Evolution of the grass leaf by primordium extension and petiole-lamina remodeling.

The sheathing leaf found in grasses and other monocots is an evolutionary innovation, yet its origin has been a subject of long-standing debate. Here, we revisit the problem in the light of developmental genetics and computational modeling. We show that the sheathing leaf likely arose through WOX-gene-dependent extension of a primordial zone straddling concentric domains around the shoot apex. Patterned growth within this zone, oriented by two polarity fields, accounts for wild-type, mutant and mosaic grass leaf development, whereas zone contraction and growth remodeling accounts for eudicot leaf development. In contrast to the prevailing view, our results suggest that the sheath derives from petiole, whereas the blade derives from the lamina of the eudicot leaf, consistent with homologies proposed in the 19th century.

Insulin-like growth factor-2 (IGF-2) activates estrogen receptor-α and -ß via the IGF-1 and the insulin receptors in breast cancer cells.

The estrogen receptor (ER) is a primary target for breast cancer (BC) treatment. As BC progresses to estrogen-independent growth, the insulin-like growth factor-1 receptor (IGF-1R) and the ER interact in synergistic cross-talk mechanisms, which result in enhanced activation of both receptors' signaling cascades. Insulin-like growth factor-2 (IGF-2) is critical in BC progression and its actions are mediated by the IGF-1R. Our previous studies showed that IGF-2 regulates survival genes that protect the mitochondria and promote chemoresistance. In this study, we analyzed BC cells by subcellular fractionation, Western-Blot, qRT-PCR, and siRNA analysis. Our results demonstrate that IGF-2 activates ER-α and ER-ß, and modulates their translocation to the nucleus, membrane organelles, and the mitochondria. IGF-2 actions are mediated by the IGF-1R and the insulin receptor. This novel mechanism of IGF-2 synergistic cross-talk signaling with ER-α and ER-ß can promote estrogen-independent BC progression and provide new therapeutic targets for the treatment of BC patients.

Life history determines biogeographical patterns of soil bacterial communities over multiple spatial scales.

The extent to which the distribution of soil bacteria is controlled by local environment vs. spatial factors (e.g. dispersal, colonization limitation, evolutionary events) is poorly understood and widely debated. Our understanding of biogeographic controls in microbial communities is likely hampered by the enormous environmental variability encountered across spatial scales and the broad diversity of microbial life histories. Here, we constrained environmental factors (soil chemistry, climate, above-ground plant community) to investigate the specific influence of space, by fitting all other variables first, on bacterial communities in soils over distances from m to 10² km. We found strong evidence for a spatial component to bacterial community structure that varies with scale and organism life history (dispersal and survival ability). Geographic distance had no influence over community structure for organisms known to have survival stages, but the converse was true for organisms thought to be less hardy. Community function (substrate utilization) was also shown to be highly correlated with community structure, but not to abiotic factors, suggesting nonstochastic determinants of community structure are important Our results support the view that bacterial soil communities are constrained by both edaphic factors and geographic distance and further show that the relative importance of such constraints depends critically on the taxonomic resolution used to evaluate spatio-temporal patterns of microbial diversity, as well as life history of the groups being investigated, much as is the case for macro-organisms.

Characterization of promoter expression patterns derived from the Pht1 phosphate transporter genes of barley (Hordeum vulgare L.).

By comparison with dicot plant species, relatively little work has been reported on the phosphate transporter (Pht1) gene family from monocot species. Initial studies have shown that barley contains at least eight homologous genes. The promoters of six of these genes were analysed for the presence of regulatory elements potentially associated with expression specificity. In particular, the P1BS-like elements (implicated in phosphorus-regulated expression of genes in plants) was identified in all HvPht1 promoters examined. For two members of the family (HvPht1;1 and HvPht1;2), promoter fusions to beta-glucuronidase and green fluorescent protein reporter genes were constructed, transformed into rice, and the expression profiles observed. The inclusion of an intron derived from Adh1 enhanced gene expression approximately 20-fold, but did not appear to affect the specificity of expression. The HvPht1;1 and HvPht1;2 promoters showed minor differences in expression patterns but, in general, expression was observed at high levels in trichoblast cells (root hairs) and stele of the nodal root, throughout secondary roots, and at a relatively low level in leaf tissues. Under phosphorus deficiency, expression was induced by up to 5-fold. These observations are consistent with a primary role for the encoded genes in the uptake of phosphate by root hairs from soil solution and further current understanding of the mechanisms involved. The promoters also have application for providing a new resource for cereal transformation, ideally suited for driving the expression of foreign genes associated with nutrient uptake.

Extracellular secretion of Aspergillus phytase from Arabidopsis roots enables plants to obtain phosphorus from phytate.

Phosphorus (P) deficiency in soil is a major constraint for agricultural production worldwide. Despite this, most soils contain significant amounts of total soil P that occurs in inorganic and organic fractions and accumulates with phosphorus fertilization. A major component of soil organic phosphorus occurs as phytate. We show that when grown in agar under sterile conditions, Arabidopsis thaliana plants are able to obtain phosphorus from a range of organic phosphorus substrates that would be expected to occur in soil, but have only limited ability to obtain phosphorus directly from phytate. In wild-type plants, phytase constituted less than 0.8% of the total acid phosphomonoesterase activity of root extracts and was not detectable as an extracellular enzyme. By comparison, the growth and phosphorus nutrition of Arabidopsis plants supplied with phytate was improved significantly when the phytase gene (phyA) from Aspergillus niger was introduced. The Aspergillus phytase was only effective when secreted as an extracellular enzyme by inclusion of the signal peptide sequence from the carrot extensin (ex) gene. A 20-fold increase in total root phytase activity in transgenic lines expressing ex::phyA resulted in improved phosphorus nutrition, such that the growth and phosphorus content of the plants was equivalent to control plants supplied with inorganic phosphate. These results show that extracellular phytase activity of plant roots is a significant factor in the utilization of phosphorus from phytate and indicate that opportunity exists for using gene technology to improve the ability of plants to utilize accumulated forms of soil organic phosphorus.

Spatial knowledge acquisition from maps and from navigation in real and virtual environments.

In this study, the nature of the spatial representations of an environment acquired from maps, navigation, and virtual environments (VEs) was assessed. Participants first learned the layout of a simple desktop VE and then were tested in that environment. Then, participants learned two floors of a complex building in one of three learning conditions: from a map, from direct experience, or by traversing through a virtual rendition of the building. VE learners showed the poorest learning of the complex environment overall, and the results suggest that VE learners are particularly susceptible to disorientation after rotation. However, all the conditions showed similar levels of performance in learning the layout of landmarks on a single floor. Consistent with previous research, an alignment effect was present for map learners, suggesting that they had formed an orientation-specific representation of the environment. VE learners also showed a preferred orientation, as defined by their initial orientation when learning the environment. Learning the initial simple VE was highly predictive of learning a real environment, suggesting that similar cognitive mechanisms are involved in the two learning situations.

A comparison of methods for estimating directions in egocentric space.

A central issue for researchers of human spatial knowledge, whether focused on perceptually guided action or cognitive-map acquisition, is knowledge of egocentric directions, directions from the body to objects and places. Several methods exist for measuring this knowledge. We compared two particularly important methods, manual pointing with a dial and whole-body rotation (body heading), under various conditions of sensory or memory access to targets. In two experiments, blindfolded body rotation resulted in the greatest variability of performance (variable error), while the manual dial resulted in greater consistent bias (constant error). The variability of performance with body rotation was no greater than that of the dial when subjects' memory loads for directions to targets was reduced by allowing them to peek at targets in between trials, point to concurrent auditory targets, or point with their eyes open. In both experiments, errors with the manual dial were greater for directions to targets that were further from the closest orthogonal axis (ahead, behind, right, left), while errors with body rotation with restricted perceptual access were greater for directions to targets that were further from an axis straight ahead of subjects. This suggests that the two methods will produce evidence of different organizational frameworks for egocentric spatial knowledge. Implications for the structures and processes that underlie egocentric spatial knowledge, and are involved in estimating directions, are discussed, as is the value of decomposing absolute errors into variable and constant errors.

Soil isolates of Pseudomonas spp. that utilize inositol phosphates.

Soil bacteria that utilize inositol hexaphosphate (IHP) were isolated from a range of soils using defined selection media. An analysis of 200 randomly selected isolates indicated that less than 0.5% of the culturable population of soil bacteria were capable of using IHP as a sole source of C and P. From a further 238 isolates obtained from enrichment culture, four unique organisms (identified by randomly amplified polymorphic DNA-polymerase chain reaction) were selected and characterized for their ability to specifically utilize IHP. These four organisms were putatively identified as either fluorescent Pseudomonas spp. (P. putida CCAR53 and CCAR59) or nonfluorescent Pseudomonas spp. (P. mendocina CCAR31 and CCAR60) as determined by partial DNA sequence analysis of 16S rRNA genes. The fluorescent Pseudomonas strains exhibited marked phytase activity and liberated up to 81% of the phosphate from IHP either in the absence or presence of arabinose as an additional C source. The nonfluorescent strains also exhibited an ability to liberate Pi from IHP but were effective only in the presence of added arabinose. Strains CCAR59 and CCAR60 could effectively utilize either Na-IHP or Ca-IHP at pH 7.0, whereas only strain CCAR59 could grow and utilize these substrates at pH 5.0.

Studies of the Physiological and Genetic Basis of Acid Tolerance in Rhizobium leguminosarum biovar trifolii.

Acid-tolerant Rhizobium leguminosarum biovar trifolii ANU1173 was able to grow on laboratory media at a pH as low as 4.5. Transposon Tn5 mutagenesis was used to isolate mutants of strain ANU1173, which were unable to grow on media at a pH of less than 4.8. The acid-tolerant strain ANU1173 maintained a near-neutral intracellular pH when the external pH was as low as 4.5. In contrast, the acid-sensitive mutants AS25 and AS28 derived from ANU1173 had an acidic intracellular pH when the external pH was less than 5.5. The acid-sensitive R. leguminosarum biovar trifolii ANU794, which was comparatively more sensitive to low pH than mutants AS25 and AS28, showed a more acidic internal pH than the two mutants when the three strains were exposed to medium buffered at a pH of less than 5.5. The two acid-sensitive mutants had an increased membrane permeability to protons but did not change their proton extrusion activities. However, the acid-sensitive strain ANU794 exhibited both a higher membrane permeability to protons and a lower proton extrusion activity compared with the acid-tolerant strain ANU1173. DNA hybridization analysis showed that mutants AS25 and AS28 carried a single copy of Tn5 located in 13.7-kb (AS25) and 10.0-kb (AS28) EcoRI DNA fragments. The wild-type DNA sequences spanning the mutation sites of mutants AS25 and AS28 were cloned from genomic DNA of strain ANU1173. Transfer of these wild-type DNA sequences into corresponding Tn5-induced acid-sensitive mutants, respectively, restored the mutants to their acid tolerance phenotypes. Mapping studies showed that the AS25 locus was mapped to a 5.6-kb EcoRI-BamHI megaplasmid DNA fragment, whilst the AS28 locus was located in an 8.7-kb BglII chromosomal DNA fragment.

Construction of an Acid-Tolerant Rhizobium leguminosarum Biovar Trifolii Strain with Enhanced Capacity for Nitrogen Fixation.

Strain ANU1173 is an acid-tolerant Rhizobium leguminosarum biovar trifolii strain that is able to nodulate subterranean clover plants growing in agar culture at pH 4.4 At pH 6.5, its symbiotic effectiveness in association with Trifolium subterraneum cv. Mt. Barker was 80% relative to that of strain ANU794, a Sm derivative of the commercial inoculant R. leguminosarum bv. trifolii TA1. Strain ANU1173 contained four indigenous megaplasmids, the smallest of these being the symbiotic (Sym) plasmid. The critical pH requirement for growth of strain ANU1173 in laboratory media was shown not to be associated with this plasmid. When the Sym plasmid of strain ANU1173(pSym-1173) was mobilized into a Nod strain of R. leguminosarum bv. viciae, the plasmid conferred to the transconjugant a level of symbiotic effectiveness in association with T. subterraneum that was similar to that observed with ANU1173. The symbiotic effectiveness of strain ANU1173 was improved by first curing pSym-1173 (generating strain ANU1184) and replacing it with another R. leguminosarum bv. trifolii Sym plasmid, pBR1AN. Subterranean clover plants inoculated with strain ANU1184 (pBR1AN) exhibited a 35 or 53% increase in acetylene reduction activity and a 20 or 17% increase in dry weight when grown at pH 6.5 and pH 4.4, respectively, compared with plants inoculated with strain ANU1173 and grown under the same pH conditions. It was further shown that pBR1AN was stably maintained in strain ANU1184 under free-living and symbiotic conditions. These results indicate that it is possible to construct an acid-tolerant strain of R. leguminosarum bv. trifolii with an enhanced capacity for nitrogen fixation.

Expression of Nodulation Genes in Rhizobium leguminosarum biovar trifolii Is Affected by Low pH and by Ca and Al Ions.

Early stages in the infection of leguminous plants by Rhizobium spp. are restricted at low pH and are further influenced by the presence of Ca and Al ions. In the experiments reported here, transcriptional and translational fusions of the Escherichia coli lacZ gene to Rhizobium leguminosarum biovar trifolii nodulation (nod) genes were used to investigate the effects of pH and of Ca and Al ions on nod gene expression. The regulatory nodD gene in R. leguminosarum biovar trifolii was constitutively expressed at a range of pH levels between 4.8 and 6.5, and expression was not affected by the addition of 22.5 muM Al or 1,250 muM Ca. Induction of expression of nodA, nodF, and region II nodulation genes in the presence of 5 x 10 M 7,4'-dihydroxyflavone was restricted at a pH of <5.7 and was extremely poor at pH 4.8. Induction of nodA expression was further restricted by 22.5 muM Al over a range of pH levels but was increased in the presence of Ca. The addition of Ca, however, only slightly alleviated the Al-mediated inhibition of nodA induction. Induction of expression of nodA was equally sensitive to low pH in three strains of R. leguminosarum biovar trifolii (ANU845, ANU815, and ANU1184), which exhibited contrasting growth abilities in solution culture at a pH of <5.0. Aluminum, however, differentially affected the induction of nodA in these three strains, with the most Al-tolerant strain for growth being the most Al-sensitive strain for nod gene induction. Poor induction of expression of nodulation genes in R. leguminosarum biovar trifolii was considered to be an important factor contributing to the acid-sensitive step of legume root infection.

The role of a (Ca2+ + Mg2+)-ATPase of the rough endoplasmic reticulum in regulating intracellular Ca2+ during cholinergic stimulation of rat pancreatic acini.

Rough endoplasmic reticulum membranes, purified from isolated rat pancreatic acini stimulated by carbachol, had a decreased Ca2+ content and increased (Ca2+ + Mg2+)-ATPase activity. Ca2+ was regained and ATPase activity reduced to control levels only after blockade by atropine. The (Ca2+ + Mg2+)-ATPase was activated by free Ca2+ (half-maximal at 0.17 microM; maximal at 0.7 microM) over the concentration range which occurs in the cell cytoplasm. Pretreatment with EGTA, at a high concentration (5 mM), inhibited ATPase activity which, our results suggest, was due to removal of a bound activator such as calmodulin. The rate of (Ca2+ + Mg2+)-ATPase actively declined during the 10-min period over which maximal active accumulation of Ca2+ by membrane vesicles occurs. In the presence of ionophore A23187, which released actively accumulated Ca2+ and stimulated the (Ca2+ + Mg2+)-ATPase, this time-dependent decline in activity was not observed. Our data provide evidence that the activity of the Ca2+-transporting ATPase of the rough endoplasmic reticulum is regulated by both extra and intravesicular Ca2+ and is consistent with a direct role of this enzyme in the release and uptake of Ca2+ during cholinergic stimulation of pancreatic acinar cells.

Solid haemangioblastomas of the posterior fossa: radiological features and results of surgery.

Cerebellar haemangioblastomas are benign neoplasms which have been reported to have a favourable prognosis. The authors report the experience of Atkinson Morley's Hospital, London with 14 cases of solid haemangioblastoma since the introduction of the CT scan. Death or a poor result occurred in 50% of cases. Incomplete excision carried a high risk of complications. The CT scan appearances were too non specific to make an accurate preoperative diagnosis, the lesions usually being mistaken for meningiomas. Vertebral angiography can be of considerable help in identifying their vascular anatomy.

The value of computerized tomography in aneurysmal subarachnoid hemorrhage. The concept of the CT score.

Of 256 patients with aneurysmal subarachnoid hemorrhage, 131 underwent computerized tomographic (CT) scanning within 7 days of the ictus. These scans were analyzed in order to assess the quantity of blood in the main subarachnoid cisterns and cerebral fissures. The method of quantification used recognized the horizontal and vertical components of the largest clot visible on the CT scan and expressed this as the "CT score." Angiographic vasospasm was assessed and graded, based on reduction in the caliber of the major cerebral vessels. The CT score was then compared to 1) the incidence of angiographic vasospasm, 2) the clinical course, and 3) the eventual outcome. Of the patients who showed no blood on the initial CT scan, 87% were admitted in good clinical grades, whereas among patients with higher CT scores the number admitted in poor clinical grades increased. The degree of angiographic vasospasm did not relate as closely as the CT score to the clinical grade on admission or to the subsequent clinical course. The final outcome was assessed on follow-up review, and those acquiring neurological deficits from ischemic neurological dysfunction (IND) were identified. Ninety percent of patients with no blood on the CT scan (CT score 0) had a good outcome, while 5% sustained the effects of IND. The incidence of IND gradually increased with a rise in the CT score until, with scores of 8 and above, 90% of patients suffered the ill effects of IND. The CT score proved to be a simple yet accurate prognostic indicator of the outcome of IND.

Calcium-ion-transporting activity in two microsomal subfractions from rat pancreatic acini. Modulation by carbamylcholine.

Two microsomal subfractions from isolated rat pancreatic acini were produced by centrifugation through a discontinuous sucrose density gradient and characterized by biochemical markers. The denser fraction ( SF2 ) was a highly purified preparation of rough endoplasmic reticulum; the less-dense fraction ( SF1 ) was heterogeneous and contained Golgi, endoplasmic reticulum and plasma membranes. 45Ca2+ accumulation in the presence of ATP and its rapid release after treatment with the bivalent-cation ionophore A23187 were demonstrated in both fractions. The pH optimum for active 45Ca2+ uptake was approx. 6.8 for the rough endoplasmic reticulum ( SF2 ) and approx. 7.5 for SF1 . Initial rate measurements were used to determine the affinity of the rough-endoplasmic-reticulum uptake system for free Ca2+. An apparent Km of 0.16 +/- 0.06 microM and Vmax. of 21.5 +/- 5.6 nmol of Ca2+/min per mg of protein were obtained. 45Ca2+ uptake by SF1 was less sensitive to Ca2+, half-maximal uptake occurring at 1-2 microM-free Ca2+. When fractions were prepared from isolated acini stimulated with 3 microM-carbamylcholine, 45Ca2+ uptake was increased in the rough endoplasmic reticulum. The increased uptake was due to a higher Vmax. with no significant change in Km. No effect was observed on 45Ca2+ uptake by SF1 . In conclusion, two distinct non-mitochondrial, ATP-dependent calcium-uptake systems have been demonstrated in rat pancreatic acini. One of these is located in the rough endoplasmic reticulum, but the precise location of the other has not been determined. We have shown that the Ca2+-transporting activity in the rough endoplasmic reticulum may have an important role in maintaining the cytosolic free Ca2+ concentration in resting acinar cells and is involved in Ca2+ movements which occur during stimulation of enzyme secretion.

Clinical correlation of urodynamic findings in patients with localized partial lesions of the spinal cord and cauda equina.

Urodynamic studies were undertaken in 70 patients with incomplete, confirmed, localized lesions involving the spinal cord and cauda equina, irrespective of clinically evident bladder disturbances. Both detrusor and urethral function were simultaneously evaluated and correlated with neurological deficits and symptoms of lower urinary tract dysfunction. It was evident that in those patients with localized spinal cord lesions, lower urinary tract disturbances were closely associated with disturbances of pinprick sensation. Clinically silent bladder and sphincter dysfunction was encountered in 14% of the patients, and the number of asymptomatic patients was significantly higher among those with upper motor neuron lesions. Incontinence was associated predominantly with conus lesions; however, the other symptoms had no relationship to the site of the lesion. Symptoms such as urgency, incontinence, and nocturnal incontinence were associated with detrusor hyperactivity, and urinary retention was associated with urethral overactivity and dyssynergia. The urodynamic findings could explain the pathogenesis of the symptoms in most instances, but were not necessarily related to them. It is concluded that the pathways subserving lower-urinary-tract function are closely associated with the pathways subserving pinprick sensation, and that a lesion along the spinal axis can affect the bladder, sphincter, or both in a variety of ways, thus leading to the widely variable clinical findings associated with neurogenic lower urinary tract dysfunction.

The long-term outcome in patients with multiple aneurysms. Incidence of late hemorrhage and implications for treatment of incidental aneurysms.

The proper treatment of multiple and incidental aneurysms remains controversial because the long-term result of different modes of management is unclear. This report evaluates the long-term outcome (follow-up period averaged 7.7 years) in 182 patients with multiple aneurysms who suffered a subarachnoid hemorrhage (SAH) to document the incidence of late bleeding. Of the 182 patients, 132 were treated by bed rest and 50 by surgery (craniotomy) directed at only the ruptured aneurysm. Seventy of the patients with bed rest were alive after 6 months. Twenty-one of these conservatively treated patients (30%) suffered a late hemorrhage, which is equal to the previously reported average yearly rebleed rate (3%) with a single aneurysm of the anterior circulation. There was no evidence that a previously intact aneurysm had ruptured in SAH patients treated with bed rest, indicating that late hemorrhage was due to rerupture from the original aneurysm. Patients who were hypertensive and who had a large aneurysm had an increased risk of late rehemorrhage. A linear discriminant analysis was developed to predict late rebleeding. The fate of intact aneurysms was evaluated by following patients with multiple aneurysms treated by craniotomy directed only at the ruptured aneurysm. Of the 50 craniotomy patients, 38 were alive after 6 months. In this group, the minimal risk of rupture of an intact aneurysm is approximately 1% per year. The presence of hypertension increased the risk of late hemorrhage. In conclusion, patients with multiple untreated aneurysms managed by bed rest have a late rehemorrhage rate equal to that observed in patients with a single cerebral aneurysm; the data indicate that rupture of intact aneurysms is not insignificant.

Urethral function in neurological disorders of the lower urinary tract.

Simultaneous measurements of intravesical pressure, urethral pressure, and electromyographic activity of perianal muscles were performed during bladder filling and voiding in 56 patients with neurogenic lower urinary tract dysfunction. In 21 patients simultaneous measurements were taken of the EMG activity of the striated-muscle external urethral sphincter. During bladder filling the urethral pressure was found to correspond to the EMG activity of both muscles only in those patients with normal urethral pressure. During voiding the EMG activity of neither muscle corresponded well with changes in urethral pressure. The EMG activity of perianal muscles corresponded to the activity of the external urethral sphincter during filling of the bladder but not during voiding.

Significance of detrusor compliance in patients with localized partial lesions of the spinal cord and cauda equina.

Detrusor compliance was measured by cystometry irrespective of any symptoms of lower urinary trace dysfunction in 70 patients with localized, partial, surgically treatable lesions of the spinal cord and cauda equina. Compliance was found to be normal in all patients with upper motor neuron lesions; reduced compliance was encountered only in those with lower motor neuron lesions. Reduced compliance was closely associated with lesions of the conus medullaris and with detrusor hyperreflexia, and it is therefore likely that the mechanism of compliance involves a significant neural component. Incontinence, particularly nocturnal incontinence, was encountered more frequently among those with reduced compliance than in those with normal compliance, whereas retention of urine was comparatively rare with reduced compliance.