Variation in heat-shock proteins among species of desert fishes (Poeciliidae, Poeciliopsis)

Analysis of the heat-shock proteins (hsps) of six closely related species of Poeciliopsis demonstrated the existence of biochemical diversity in the hsp100, hsp70, hsp60, and hsp30 protein families among species. Each species expressed five to seven hsp70-related isoforms. Constitutive 70-kD isoforms were identical among species, but four different patterns of heat-inducible isoforms were seen in these six species. Members of the hsp70 family of molecular chaperones are included among the most highly conserved proteins known, and the possibility of variation in hsp70 among closely related species has rarely been addressed. The hsp30 family is known to be less conserved than the hsp70 family, and, as expected, the Poeciliopsis hsp30 patterns showed more variation. Most of the hsp30 isoforms characteristic of a particular species were unique to that species. Hsp100 and hsp60 were identical in five of the species, but alternate isoforms were found in P. monacha. The small size and limited geographical distribution of the P. monacha population have probably contributed to the uniqueness of the monacha pattern. Two of the species were shown to acquire thermotolerance, the ability to withstand normally lethal temperatures when subjected to a gradual temperature increase. Rapid-heating protocols commonly used to establish critical thermal maxima of organisms do not include this inducible component of thermoresistance and therefore do not adequately assess an organism's capacity to withstand thermal stress.      

Cis-dominant regulatory mutations affecting the expression of GABA permease in Aspergllus nidulans

Summary In Aspergillus nidulans expression of the gabA gene, the probable structural gene for the -amino-n-butyrate (GABA) permease, is controlled by induction, via the intA gene, ammonium repression, mediated by the areA gene, and probably carbon catabolite repression. Regulatory mutations, tightly linked to gabA, were selected by reverting an areA-2 strain on GABA as nitrogen source. These mutations, gabI-1, gabI-2, and gabI-3 result in increased gabA expression and are cis-dominant in their effects on the gabA gene. Mapping data show that the regulatory mutations map on one side of all gabA^- alleles tested.     

Specular reflection of neutrons at phospholipid monolayers. Changes of monolayer structure and headgroup hydration at the transition from the expanded to the condensed phase state.

The specular neutron reflection technique has been applied for the first time to study the structure and head group hydration of a phospholipid monolayer (dimyristoylphosphatidylcholine containing negatively charged phospholipids) in the condensed and expanded state. By variation of the contrast of the subphase to that of the air it is shown that at the transition from the condensed to the expanded state the carboxyl bonds of the glycerol backbone are hydrated leading to a strong structural change of the headgroup. The total monolayer thickness in the condensed state is 22.5 +/- 1 A (tilt angle 32 +/- 6 degrees) and decreases to 19.5 +/- 1 A in the expanded state. The mean molecular volume increases from 1190 +/- 50 A3 to 1250 +/- 50 A3.     

Distribution of uterine histological changes in aged captive cheetahs (Acinonyx jubatus)

The histological effect on the felid uterus of sterilization, via ovariectomy or salpingectomy, is currently unknown. To investigate the association of ovariectomy or salpingectomy with uterine health, it is first necessary to establish if changes are distributed evenly throughout the uterus. Both laparoscopic ovariectomy and salpingectomy with concurrent sampling of the tip of the uterine horn are possible in the cheetah. Currently accepted practice for histopathological screening of the uterus utilizes four biopsy samples. It is not known whether this method accurately reflects the status of the entire uterus. In this study we histologically examined the uteri of six older cheetahs (one 7-year-old and five 10–10.5-year-old animals) via 21 tissue samples (three samples from seven different anatomical regions) per cheetah to determine overall uterine health. Although no defined lesions were detected, mild endometrial gland dilation, assumed to be of no functional consequence, was observed in multiple samples. The odds of observing this dilation was lowest in the uterine body and progressively increased in a cranial direction, being significantly higher at the tip of the uterine horns (OR = 11.5; 95% CI, 2.0-65.1; p = 0.006). This supported the reliability of sampling the tip of the uterine horn to screen for endometrial gland dilation.     

Differences in Fecal Androgen Patterns of Breeding and Nonbreeding Kori Bustards (Ardeotis kori)

To better understand breeding conditions to promote reproduction in captive kori bustards, fundamental endocrine studies measuring fecal androgen metabolites in male and female kori bustards were conducted. Feces collected weekly from males and females were analyzed for testosterone using enzyme‐linked immunoassay. Results from adult males (n = 5), adult females (n = 10), immature males (n = 10), and immature females (n = 10) revealed seasonally elevated testosterone concentrations in fertile, but not nonfertile adult males and females (P > 0.05). Adult females that were not maintained in a breeding group, or that did not produce eggs, did not demonstrate increases in testosterone compared to egg laying counterparts. In males, but not females, seasonal testosterone increases were accompanied by weight gain. Peaks in male fecal androgen metabolites ranged from 10‐ to 22‐fold higher than nonbreeding season (181.5 ± 19.1 vs. 17.0 ± 0.94 ng/g; P < 0.05). Mean breeding season values for adult males were 83.6 ± 6.1 ng/g vs. nonbreeding season values of 12.3 ± 0.73 ng/g (P < 0.05). In females, average breeding season testosterone concentrations were approximately 4‐fold higher than nonbreeding season (55.9 ± 6.0 vs. 14.5 ± 1.8 ng/g), with peaks 10‐ to 30‐fold higher. Results show that noninvasive fecal androgen metabolite analysis can provide a means of predicting fertility potential of male and female kori bustards and might be utilized to assess effects of modifying captive environments to promote reproduction in this species. Zoo Biol. 32:54‐62, 2013. © 2012 Wiley Periodicals, Inc.     

Investigating DNA Radiation Damage Using X-Ray Absorption Spectroscopy

The biological influence of radiation on living matter has been studied for years; however, several questions about the detailed mechanism of radiation damage formation remain largely unanswered. Among all biomolecules exposed to radiation, DNA plays an important role because any damage to its molecular structure can affect the whole cell and may lead to chromosomal rearrangements resulting in genomic instability or cell death. To identify and characterize damage induced in the DNA sugar-phosphate backbone, in this work we performed x-ray absorption spectroscopy at the P K-edge on DNA irradiated with either UVA light or protons. By combining the experimental results with theoretical calculations, we were able to establish the types and relative ratio of lesions produced by both UVA and protons around the phosphorus atoms in DNA.     

OmpF enhances the ability of BtuB to protect susceptible Escherichia coli cells from colicin E9 cytotoxicity

The outer membrane (OM) vitamin B(12) receptor, BtuB, is the primary receptor for E group colicin adsorption to Escherichia coli. Cell death by this family of toxins requires the OM porin OmpF but its role remains elusive. We show that OmpF enhances the ability of purified BtuB to protect bacteria against the endonuclease colicin E9, demonstrating either that the two OM proteins form the functional receptor or that OmpF is recruited for subsequent translocation of the bacteriocin. While stable binary colicin E9-BtuB complexes could be readily shown in vitro, OmpF-containing complexes could not be detected, implying that OmpF association with the BtuB-colicin complex, while necessary, must be weak and/or transient in nature.     

A 76-residue polypeptide of colicin E9 confers receptor specificity and inhibits the growth of vitamin B12-dependent Escherichia coli 113/3 cells

The mechanism by which E colicins recognize and then bind to BtuB receptors in the outer membrane of Escherichia coli cells is a poorly understood first step in the process that results in cell killing. Using N- and C-terminal deletions of the N-terminal 448 residues of colicin E9, we demonstrated that the smallest polypeptide encoded by one of these constructs that retained receptor-binding activity consisted of residues 343-418. The results of the in vivo receptor-binding assay were supported by an alternative competition assay that we developed using a fusion protein consisting of residues 1-497 of colicin E9 fused to the green fluorescent protein as a fluorescent probe of binding to BtuB in E. coli cells. Using this improved assay, we demonstrated competitive inhibition of the binding of the fluorescent fusion protein by the minimal receptor-binding domain of colicin E9 and by vitamin B12. Mutations located in the minimum R domain that abolished or reduced the biological activity of colicin E9 similarly affected the competitive binding of the mutant colicin protein to BtuB. The sequence of the 76-residue R domain in colicin E9 is identical to that found in colicin E3, an RNase type E colicin. Comparative sequence analysis of colicin E3 and cloacin DF13, which is also an RNase-type colicin but uses the IutA receptor to bind to E. coli cells, revealed significant sequence homology throughout the two proteins, with the exception of a region of 92 residues that included the minimum R domain. We constructed two chimeras between cloacin DF13 and colicin E9 in which (i) the DNase domain of colicin E9 was fused onto the T+R domains of cloacin DF13; and (ii) the R domain and DNase domain of colicin E9 were fused onto the T domain of cloacin DF13. The killing activities of these two chimeric colicins against indicator strains expressing BtuB or IutA receptors support the conclusion that the 76 residues of colicin E9 confer receptor specificity. The minimum receptor-binding domain polypeptide inhibited the growth of the vitamin B12-dependent E. coli 113/3 mutant cells, demonstrating that vitamin B12 and colicin E9 binding is mutually exclusive.