Evolution of base composition in the insulin and insulin-like growth factor genes

The genomes of homeothermic (warm-blooded) vertebrates are mosaic interspersions of homogeneously GC-rich and GC-poor regions (isochores). Evolution of genome compartmentalization and GC-rich isochores is hypothesized to reflect either selective advantages of an elevated GC content or chromosome location and mutational pressure associated with the timing of DNA replication in germ cells. To address the present controversy regarding the origins and maintenance of isochores in homeothermic vertebrates, newly obtained as well as published nucleotide sequences of the insulin and insulin-like growth factor (IGF) genes, members of a well-characterized gene family believed to have evolved by repeated duplication and divergence, were utilized to examine the evolution of base composition in nonconstrained (flanking) and weakly constrained (introns and fourfold degenerate sites) regions. A phylogeny derived from amino acid sequences supports a common evolutionary history for the insulin/IGF family genes. In cold- blooded vertebrates, insulin and the IGFs were similar in base composition. In contrast, insulin and IGF-II demonstrate dramatic increases in GC richness in mammals, but no such trend occurred in IGF- I. Base composition of the coding portions of the insulin and IGF genes across vertebrates correlated (r = 0.90) with that of the introns and flanking regions. The GC content of homologous introns differed dramatically between insulin/IGF-II and IGF-I genes in mammals but was similar to the GC level of noncoding regions in neighboring genes. Our findings suggest that the base composition of introns and flanking regions is determined by chromosomal location and the mutational pressure of the isochore in which the sequences are embedded. An elevated GC content at codon third positions in the insulin and the IGF genes may reflect selective constraints on the usage of synonymous codons.      

Sources of error in neurotransmitter analysis

A compound having fluorescence characteristics similar to those of 5-hydroxytryptamine (5-HT), when reacted with ninhydrin, is present in several invertebrates (Fasciola hepatica, Aplysia californica, Tritonia diomedia, and Hirudo medicinalis). However, this substance is not identical with 5-HT [ Andreini, G. C., Beretta, C., Faustini, R., and Gallina, G. (1970)Experientia26, 166–167]. We confirmed these findings for Fasciola and also observed this substance in Spirometra mansonoides and mouse brain. Using chromatography and amino acid analysis we identified this substance as lysine. While 5-HT is also demonstrable in Spirometra, no 5-HT is present in Fasciola. Although epinephrine and dopamine seem to be present in Spirometra when the hydroxyindole technique of Laverty and Taylor [ Laverty, R., and Taylor, K. M. (1968)Anal. Biochem.22, 269–279] is used, neither amine is detectable in adult Spirometra by mass spectrometric analysis. A relatively high concentration of tyrosine in Spirometra could account for the apparent presence of dopamine. Therefore, lysine, tyrosine, and possibly other substances can be sources of error in the detection of biogenic amines in invertebrates.     

Using stable isotope analysis with telemetry or mark-recapture data to identify fish movement and foraging

Information about animal movements has often been inferred from stable isotope analysis (SIA), but is dependent on animals assimilating site-specific isotopic signatures via diet. This potential weakness in ecological interpretation can be overcome by using other investigative tools that provide precise information about individual movement patterns. In this paper, we demonstrate the value of combining SIA with telemetry or mark-recapture data from trapping, electrofishing and remote detection of individuals to study the movement and feeding ecology of fishes in different habitats. In a fjord lake system in Newfoundland, Canada, juvenile Atlantic salmon delayed downstream migration (smolts) or actively moved into a large lake (parr) where they foraged for periods reflecting different life history strategies. In the Miramichi River (New Brunswick, Canada), SIA provided evidence of distinct foraging habitats (tributary versus large river). By tracking fish implanted with passive integrated transponder (PIT) tags, we distinguished between movements related to foraging versus seeking cool water refugia during high temperature events. Finally, site fidelity and limited mobility of slimy sculpin, a small benthic fish, was established where δ¹³C in muscle tissue showed a progressive enrichment downstream and where a median displacement of <10 m was estimated for sculpin tagged with PIT tags. Technological improvements have permitted non-destructive tissue sampling of wild fishes for SIA, and the tagging and remote detection of animals smaller than was previously possible. These advancements and the combination of investigative tools promise new insights into animal ecology.     

Erythrocyte spectrin is an E2 ubiquitin conjugating enzyme

The involvement of red blood cell spectrin in the ubiquitination process was studied. Spectrin was found to form two ubiquitin-associated derivatives, a DTT-sensitive ubiquitin adduct and a DTT-insensitive conjugate, characteristic intermediate and final products of the ubiquitination reaction cascade. In addition to spectrin and ubiquitin, ubiquitin-activating enzyme (E1) and ATP were necessary and sufficient to form both the spectrin-ubiquitin adduct and conjugate. No exogenous ubiquitin-conjugating (E2) or ligase (E3) activities were required, suggesting that erythrocyte spectrin is an E2 ubiquitin-conjugating enzyme able to target itself. Both ubiquitin adduct and conjugate were linked to the alpha subunit of spectrin, suggesting that the ubiquitin-conjugating (UBC) domain and its target regions reside on the same subunit.     

Localisation of the Vacuolar Proton Pump (V-H+-ATPase) and Carbonic Anhydrase II in the Human Eccrine Sweat Gland

The localisation of the vacuolar proton pump (V-H+ -ATPase) and the enzyme carbonic anhydrase II (CAII) was investigated in the human eccrine sweat gland employing standard immunohistochemical techniques after antigen retrieval using microwave heat treatment and high pressure. The high-pressure antigen retrieval unmasked the presence of V-H+ -ATPase in the clear cells of the secretory coil, with a distribution similar to that previously observed for CAII. However, the dark cells were unreactive to both antibodies. In addition, heat and high-pressure antigen retrieval demonstrated the presence of CAII in the apical zone of luminal cells of the reabsorptive duct, a location not previously reported. The localisation of V-H+ -ATPase and CAII in the secretory coil clear cells suggests that the formation of HCO3- and H+ by carbonic anhydrase II and the transport of H+ by V-H+ -ATPase may play an role in sweat fluid secretion. Their presence at the apex of the duct cells indicates involvement in ductal ion reabsorption.     

Small Tails Tell Tall Tales – Intra-Individual Variation in the Stable Isotope Values of Fish Fin

Background

Fish fin is a widely used, non-lethal sample material in studies using stable isotopes to assess the ecology of fishes. However, fish fin is composed of two distinct tissues (ray and membrane) which may have different stable isotope values and are not homogeneously distributed within a fin. As such, estimates of the stable isotope values of a fish may vary according to the section of fin sampled.

Methods

To assess the magnitude of this variation, we analysed carbon (δ ¹³C), nitrogen (δ ¹⁵N), hydrogen (δ ²H) and oxygen (δ ¹⁸O) stable isotopes of caudal fin from juvenile, riverine stages of Atlantic salmon (Salmo salar) and brown trout (Salmo trutta). Individual fins were sub-sectioned into tip, mid and base, of which a further subset were divided into ray and membrane.

Findings

Isotope variation between fin sections, evident in all four elements, was primarily related to differences between ray and membrane. Base sections were¹³C depleted relative to tip (~ 1 ‰) with equivalent variation evident between ray and membrane. A similar trend was evident in δ ²H, though the degree of variation was far greater (~ 10 ‰). Base and ray sections were ¹⁸O enriched (~ 2 ‰) relative to tip and membrane, respectively. Ray and membrane sections displayed longitudinal variation in ¹⁵N mirroring that of composite fin (~ 1 ‰), indicating that variation in¹⁵N values was likely related to ontogenetic variation.

Conclusions

To account for the effects of intra-fin variability in stable isotope analyses we suggest that researchers sampling fish fin, in increasing priority, 1) also analyse muscle (or liver) tissue from a subsample of fish to calibrate their data, or 2) standardize sampling by selecting tissue only from the extreme tip of a fin, or 3) homogenize fins prior to analysis.