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D Clegg

Publications and source records attributed to D Clegg.

7 recordsLinked to original sources
PubMed2001

Inward rectifier potassium channel Kir2.2 is associated with synapse-associated protein SAP97.

The strong inwardly rectifying potassium channels Kir2.x are involved in maintenance and control of cell excitability. Recent studies reveal that the function and localization of ion channels are regulated by interactions with members of the membrane-associated guanylate kinase (MAGUK) protein family. To identify novel interacting MAGUK family members, we constructed GST-fusion proteins with the C termini of Kir2.1, Kir2.2 and Kir2.3. GST affinity-pulldown assays from solubilized rat cerebellum and heart membrane proteins revealed an interaction between all three Kir2.x C-terminal fusion proteins and the MAGUK protein synapse-associated protein 97 (SAP97). A truncated form of the C-terminal GST-Kir2.2 fusion protein indicated that the last three amino acids (S-E-I) are essential for association with SAP97. Affinity interactions using GST-fusion proteins containing the modular domains of SAP97 demonstrate that the second PSD-95/Dlg/ZO-1 (PDZ) domain is sufficient for interaction with Kir2.2. Coimmunoprecipitations demonstrated that endogenous Kir2.2 associates with SAP97 in rat cerebellum and heart. Additionally, phosphorylation of the Kir2.2 C terminus by protein kinase A inhibited the association with SAP97. In rat cardiac ventricular myocytes, Kir2.2 and SAP97 colocalized in striated bands corresponding to T-tubules. In rat cerebellum, Kir2.2 was present in a punctate pattern along SAP97-positive processes of Bergmann glia in the molecular layer, and colocalized with astrocytes and granule cells in the granule cell layer. These results identify a direct association of Kir2.1, Kir2.2 and Kir2.3 with the MAGUK family member SAP97 that may form part of a macromolecular signaling complex in many different tissues.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Developmental changes in nicotinic receptor mRNAs and responses to nicotine in the suprachiasmatic nucleus and other brain regions.

Our previous studies demonstrated that nicotine induces c-fos expression in the suprachiasmatic nucleus (SCN) of the rat during a narrow developmental window occurring in the perinatal period. We have extended these observations by showing that c-fos cannot be induced in the adult SCN by nicotine even during the subjective night, when phase shifts do occur. In contrast to the SCN, significant induction of c-fos and NGFI-A was observed in the medial habenula and paraventricular nucleus at all circadian times. In the fetal rat SCN we show that NGFI-A and junB are also induced by nicotine, but not c-jun. To investigate whether changes in nicotinic acetylcholine receptor (nAChR) expression in the SCN may underlie this change in sensitivity during the perinatal period, we examined nAChR mRNAs across this developmental period. By Northern analyses, alpha2, alpha3 and alpha4 subunit mRNAs are relatively abundant in the fetal SCN but decline substantially in the adult. alpha7 mRNA increases substantially while beta2 mRNA is relatively abundant throughout development. We also examine expression in the whole mouse brain beginning at embryonic day 11. Many mRNA sizes for nAChR subunits in both the rat and mouse are characterized here for the first time by Northern analyses and some show very large changes in expression across development. In particular, a small 1.4 kb alpha2-related mRNA is highly expressed during early development, perhaps indicating an important novel function for this subunit.

Acetylcholine↗
PubMed1998

The alpha4 subunit of integrin is important for neural crest cell migration.

We identify the alpha4 subunit of integrin as a predominant integrin expressed by neural crest cells in both avian and murine embryos. Using degenerate primers, we obtained a PCR fragment of the chick integrin alpha4 subunit that was subsequently used to clone the full-length subunit with a predicted amino acid sequence 60% identical to human and mouse alpha4 subunits. In situ hybridization demonstrates that chick integrin alpha4 mRNA is expressed at high levels by migrating neural crest cells and neural crest-derived ganglia at both cranial and trunk levels. An antibody against the murine alpha4 subunit revealed similar distribution patterns in mouse to chick. In addition to neural crest cells, the integrin alpha4 subunit was later observed on the muscle masses of the limb, the apical ectodermal ridge, and the developing liver. To examine the functional role of the integrin alpha4 subunit in neural crest cell migration, we used an explant preparation that allows visualization of neural crest cells in their normal environment with or without perturbing reagents. In the presence of a blocking antibody against the mouse integrin alpha4 subunit, there was a profound abrogation of neural crest cell migration at trunk and hindbrain levels. Both the numbers of migrating neural crest cells and the total distance traversed were markedly reduced. Similarly, avian embryos injected with synthetic peptides that contain the integrin alpha4 binding site in fibronectin displayed abnormal neural crest cell migration. Our results suggest that the integrin alpha4 subunit is important for normal neural crest cell migration and may be one of the primary alpha subunits used for neural crest cell migration in vivo. Furthermore, the integrin alpha4 subunit represents a useful neural crest marker in the mouse.

Amino Acid Sequence↗
PubMed1986

Toxicity of toxaphene in the rat and beagle dog.

Residues of the insecticidal mixture, toxaphene, have been found in Great Lakes fish. The purpose of the present study was to assess the subchronic toxicity of toxaphene in the rat and beagle dog. In the rat study, groups of 10 male and 10 female animals were fed diets containing 0, 4, 20, 100, or 500 ppm of the test compound for 13 weeks. No clinical signs of toxicity or spontaneous deaths were observed. Toxaphene treatment up to 500 ppm had no effects on weight gain or food consumption. The liver/body weight ratio and hepatic microsomal enzyme activities (phenobarbital type) were increased in both sexes fed 500 ppm of the test compound. Toxaphene at the highest dose also caused kidney enlargement in male but not in female rats. Dose-dependent histological changes were seen in the kidney, thyroid, and liver. Changes in the liver and thyroid were considered to be adaptative but the injury in the proximal tubules of the kidney was focally severe. Groups of six male and six female beagle dogs were fed toxaphene in gelatin capsules at 0, 0.2, 2.0, and 5.0 mg/kg body wt/kg body wt/day for 13 weeks. Food consumption and growth rate were not affected. All animals survived the entire treatment period. No clinical signs of toxicity were observed. The liver/body weight ratio and serum alkaline phosphatase were increased in dogs of both sexes fed 5.0 mg/kg. Mild to moderate dose-dependent histological changes were observed in the liver and thyroid. Toxaphene was accumulated in a dose-dependent manner in the fat and liver of dogs and rats. Based on the biochemical, histological, and residue data, it was concluded that the no-adverse-effect levels of the pesticide were 4.0 ppm (0.35 mg/kg) for the rat and 0.2 mg/kg for the dog.

Animals↗