Failure of prostaglandin F2α to affect LH secretion in the ovariectomized ewe

Prostaglandin F_2α (PGF_2α) when administered to ovariectomized ewes by intra-carotid infusion did not alter either the pattern of tonic LH secretion or the LH surge evoked by estradiol, indicating that, in the sheep, the luteolytic action of PGF_2α does not involve alteration of LH secretion by the pituitary gland.     

An investigation of the molecular properties and stability of intermediates of proenkephalin in isolated bovine adrenal medullary chromaffin granules

An antiserum to a synthetic peptide corresponding to residues 95-117 of bovine proenkephalin recognizes all the major intermediates of this prohormone in bovine adrenal medulla (Birch, N. P. and Christie, D. L. (1986) J. Biol. Chem. 261, 12213-12221). This antiserum enabled an investigation of the stability and molecular properties of intermediates in the processing of proenkephalin in bovine adrenal medullary chromaffin granules. Intact and hypotonic lysates of chromaffin granules were incubated at 37 degrees C and the stability of intermediates assessed by gel filtration followed by radioimmunoassay and gel electrophoresis in combination with immunoblotting. Processing was slow in intact granules compared with incubations of hypotonic lysates which resulted in the selective cleavage of an Mr 27,000 intermediate and increases in the amounts of immunoreactivity of lower molecular weight. Protease inhibitors increased the stability of the 27-kilodalton intermediate, the most effective being p-chloromercuribenzoate. Preliminary evidence was obtained for the regulation of the processing of this intermediate by soluble factors present in chromaffin granules. It appears that membrane-associated intermediates of proenkephalin are relatively stable, although analysis of soluble immunoreactivity released during the incubation of chromaffin granule membranes showed a decrease in the 27-kilodalton intermediate and increased amounts of lower molecular weight intermediates. Analysis of hypotonic lysates by two-dimensional gel-electrophoresis showed that proenkephalin intermediates exhibit significant microheterogeneity. It will be important to compare the products of proenkephalin generated by purified proteases with a putative role in the processing of this prohormone with the properties of endogenous intermediates as revealed in this study.     

A Co(III) derivative of concanavalin A

Co(III) has been stoichiometrically incorporated into jack bean concanavalin A. The Co(III) protein still possesses a binding site for an additional divalent transition metal ion which together with Ca(II) can induce the sugar binding ability. No H₂O₂ oxidation of Co(II) occurs with demetallized concanavalin A activated with Ca(II) and Co(II) unless Co(II) is present in a stoichiometric excess. Evidence is presented to indicate that kinetically stable Co(III) is bound to a completely different location than the thermodynamically stable Co(II) protein site.     

Ecological genomics of mutualism decline in nitrogen-fixing bacteria

Anthropogenic changes can influence mutualism evolution; however, the genomic regions underpinning mutualism that are most affected by environmental change are generally unknown, even in well-studied model mutualisms like the interaction between legumes and their nitrogen (N)-fixing rhizobia. Such genomic information can shed light on the agents and targets of selection maintaining cooperation in nature. We recently demonstrated that N-fertilization has caused an evolutionary decline in mutualistic partner quality in the rhizobia that form symbiosis with clover. Here, population genomic analyses of N-fertilized versus control rhizobium populations indicate that evolutionary differentiation at a key symbiosis gene region on the symbiotic plasmid (pSym) contributes to partner quality decline. Moreover, patterns of genetic variation at selected loci were consistent with recent positive selection within N-fertilized environments, suggesting that N-rich environments might select for less beneficial rhizobia. By studying the molecular population genomics of a natural bacterial population within a long-term ecological field experiment, we find that: (i) the N environment is indeed a potent selective force mediating mutualism evolution in this symbiosis, (ii) natural variation in rhizobium partner quality is mediated in part by key symbiosis genes on the symbiotic plasmid, and (iii) differentiation at selected genes occurred in the context of otherwise recombining genomes, resembling eukaryotic models of adaptation.     

Thermal unfolding studies show the disease causing F508del mutation in CFTR thermodynamically destabilizes nucleotide-binding domain 1

Misfolding and degradation of CFTR is the cause of disease in patients with the most prevalent CFTR mutation, an in-frame deletion of phenylalanine (F508del), located in the first nucleotide-binding domain of human CFTR (hNBD1). Studies of (F508del)CFTR cellular folding suggest that both intra- and inter-domain folding is impaired. (F508del)CFTR is a temperature-sensitive mutant, that is, lowering growth temperature, improves both export, and plasma membrane residence times. Yet, paradoxically, F508del does not alter the fold of isolated hNBD1 nor did it seem to perturb its unfolding transition in previous isothermal chemical denaturation studies. We therefore studied the in vitro thermal unfolding of matched hNBD1 constructs ±F508del to shed light on the defective folding mechanism and the basis for the thermal instability of (F508del)CFTR. Using primarily differential scanning calorimetry (DSC) and circular dichroism, we show for all hNBD1 pairs studied, that F508del lowers the unfolding transition temperature (T_m) by 6–7°C and that unfolding occurs via a kinetically-controlled, irreversible transition in isolated monomers. A thermal unfolding mechanism is derived from nonlinear least squares fitting of comprehensive DSC data sets. All data are consistent with a simple three-state thermal unfolding mechanism for hNBD1 ± F508del: N(±MgATP) ⇄ I_T(±MgATP) → A_T → (A_T)_n. The equilibrium unfolding to intermediate, I_T, is followed by the rate-determining, irreversible formation of a partially folded, aggregation-prone, monomeric state, A_T, for which aggregation to (A_T)_n and further unfolding occur with no detectable heat change. Fitted parameters indicate that F508del thermodynamically destabilizes the native state, N, and accelerates the formation of A_T.     

The Effects of Temperature and Body Mass on Jump Performance of the Locust Locusta migratoria

Locusts jump by rapidly releasing energy from cuticular springs built into the hind femur that deform when the femur muscle contracts. This study is the first to examine the effect of temperature on jump energy at each life stage of any orthopteran. Ballistics and high-speed cinematography were used to quantify the energy, distance, and take-off angle of the jump at 15, 25, and 35°C in the locust Locusta migratoria. Allometric analysis across the five juvenile stages at 35°C reveals that jump distance (D; m) scales with body mass (M; g) according to the power equation D = 0.35M ^{0.17±0.08 (95% CI)}, jump take-off angle (A; degrees) scales as A = 52.5M ^0.00±0.06, and jump energy (E; mJ per jump) scales as E = 1.91M ^1.14±0.09. Temperature has no significant effect on the exponent of these relationships, and only a modest effect on the elevation, with an overall Q₁₀ of 1.08 for jump distance and 1.09 for jump energy. On average, adults jump 87% farther and with 74% more energy than predicted based on juvenile scaling data. The positive allometric scaling of jump distance and jump energy across the juvenile life stages is likely facilitated by the concomitant relative increase in the total length (L _f+t; mm) of the femur and tibia of the hind leg, L _f+t = 34.9M ^0.37±0.02. The weak temperature-dependence of jump performance can be traced to the maximum tension of the hind femur muscle and the energy storage capacity of the femur''s cuticular springs. The disproportionately greater jump energy and jump distance of adults is associated with relatively longer (12%) legs and a relatively larger (11%) femur muscle cross-sectional area, which could allow more strain loading into the femur''s cuticular springs. Augmented jump performance in volant adult locusts achieves the take-off velocity required to initiate flight.     

Rapid isolation of moderate and highly polymorphic DNA fragments mapping close to WT (Wilms' tumour) and AN2 (aniridia) on chromosome 11

Summary Genes implicated in the development of Wilms' tumour (WT) and aniridia (AN2) have been localised to a subregion of band p13 on chromosome 11 by molecular and cytogenetic characterisation of WAGR syndrome patients carrying variable constitutional deletions. Polymorphic markers for the region would be valuable for linkage analysis in the familial forms of both Wilms' tumour and aniridia, as well as for studying somatic rearrangements of chromosome 11 in a variety of tumour types. Here we describe the isolation and characterisation of three frequently polymorphic arbitrary DNA fragments that map proximal to the AN2 and WT loci.