluminescence assay. This assay is universally applicable to protein tyrosine kinases using a BV-tag-labeled monoclonal antibody against phosphotyrosine. Furthermore, this assay can be extended to the evaluation of Ser/Thr kinases in those cases where an antibody recognizing the phospho-product is available.     

ScrepYard: An online resource for disulfide‐stabilized tandem repeat peptides

Receptor avidity through multivalency is a highly sought‐after property of ligands. While readily available in nature in the form of bivalent antibodies, this property remains challenging to engineer in synthetic molecules. The discovery of several bivalent venom peptides containing two homologous and independently folded domains (in a tandem repeat arrangement) has provided a unique opportunity to better understand the underpinning design of multivalency in multimeric biomolecules, as well as how naturally occurring multivalent ligands can be identified. In previous work, we classified these molecules as a larger class termed secreted cysteine‐rich repeat‐proteins (SCREPs). Here, we present an online resource; ScrepYard, designed to assist researchers in identification of SCREP sequences of interest and to aid in characterizing this emerging class of biomolecules. Analysis of sequences within the ScrepYard reveals that two‐domain tandem repeats constitute the most abundant SCREP domain architecture, while the interdomain “linker” regions connecting the functional domains are found to be abundant in amino acids with short or polar sidechains and contain an unusually high abundance of proline residues. Finally, we demonstrate the utility of ScrepYard as a virtual screening tool for discovery of putatively multivalent peptides, by using it as a resource to identify a previously uncharacterized serine protease inhibitor and confirm its predicted activity using an enzyme assay.     

Ehd4 is required to attain normal prepubertal testis size but dispensable for fertility in male mice

The four highly homologous members of the C‐terminal EH domain‐containing (EHD) protein family (EHD1‐4) regulate endocytic recycling. To delineate the role of EHD4 in normal physiology and development, mice with a conditional knockout of the Ehd4 gene were generated. PCR of genomic DNA and Western blotting of organ lysates from Ehd4^−/− mice confirmed EHD4 deletion. Ehd4^−/− mice were viable and born at expected Mendelian ratios; however, males showed a 50% reduction in testis weight, obvious from postnatal day 31. An early (Day 10) increase in germ cell proliferation and apoptosis and a later increase in apoptosis (Day 31) were seen in the Ehd4^−/− testis. Other defects included a progressive reduction in seminiferous tubule diameter, dysregulation of seminiferous epithelium, and head abnormalities in elongated spermatids. As a consequence, lower sperm counts and reduced fertility were observed in Ehd4^−/− males. Interestingly, EHD protein expression was seen to be temporally regulated in the testis and EHD4 levels peaked between days 10 and 15. In the adult testis, EHD4 was highly expressed in primary spermatocytes and EHD4 deletion altered the levels of other EHD proteins in an age‐dependent manner. We conclude that high levels of EHD1 in the adult Ehd4^−/− testis functionally compensate for lack of EHD4 and prevents the development of severe fertility defects. Our results suggest a role for EHD4 in the proper development of postmitotic and postmeiotic germ cells and implicate EHD protein‐mediated endocytic recycling as an important process in germ cell development and testis function. genesis 48:328–342, 2010. © 2010 Wiley‐Liss, Inc.     

(+)-Isoshinanolone and 2-methylbenzofuran-4-carbaldehyde from the fish-stunning plant Habropetalum dawei

(+)-Isoshinanolone was isolated from an aqueous extract of the leaves of Habropetalum dawei. After isolation of (+)-isoshinanolone, the aqueous extract of the leaves was acidified, refluxed distilled to give a new benzofuran, 2-methylbenzofuran-4-carbaldehyde. (+)-Isoshinanolone was found to have fish-stunning activity.     

A radioimmunoassay using labeled antibody for polyhedron protein from a nuclear polyhedrosis virus of Wiseana cervinata

A radioimmunoassay (RIA) for polyhedron protein based on precipitation of the antibody-antigen complex by sodium acetate buffer, pH 5.0, was studied. The effects of varying assay conditions and of using [³H]acetate- or ¹²⁵I-labeled antibodies were examined. The RIA could detect specific differences between the polyhedron protein from the nuclear polyhedrosis viruses of Bombyx mori and Wiseana cervinata at the level of 3 × 10⁴ polyhedral/ml when ¹²⁵I-labeled antibodies were used. This procedure could provide the basis for making serological comparisons of polyhedron proteins and for detecting viruses in samples such as soil and bird feces.