Retinoscleral control of scleral remodelling in refractive development: a role for endogenous FGF-2?

AIMS: Studies in avian models of myopia have shown that refractive error development can be influenced by exogenously delivered fibroblast growth factor (FGF)-2. The present study sought to determine whether endogenous FGF-2 was associated with retinoscleral signalling or scleral remodelling during changes in refractive error in a mammalian model of myopia. METHODS: Myopia was induced in tree shrews over a 5-day period. One group of animals was then allowed 3 days of recovery from the induced myopia. Endogenous levels of FGF-2 were measured in scleral and retinal homogenates using ELISA. Real-time PCR was used to investigate scleral FGF-2 and FGF receptor (FGFR)-1 mRNA expression. RESULTS: No difference in FGF-2 content was found in posterior scleral or retinal extracts of myopic eyes (scleral -4+/-9%, retinal +23+/-17%) or recovering eyes (scleral -10+/-18%, retinal +1+/-13%), when compared with contralateral control eyes. In addition, no significant changes were found in scleral FGF-2 mRNA expression in myopic or recovering eyes (+106+/-56% and +14+/-12% respectively, P=0.21). However, FGF-2 concentration was significantly higher in anterior, relative to posterior, scleral regions in all animals (1602+/-105 vs 1030+/-50pg/mg respectively P<0.001). Expression of scleral FGFR-1 mRNA was upregulated in myopic eyes (+186+/-32%, P=0.01) but returned to control eye levels during recovery (+63+/-20%). CONCLUSIONS: The findings indicate that alterations in endogenous retinal or scleral FGF-2 levels are not associated with changes in scleral remodelling in this mammalian model of myopia. However, the reversible changes found in FGFR-1 expression in the sclera of myopic eyes mean that an indirect role for FGF-2 in the control of scleral remodelling is implicated. The anteroposterior difference found in scleral FGF-2 concentration indicates a role for this cytokine in the control of normal scleral growth and development and, presumably, eye size.     

Does water play a structural role in the folding of small nucleic acids?

Nucleic acid structure and dynamics are known to be closely coupled to local environmental conditions and, in particular, to the ionic character of the solvent. Here we consider what role the discrete properties of water and ions play in the collapse and folding of small nucleic acids. We study the folding of an experimentally well-characterized RNA hairpin-loop motif (sequence 5'-GGGC[GCAA]GCCU-3') via ensemble molecular dynamics simulation and, with nearly 500 micros of aggregate simulation time using an explicit representation of the ionic solvent, report successful ensemble folding simulations with a predicted folding time of 8.8(+/-2.0) micros, in agreement with experimental measurements of approximately 10 micros. Comparing our results to previous folding simulations using the GB/SA continuum solvent model shows that accounting for water-mediated interactions is necessary to accurately characterize the free energy surface and stochastic nature of folding. The formation of the secondary structure appears to be more rapid than the fastest ionic degrees of freedom, and counterions do not participate discretely in observed folding events. We find that hydrophobic collapse follows a predominantly expulsive mechanism in which a diffusion-search of early structural compaction is followed by the final formation of native structure that occurs in tandem with solvent evacuation.     

A circumventing role for the non-native intermediate in the folding of β-lactoglobulin

Folding experiments have suggested that some proteins have kinetic intermediates with a non-native structure. A simple G ?o model does not explain such non-native intermediates. Therefore, the folding energy landscape of proteins with non-native intermediates should have characteristic properties. To identify such properties, we investigated the folding of bovine β-lactoglobulin (βLG). This protein has an intermediate with a non-native α-helical structure, although its native form is predominantly composed of β-structure. In this study, we prepared mutants whose α-helical and β-sheet propensities are modified and observed their folding using a stopped-flow circular dichroism apparatus. One interesting finding was that E44L, whose β-sheet propensity was increased, showed a folding intermediate with an amount of β-structure similar to that of the wild type, though its folding took longer. Thus, the intermediate seems to be a trapped intermediate. The high α-helical propensity of the wild-type sequence likely causes the folding pathway to circumvent such time-consuming intermediates. We propose that the role of the non-native intermediate is to control the pathway at the beginning of the folding reaction.     

Microbial control in Asia: a bellwether for the future?

Advances and barriers faced by microbial control efforts in Asia offer instructive insights for microbial control in general. The papers in this series, which are based on plenary lectures given at the Society for Invertebrate Pathology 2006 meeting in Wuhan, China, explore the history and current status of microbial control in China, Japan, and Southeast Asia, and in doing so, bring to light the following key assumptions that deserve further examination; (1) the adoption rate of microbial control is well documented; (2) microbial control agents can compete directly with conventional insecticides; (3) microbial control agents are relatively easy and inexpensive to produce and develop; (4) patents will promote innovation and investor interest in microbial control. Alternative viewpoints are presented that can hopefully aid in future efforts to develop more safe and effective microbial control agents.     

Glycoprotein folding in the endoplasmic reticulum: a tale of three chaperones?

The endoplasmic reticulum (ER) is a major site of protein synthesis and its inside, or lumen, is a major site of protein folding. The lumen of the ER contains many folding factors and molecular chaperones, which facilitate protein folding by increasing both the rate and the efficiency of this process. Amongst the many ER folding factors, there are three components that specifically modulate the folding glycoproteins bearing N-linked carbohydrate side chains. These components are calnexin, calreticulin and ERp57, and this review focuses on the molecular basis for their capacity to influence glycoprotein folding.     

Does inositol tetrakisphosphate play a role in the receptor-mediated control of calcium mobilization?

The evidence for and against an important role for inositol 1,3,4,5 tetrakisphosphate (Ins 1,3,4,5 P4) in receptor-mediated Ca2+ mobilization is reviewed. Data obtained from patch-clamp whole-cell current recording studies on internally perfused exocrine acinar cells show that the acetylcholine (ACh)-evoked sustained increase in Ca2+-dependent K+ current caused by an increase in [Ca2+]i cannot be mimicked by internal application of inositol 1,4,5-trisphosphate (Ins 1,4,5 P3), but only by a combination of Ins 1,4,5 P3 and Ins 1,3,4,5 P4. The sustained response evoked by Ins 1,4,5 P3 + Ins 1,3,4,5 P4 is dependent on the presence of external Ca2+ as is the effect of ACh. Only those inositol trisphosphates able to evoke Ca2+ release from internal stores can support the action of Ins 1,3,4,5 P4 in evoking responses that are acutely dependent on extracellular Ca2+ (Ca2+ influx). The various arguments presented against an involvement of Ins 1,3,4,5 P4 are discussed. The main point emerging is that most studies are inadequately controlled and it is concluded that there is a strong need for whole-cell current recording studies combined with pipette fluid exchange to be carried out in many more systems. The major problem in this field is that the precise site and mechanism of action of Ins 1,3,4,5 P4 are unknown and that the pathway for Ca2+ uptake during receptor activation is inadequately defined.     

Nitric oxide's role in the heart: control of beating or breathing?

Beneficial actions of nitric oxide (NO) in failing myocardium have frequently been overshadowed by poorly documented negative inotropic effects mainly derived from in vitro cardiac preparations. NO's beneficial actions include control of myocardial energetics and improvement of left ventricular (LV) diastolic distensibility. In isolated cardiomyocytes, administration of NO increases their diastolic cell length consistent with a rightward shift of the passive length-tension relation. This shift is explained by cGMP-induced phosphorylation of troponin I, which prevents calcium-independent diastolic cross-bridge cycling and concomitant diastolic stiffening of the myocardium. Similar improvements in diastolic stiffness have been observed in isolated guinea pig hearts, in pacing-induced heart failure dogs, and in patients with dilated cardiomyopathy or aortic stenosis and have been shown to result in higher LV preload reserve and stroke work. NO also controls myocardial energetics through its effects on mitochondrial respiration, oxygen consumption, and substrate utilization. The effects of NO on diastolic LV performance appear to be synergistic with its effects on myocardial energetics through prevention of myocardial energy wastage induced by LV contraction against late-systolic reflected arterial pressure waves and through prevention of diastolic LV stiffening, which is essential for the maintenance of adequate subendocardial coronary perfusion. A drop in these concerted actions of NO on diastolic LV distensibility and on myocardial energetics could well be instrumental for the relentless deterioration of failing myocardium.     

Is counterion delocalization responsible for collapse in RNA folding?

Although energetic and phylogenetic methods have been very successful for prediction of nucleic acid secondary structures, arrangement of these secondary structure elements into tertiary structure has remained a difficult problem. Here we explore the packing arrangements of DNA, RNA, and DNA/RNA hybrid molecules in crystals. In the conventional view, the highly charged double helix will be pushed toward isolation by favorable solvation effects; interactions with other like-charged stacks would be strongly disfavored. Contrary to this expectation, we find that most of the cases analyzed ( approximately 80%) exhibit specific, preferential packing between elements of secondary structure, which falls into three categories: (i) interlocking of major grooves of two helices, (ii) side-by-side parallel packing of helices, and (iii) placement of the ribose-phosphate backbone ridge of one helix into the major groove of another. The preponderance of parallel packing motifs is especially surprising. This category is expected to be maximally disfavored by charge repulsion. Instead, it comprises in excess of 50% of all packing interactions in crystals of A-form RNA and has also been observed in crystal structures of large RNA molecules. To explain this puzzle, we introduce a novel model for RNA folding. A simple calculation suggests that the entropy gained by a cloud of condensed cations surrounding the helices more than offsets the Coulombic repulsion of parallel arrangements. We propose that these condensed counterions are responsible for entropy-driven RNA collapse, analogous to the role of the hydrophobic effect in protein folding.     

Is there a role for renal alpha 2-adrenoceptors in the pathogenesis of hypertension?

Current information suggests that alpha 2-adrenoceptors do not directly influence vascular resistance or Na reabsorption in the rat kidney. To reexamine the effects of alpha 2-agonists we used isolated rat kidneys perfused at 37.5 degrees C with precise measurement of renal artery pressure and flow. The recirculating perfusate contained pyruvate as the sole metabolic substrate which enabled us to use gluconeogenesis as an index of proximal tubular alpha 1-responses. Clonidine and guanfacine in 100 nM concentrations decreased phosphate excretion without altering Na, Cl, or K reabsorption or gluconeogenesis; 500 nM concentrations increased vascular resistance and decreased glomerular filtration rate and Na, Cl, and K excretion with no significant effect on gluconeogenesis. Prior thyroparathyroidectomy prevented the antiphosphaturic but not the antinatriuretic or vascular responses. Clonidine, an alpha 2-agonist with some alpha 1-activity, was a more potent vasoconstrictor than methoxamine or guanfacine. In the presence of prazosin (1 microM), norepinephrine (60 nM) stimulated phosphate reabsorption; norepinephrine alone did not stimulate phosphate reabsorption which indicates alpha 1-antagonism of this alpha 2-response to NE. These results and a literature review suggest that increased renal alpha 2-adrenoceptors could raise renal vascular resistance, reduce renin secretion, and antagonize parathyroid hormone effects on Pi, Ca, HCO3, and Na reabsorption to produce a low renin type of hypertension with increased proximal Na reabsorption and abnormal Ca and Pi excretion.     

River blindness: a role for parasite retinoid-binding proteins in the generation of pathology?

A new family of fatty acid- and retinoid-binding proteins has recently been identified in nematodes. These are apparently nematode specific and have very different structures and binding characteristics to their mammalian counterparts. Retinoids have important roles in vision, tissue differentiation and repair, and can profoundly affect collagen synthesis. Binding proteins released by a parasite might therefore play a part in the generation of the skin and eye pathology seen in river blindness. They might also be involved in the formation of the subcutaneous nodules induced by this parasite.     

Obstruction of the fetal urinary tract: a role for surgical intervention in utero?

Obstruction of the lower urinary tract was diagnosed by ultrasound in 11 fetuses. One pregnancy was therapeutically aborted. Four of the neonates died within 48 hours because of pronounced pulmonary hypoplasia, which is associated with obstruction of the urinary tract. The remaining six survived with adequate renal function but one, now aged 4, is obviously too small for his age. Intervention in utero for obstruction of the urinary tract is safe, but those fetuses for whom it is appropriate cannot yet be identified because of difficulties in diagnosing the condition of the whole fetus.     

Photoentrainment in mammals: a role for cryptochrome?

There is growing evidence in support of the hypothesis that, in mammals, photoreceptive tasks are segregated into those associated with creating a detailed visual image of the environment and those involved in the photic regulation of temporal biology. The hypothesis that this segregation extends to the use of different photoreceptors remains unproven, but published reports from several mammalian species that circadian photoentrainment survives a degree of retinal degeneration sufficient to induce visual blindness suggest that this may be so. This has lead to speculation that mammals might employ a dedicated 'circadian photoreceptor' distinct from the rod and cone cells of the visual system. The location and nature of this putative circadian photoreceptor has become a matter of conjecture. The latest candidates to be put forward as potential circadian photopigments are the mammalian cryptochrome proteins (CRY1 and 2), putative vitamin-B2 based photopigments. To date, published experimental evidence falls short of a definitive demonstration that these proteins form the basis of circadian photoreception in mammals. Consequently, this review aims to assess their suitability for this task in light of what we know regarding the biology of the cyrptochromes and the nature of mammalian photoentrainment.