Roles of intraloops-2 and -3 and the proximal C-terminus in signalling pathway selection from the human calcium-sensing receptor

AbstractThe calcium-sensing receptor (CaSR) couples to signalling pathways via intracellular loops 2 and 3, and the C-terminus. However, the requirements for signalling are largely undefined. We investigated the impacts of selected point mutations in iL-2 (F706A) and iL-3 (L797A and E803A), and a truncation of the C-terminus (R866X) on extracellular Ca2+ (Ca2+o)-stimulated phosphatidylinositol-specific phospholipase-C (PI-PLC) and various other signalling responses. CaSR-mediated activation of PI-PLC was markedly attenuated in all four mutants and similar suppressions were observed for Ca2+o-stimulated ERK1/2 phosphorylation. Ca2+o-stimulated intracellular Ca2+ (Ca2+i) mobilization, however, was relatively preserved for the iL-2 and iL-3 mutants and suppression of adenylyl cyclase was unaffected by either E803A or R866X. The CaSR selects for specific signalling pathways via the proximal C-terminus and key residues in iL-2, iL-3

Testing Protein Leverage in Lean Humans: A Randomised Controlled Experimental Study

A significant contributor to the rising rates of human obesity is an increase in energy intake. The ‘protein leverage hypothesis’ proposes that a dominant appetite for protein in conjunction with a decline in the ratio of protein to fat and carbohydrate in the diet drives excess energy intake and could therefore promote the development of obesity. Our aim was to test the ‘protein leverage hypothesis’ in lean humans by disguising the macronutrient composition of foods offered to subjects under ad libitum feeding conditions. Energy intakes and hunger ratings were measured for 22 lean subjects studied over three 4-day periods of in-house dietary manipulation. Subjects were restricted to fixed menus in random order comprising 28 foods designed to be similar in palatability, availability, variety and sensory quality and providing 10%, 15% or 25% energy as protein. Nutrient and energy intake was calculated as the product of the amount of each food eaten and its composition. Lowering the percent protein of the diet from 15% to 10% resulted in higher (+12±4.5%, p = 0.02) total energy intake, predominantly from savoury-flavoured foods available between meals. This increased energy intake was not sufficient to maintain protein intake constant, indicating that protein leverage is incomplete. Urinary urea on the 10% and 15% protein diets did not differ statistically, nor did they differ from habitual values prior to the study. In contrast, increasing protein from 15% to 25% did not alter energy intake. On the fourth day of the trial, however, there was a greater increase in the hunger score between 1–2 h after the 10% protein breakfast versus the 25% protein breakfast (1.6±0.4 vs 25%: 0.5±0.3, p = 0.005). In our study population a change in the nutritional environment that dilutes dietary protein with carbohydrate and fat promotes overconsumption, enhancing the risk for potential weight gain

The Calcium-Sensing Receptor and the Parathyroid: Past, Present, Future

Parathyroid hormone (PTH) defends the extracellular fluid from hypocalcemia and has powerful and well-documented actions on the skeleton and renal tubular system. To achieve a satisfactory stable plasma calcium level, the secretion of PTH, and the resulting serum PTH level, is titrated carefully to the prevailing plasma ionized Ca2+ concentration via a Ca2+ sensing mechanism that mediates feedback inhibition of PTH secretion. Herein, I consider the properties of the parathyroid Ca2+ sensing mechanism, the identity of the Ca2+ sensor, the intracellular biochemical mechanisms that it controls, the manner of its integration with other components of the PTH secretion control mechanism, and its modulation by other nutrients. Together the well established, recently elucidated, and yet-to-be discovered elements of the story constitute the past, present, and future of the parathyroid and its calcium-sensing receptor (CaSR)

Thiophosphate induces apoptosis in human leukemia cell lines

Apoptosis is a form of self-regulated cell death which differs from necrosis (1). The characteristic morphological changes of apoptosis include membrane blebbing, chromatin condensation and the formation of apoptotic bodies. In addition, apoptotic cells undergo DNA double strand cleavage into fragments of multiples of about 185–200 bp resulting in a characteristic laddering pattern on agarose gel electrophoresis (2–4). Apoptosis is necessary for normal embryonic and tissue differentiation (3, 5, 6). Cell death in tumors, whether spontaneous or treatment-induced, occurs predominantly via apoptosis rather than necrosis (7, 8). Thiophosphate is a sulfur-substituted analog of orthophosphate that is an essential component of commonly used hydrolysis-resistant nucleotide analogs such as ATPgS and GTPgS as well as thiophosphorylated forms of DNA that are popular in antisense transfection studies. In a study of the growth suppressing effect of ATPgS on human promyelocytic HL-60 cells (9), we noted that thiophosphate, a breakdown product of ATPgS, independently inhibited cell growth. We investigated the nature of the thiophosphate effect on HL-60 cell proliferation and found that this effect occurs via the induction of apoptosis

Controversies in calcium-sensing in bone

Oral presentation abstrac

Invasive toxic prey may imperil the survival of an iconic giant lizard, the Komodo dragon

Although invasive species constitute a major threat to global biodiversity, the introduction of toxic prey into na ve predator faunas may become particularly destructive. An example of such an introduction was the deliberate release of the highly toxic cane toad (Bufo marinus) to the toad-free Australian continent in 1935. Na ve large Australian varanid lizards have recently been shown to suffer a massive increase in mortality (\u3e 95%) when attempting to feed on this toxic amphibian. The high susceptibility of Australian varanids to toad toxin is caused by minor mutations in the sodiumpotassium- ATPase enzyme. In the present study we show that Komodo dragons (Varanus komodoensis) have similar mutations within this enzyme as observed in Australian varanids demonstrating that dragons are extremely susceptible to toad toxin. During the last decade the black-spined toad (Bufo melanostictus) has been able to invade areas close to the five toad-free islands constituting the habitat of Komodo dragons. An invasion of highly toxic black-spined toads into dragon habitats may therefore cause similar dramatic increase in dragon mortality as recorded in Australian varanids imperiling the long-term survival of this giant and iconic lizard