Clostridium scindens: a human gut microbe with a high potential to convert glucocorticoids into androgens

Clostridium scindens American Type Culture Collection 35704 is capable of converting primary bile acids to toxic secondary bile acids, as well as converting glucocorticoids to androgens by side-chain cleavage. The molecular structure of the side-chain cleavage product of cortisol produced by C. scindens was determined to be 11β-hydroxyandrost-4-ene-3,17-dione (11β-OHA) by high-resolution mass spectrometry, ¹H and ¹³C NMR spectroscopy, and X-ray crystallography. Using RNA-Seq technology, we identified a cortisol-inducible (∼1,000-fold) operon (desABCD) encoding at least one enzyme involved in anaerobic side-chain cleavage. The desC gene was cloned, overexpressed, purified, and found to encode a 20α-hydroxysteroid dehydrogenase (HSDH). This operon also encodes a putative “transketolase” (desAB) hypothesized to have steroid-17,20-desmolase/oxidase activity, and a possible corticosteroid transporter (desD). RNA-Seq data suggests that the two-carbon side chain of glucocorticords may feed into the pentose-phosphate pathway and are used as a carbon source. The 20α-HSDH is hypothesized to function as a metabolic “rheostat” controlling rates of side-chain cleavage. Phylogenetic analysis suggests this operon is rare in nature and the desC gene evolved from a gene encoding threonine dehydrogenase. The physiological effect of 11β-OHAD on the host or other gut microbes is currently unknown.     

On hybridization between Theropithecus gelada and Papio anubis in the wild

Evidence is presented which suggests that Theropithecus gelada and Papio anubis may interbreed in the wild. The behavioural and ecological circumstances which give rise to this are discussed. The implications of these observations to the taxonomic status of these genera are assessed.     

Nuclear envelope proteins: Identification of lamin B subtypes

The lamins are a group of nuclear envelope proteins thought to form a structural layer at the nuclear periphery. Lamins A, B and C occur in many cell types although lamin C, a subtype of lamin A, is relatively decreased in chicken cells. A subtype of lamin B has been found in chicken cells. This subtype, called lamin B1, is slightly larger and more acidic than the quantitatively major subtype now called lamin B2. The lamin B subtypes have very similar primary sequences and share a distinctive topology. Two lamin B subtypes have not been observed in mammalian tissues but have been found in three avian tissues, erythrocytes of mature chickens and liver and erythrocytes of 11- to 13-day-old embryos. As these tissues differ widely in metabolic activity, both subtypes appear to be constitutive nuclear proteins.     

Prefoldin 5 is required for normal sensory and neuronal development in a murine model

Molecular chaperones and co-chaperones are crucial for cellular development and maintenance as they assist in protein folding and stabilization of unfolded or misfolded proteins. Prefoldin (PFDN), a ubiquitously expressed heterohexameric co-chaperone, is necessary for proper folding of nascent proteins, in particular, tubulin and actin. Here we show that a genetic disruption in the murine Pfdn5 gene, a subunit of prefoldin, causes a syndrome characterized by photoreceptor degeneration, central nervous system abnormalities, and male infertility. Our data indicate that a missense mutation in Pfdn5, may cause these phenotypes through a reduction in formation of microtubules and microfilaments, which are necessary for the development of cilia and cytoskeletal structures, respectively. The diversity of phenotypes demonstrated by models carrying mutations in different PFDN subunits suggests that each PFDN subunit must confer a distinct substrate specificity to the prefoldin holocomplex.     

Influences of soil volume and an elevated CO2 level on growth and CO2 exchange for the Crassulacean acid metabolism plant Opuntia ficus-indica

Effects of the current (38 Pa) and an elevated (74 Pa) CO₂ partial pressure on root and shoot areas, biomass accumulation and daily net CO₂ exchange were determined for Opuntia ficus-indica (L.) Miller, a highly productive Crassulacean acid metabolism species cultivated worldwide. Plants were grown in environmentally controlled rooms for 18 weeks in pots of three soil volumes (2 600, 6 500 and 26 000 cm³), the smallest of which was intended to restrict root growth. For plants in the medium-sized soil volume, basal cladodes tended to be thicker and areas of main and lateral roots tended to be greater as the CO₂ level was doubled. Daughter cladodes tended to be initiated sooner at the current compared with the elevated CO₂ level but total areas were similar by 10 weeks. At 10 weeks, daily net CO₂ uptake for the three soil volumes averaged 24% higher for plants growing under elevated compared with current CO₂ levels, but at 18 weeks only 3% enhancement in uptake occurred. Dry weight gain was enhanced 24% by elevated CO₂ during the first 10 weeks but only 8% over 18 weeks. Increasing the soil volume 10-fold led to a greater stimulation of daily net CO₂ uptake and biomass production than did doubling the CO₂ level. At 18 weeks, root biomass doubled and shoot biomass nearly doubled as the soil volume was increased 10-fold; the effects of soil volume tended to be greater for elevated CO₂. The amount of cladode nitrogen per unit dry weight decreased as the CO₂ level was raised and increased as soil volume increased, the latter suggesting that the effects of soil volume could be due to nitrogen limitations.     

The interaction of DNA-targeted 9-aminoacridine-4-carboxamide platinum complexes with DNA in intact human cells

As part of an ongoing drug development programme, this paper describes the sequence specificity and time course of DNA adduct formation for a series of novel DNA-targeted analogues of cis-diaminedichloroplatinum(II) (cisplatin) (9-aminoacridine-4-carboxamide Pt complexes) in intact HeLa cells. The sequence specificity of DNA damage caused by cisplatin and analogues in human (HeLa) cells was studied using Taq DNA polymerase and a linear amplification/polymerase stop assay. Primer extension is inhibited by a Pt-DNA adduct, and hence the sites of these lesions can be analysed on DNA sequencing gels. The repetitive alphoid DNA sequence was used as the target DNA in human cells. The 9-aminoacridine-4-carboxamide Pt complexes exhibited a markedly different sequence specificity relative to cisplatin and other analogues. The sequence specificity of the 9-aminoacridine-4-carboxamide Pt complexes is shifted away from a preference for runs of guanines. The 9-aminoacridine-4-carboxamide Pt complexes have an enhanced preference for GA dinucleotides. This is the first occasion that an altered DNA sequence specificity has been demonstrated for a cisplatin analogue in human cells. A time course of DNA damage revealed that the DNA-targeted Pt complexes, consisting of four 9-aminoacridine-4-carboxamide Pt complexes and one acridine-4-carboxamide Pt complex, damaged DNA more rapidly compared to cisplatin and non-targeted analogues. A comparison of the time taken to reach half the maximum relative intensity indicated that the DNA-targeted Pt complexes reacted approximately 4-fold faster than cisplatin and the non-targeted analogues.     

High-mass-resolution direct-tissue MALDI-FTMS reveals broad conservation of three neuropeptides (APSGFLGMRamide, GYRKPPFNGSIFamide and pQDLDHVFLRFamide) across members of seven decapod crustaean infraorders

Matrix-assisted laser desorption/ionization Fourier transform mass spectrometry (MALDI-FTMS) has become an important method for identifying peptides in neural tissues. The ultra-high-mass resolution and mass accuracy of MALDI-FTMS, in combination with in-cell accumulation techniques, can be used to advantage for the analysis of complex mixtures of peptides directly from tissue fragments or extracts. Given the diversity within the decapods, as well as the large number of extant species readily available for analysis, this group of animals represents an optimal model in which to examine phylogenetic conservation and evolution of neuropeptides and neuropeptide families. Surprisingly, no large comparative studies have previously been undertaken. Here, we have initiated such an investigation, which encompasses 32 species spanning seven decapod infraorders. Two peptides, APSGFLGMRamide and pQDLDHVFLRFamide, were detected in all species. A third peptide, GYRKPPFNGSIFamide, was detected in all species except members of the Astacidean genus Homarus, where a Val(1) variant was present. Our finding that these peptides are ubiquitously (or nearly ubiquitously) conserved in decapod neural tissues not only suggests important conserved functions for them, but also provides an intrinsic calibrant set for future MALDI-FTMS assessments of other peptides in this crustacean order.     

Effects of parental survival on clutch size decisions in fluctuating environments

Whereas in constant environments parental survival has no effect on optimal clutch size in the absence of trade-offs between juvenile and parental survival, the situation is drastically different in fluctuating environments. We consider a model in which, with respect to reproduction, parents and offspring are equivalent at the start of the next breeding season. When generations are non-overlapping, the clutch size maximizing geometric mean surviving number of offspring is optimal among all pure clutch size strategies. We prove that, as parental survival increases relative to that of the offspring, the optimal clutch size converges to the arithmetic mean maximizing clutch size (the so-called ‘Lack clutch size’). We also give a numerical procedure for calculating optimal mixed strategies and we show that, as environmental variance increases and/or parental survival decreases, mixed rather than pure strategies become optimal. Furthermore, we explain how to estimate fitness from empirical data under the assumptions of our model. This revised version was published online in July 2006 with corrections to the Cover Date.     

Changes in fat body urate synthesizing capacity during the larval-pupal transformation of the tobacco hornworm, Manduca sexta

Large quantities of uric acid or urates are deposited in the fat body of tobacco hornworms, Manduca sexta, between the larval and pupal stages in development. The cause of this increased deposition was investigated by measuring fat body urate synthesizing capacity (USC) during the larval-pupal transformation (LPT). An 85% loss in USC occurs between the late-feeding larval and newly-ecdysed pupal stages. Urate synthesizing capacity, per se, is not responsible for the increase in fat body urate deposition, as evidenced by comparable rates of urate deposition in insects whose USCs differ by a factor of three. Rather, the increased deposition is caused by an increase in substrate availability. The loss in USC is programmed in two steps. The first programmed loss occurs by the end of the feeding fifth larval instar, since hornworms ligated at the pink stripe (PS) stage and measured at the time of the larval-pupal ecdysis (LPE) exhibit an increased retention of USC relative to controls. The second programmed loss in USC occurs between PS + one and PS + two day stages in development. A single administration of 20-hydroxyecdysone to hornworms ligated at the PS stage causes a restoration of this loss in USC by PS + two days, which is further sustained until the LPE. Unexpectedly, when measured immediately after the LPE, the second programmed loss in USC can be delayed until PS + 3 days if non-ligated hornworms are daily administered 20-hydroxyecdysone. The possibility is raised that 20-hydroxyecdysone does not act alone in causing the loss in fat body USC.     

Impairment of pancreatic acinar function by reserpine in vivo and in vitro

Summary Chronic reserpine treatment of animals, an experimental model for cystic fibrosis (CF), results in generalized exocrinopathy, impaired pancreatic secretion, and decreased pancreatic content of amylase. The mechanisms of altered acinar function and decreased amylase content in both CF and the reserpine-treated rat are unknown. To examine this alteration, the rate of [³H]phenylalanine (phe) incorporation into cellular protein was determined in pancreatic acinar cells after reserpine treatment of rats in vivo (7 d) and of cells in vitro (1 to 24 h). Acinar cells isolated from control, chronic reserpine-treated, and pair-fed rats were incubated in vitro with 0, 30, 50, or 100 μM reserpine. Reserpine treatment in vitro for 24 h of acinar cells from control rats significantly decreased amylase activity (20 to 70%), an effect similar to that of reserpine treatment in vivo. In vivo, reserpine treatment decreased [³H]phe incorporation (disintegrations per minute per milligram protein) 56% in freshly isolated cells, but did not alter intracellular specific activity (disintegrations per minute per nanomole phe, SA) of [³H]phe. Reserpine treatment (30 and 50 μM) in vitro for 1 h also decreased [³H]phe incorporation by freshly isolated cells from control (53 to 85%) and pair-fed (40 to 68%) rats. Reserpine treatment for 24 h in vitro significantly decreased [³H]phe incorporation by cells from control (82 and 98%), pair-fed (80 and 95%), and chronic reserpine-treated (90 and 97%) rats as compared with cells from respective in vivo treatments cultured with no reserpine. In vitro reserpine treatment also decreased the intracellular SA of [³H]phe in freshly isolated cells from control (14 and 36%) and pair-fed (35 and 39%) rats and in cultured cells from control (11 and 86%), pair-fed (60 and 88%), and chronic reserpine-treated (49 and 76%) rats. However, these alterations of SA by reserpine did not account for the decreased incorporation of [³H]phe into acinar protein, which remained significantly lower (70 to 88%) when expressed as total phe incorporation. These results suggest (a) that reserpine acts directly on acinar cells to alter function and (b) that the ability of the pancreas to synthesize digestive enzymes may be impaired in this model of cystic fibrosis. This study was supported in part by the Cystic Fibrosis Foundation, Bethesda, MD.