Crif1 Deficiency Reduces Adipose OXPHOS Capacity and Triggers Inflammation and Insulin Resistance in Mice

Impaired mitochondrial oxidative phosphorylation (OXPHOS) has been proposed as an etiological mechanism underlying insulin resistance. However, the initiating organ of OXPHOS dysfunction during the development of systemic insulin resistance has yet to be identified. To determine whether adipose OXPHOS deficiency plays an etiological role in systemic insulin resistance, the metabolic phenotype of mice with OXPHOS–deficient adipose tissue was examined. Crif1 is a protein required for the intramitochondrial production of mtDNA–encoded OXPHOS subunits; therefore, Crif1 haploinsufficient deficiency in mice results in a mild, but specific, failure of OXPHOS capacity in vivo. Although adipose-specific Crif1-haploinsufficient mice showed normal growth and development, they became insulin-resistant. Crif1-silenced adipocytes showed higher expression of chemokines, the expression of which is dependent upon stress kinases and antioxidant. Accordingly, examination of adipose tissue from Crif1-haploinsufficient mice revealed increased secretion of MCP1 and TNFα, as well as marked infiltration by macrophages. These findings indicate that the OXPHOS status of adipose tissue determines its metabolic and inflammatory responses, and may cause systemic inflammation and insulin resistance.     

A functional interaction between chromogranin B and the inositol 1,4,5-trisphosphate receptor/Ca2+ channel

Chromogranins A and B (CGA and CGB) are high capacity, low affinity calcium (Ca2+) storage proteins found in many cell types most often associated with secretory granules of secretory cells but also with the endoplasmic reticulum (ER) lumen of these cells. Both CGA and CGB associate with inositol 1,4,5-trisphosphate receptor (InsP3R) in a pH-dependent manner. At an intraluminal pH of 5.5, as found in secretory vesicles, both CGA and CGB bind to the InsP3R. When the intraluminal pH is 7.5, as found in the ER, CGA totally dissociates from InsP3R, whereas CGB only partially dissociates. To investigate the functional consequences of the interaction between the InsP3R and CGB monomers or CGA/CGB heteromers, purified mouse InsP3R type I were fused to planar lipid bilayers and activated by 2 microM InsP3. In the presence of luminal CGB monomers or CGA/CGB heteromers the InsP3R/Ca2+ channel open probability and mean open time increased significantly. The channel activity remained elevated when the pH was changed to 7.5, a reflection of CGB binding to the InsP3R even at pH 7.5. These results suggest that CGB may play an important modulatory role in the control of Ca2+ release from the ER. Furthermore, the difference in the ability of CGA and CGB to regulate the InsP3R/Ca2+ channel and the variability of CGA/CGB ratios could influence the pattern of InsP3-mediated Ca2+ release.     

Role of Cbl-PI3K Interaction during Skeletal Remodeling in a Murine Model of Bone Repair

Mice in which Cbl is unable to bind PI3K (YF mice) display increased bone volume due to enhanced bone formation and repressed bone resorption during normal bone homeostasis. We investigated the effects of disrupted Cbl-PI3K interaction on fracture healing to determine whether this interaction has an effect on bone repair. Mid-diaphyseal femoral fractures induced in wild type (WT) and YF mice were temporally evaluated via micro-computed tomography scans, biomechanical testing, histological and histomorphometric analyses. Imaging analyses revealed no change in soft callus formation, increased bony callus formation, and delayed callus remodeling in YF mice compared to WT mice. Histomorphometric analyses showed significantly increased osteoblast surface per bone surface and osteoclast numbers in the calluses of YF fractured mice, as well as increased incorporation of dynamic bone labels. Furthermore, using laser capture micro-dissection of the fracture callus we found that cells lacking Cbl-PI3K interaction have higher expression of Osterix, TRAP, and Cathepsin K. We also found increased expression of genes involved in propagating PI3K signaling in cells isolated from the YF fracture callus, suggesting that the lack of Cbl-PI3K interaction perhaps results in enhanced PI3K signaling, leading to increased bone formation, but delayed remodeling in the healing femora.     

Tarantula toxins interact with voltage sensors within lipid membranes

Voltage-activated ion channels are essential for electrical signaling, yet the mechanism of voltage sensing remains under intense investigation. The voltage-sensor paddle is a crucial structural motif in voltage-activated potassium (K(v)) channels that has been proposed to move at the protein-lipid interface in response to changes in membrane voltage. Here we explore whether tarantula toxins like hanatoxin and SGTx1 inhibit K(v) channels by interacting with paddle motifs within the membrane. We find that these toxins can partition into membranes under physiologically relevant conditions, but that the toxin-membrane interaction is not sufficient to inhibit K(v) channels. From mutagenesis studies we identify regions of the toxin involved in binding to the paddle motif, and those important for interacting with membranes. Modification of membranes with sphingomyelinase D dramatically alters the stability of the toxin-channel complex, suggesting that tarantula toxins interact with paddle motifs within the membrane and that they are sensitive detectors of lipid-channel interactions.     

Requirement of hCenexin for proper mitotic functions of polo-like kinase 1 at the centrosomes

Outer dense fiber 2 (Odf2) was initially identified as a major component of sperm tail cytoskeleton and later was suggested to be a widespread component of centrosomal scaffold that preferentially associates with the appendages of the mother centrioles in somatic cells. Here we report the identification of two Odf2-related centrosomal components, hCenexin1 and hCenexin1 variant 1, that possess a unique C-terminal extension. Our results showed that hCenexin1 is the major isoform expressed in HeLa cells, whereas hOdf2 is not detectably expressed. Mammalian polo-like kinase 1 (Plk1) is critical for proper mitotic progression, and its association with the centrosome is important for microtubule nucleation and function. Interestingly, depletion of hCenexin1 by RNA interference (RNAi) delocalized Plk1 from the centrosomes and the C-terminal extension of hCenexin1 was crucial to recruit Plk1 to the centrosomes through a direct interaction with the polo-box domain of Plk1. Consistent with these findings, the hCenexin1 RNAi cells exhibited weakened gamma-tubulin localization and chromosome segregation defects. We propose that hCenexin1 is a critical centrosomal component whose C-terminal extension is required for proper recruitment of Plk1 and other components crucial for normal mitosis. Our results further suggest that the anti-Odf2 immunoreactive centrosomal antigen previously detected in non-germ line cells is likely hCenexin1.     

Soy isoflavones prevent the ovarian hormone deficiency-associated rise in leukocytes in rats.

Recent reports indicate that ovariectomy (ovx) increases lymphopoiesis. Ipriflavone, a synthetic isoflavone, has been reported to reduce lymphocytes in postmenopausal women. The aim of this study was to investigate whether naturally occurring isoflavones also affect lymphopoiesis in ovarian hormone deficiency. The present study was carried out using an ovariectomized (ovx) rat model. To mimic early menopause, forty-eight 12-month-old Sprague-Dawley rats were either sham-operated (sham; 1 group) or ovx (3 groups) and were fed a standard semi-purified diet for 120 days. Thereafter, the ovx groups received one of the three doses of isoflavones: 0 (ovx), 500 (ISO500), or 1000 (ISO1000) mg/kg diet for 100 days. Ovariectomy increased total leukocyte counts significantly (p < 0.05) as a result of increased (p < 0.05) lymphocyte, monocyte, eosinophil, and basophil differential counts. Isoflavones at 500 and 1000 mg/kg diet returned the total leukocyte counts, as well as leukocyte subpopulations, to levels comparable to that of sham-operated rats. No other hematological parameters, e.g., red blood cell counts or red cell indices, were affected by ovariectomy or isoflavones. We conclude that soy isoflavones restore normal leukocyte counts elevated in ovarian hormone deficiency.